Biochemistry

An introduction

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Contents
Articles
Cells and water
Biochemistry Cells Water 1 1 10 21 43 44 44 47 55 79 79 93 108 118 124 141 141 148 148 163 163 174 175 179 186 186 193

Structural Biochemistry Nucleic acids

Nucleic acid RNA DNA

Proteins and amino acids

Protein Amino acid Properties of the twenty amino acids Myoglobin Hemoglobin

Enzyme mechanisms
Enzyme catalysis

Enzyme kinetics
Enzyme kinetics

Lipids and membranes

Lipid Biological membrane Membrane protein Cell membrane

Carbohydrate structure
Carbohydrate Polysaccharide

Intermediary metabolism Metabolism

Overview of metabolism

200 201 201 221 221 234 237 241 245 245 253 253 269 269 283 283 290 292 299 301 305 305 310 313 318 322 322 326 335 349

Carbohydrate metabolism
Glycolysis Gluconeogenesis Glycogen Pentose phosphate pathway

Citric acid cycle

Citric acid cycle

Oxidative phosphorylation
Oxidative phosphorylation

Photosynthesis
Photosynthesis

Lipid metabolism
Fatty acid synthesis Lipogenesis Acetyl-CoA carboxylase Fatty acid degradation Beta oxidation

Nitrogen metabolism
Nitrogen fixation Amino acid synthesis Nucleotide Urea cycle

Integration of metabolism
Hormone Signal transduction Diabetes mellitus

Informational Macromolecules

DNA synthesis and repair

DNA replication DNA repair Oncogenes

350 350 358 370 374 374 380 385 385 389 393 395

RNA synthesis and processing

Transcription Regulation of gene expression

Protein synthesis and modifications

Translation Posttranslational modification Proteolysis Proteasome

References
Article Sources and Contributors Image Sources, Licenses and Contributors 409 420

Article Licenses
License 427

Cells and water

Biochemistry
Biochemistry, sometimes called biological chemistry, is the study of chemical processes in living organisms, including, but not limited to, living matter. Biochemistry governs all living organisms and living processes. By controlling information flow through biochemical signalling and the flow of chemical energy through metabolism, biochemical processes give rise to the incredible complexity of life. Much of biochemistry deals with the structures and functions of cellular components such as proteins, carbohydrates, lipids, nucleic acids and other biomolecules although increasingly processes rather than individual molecules are the main focus. Over the last 40 years biochemistry has become so successful at explaining living processes that now almost all areas of the life sciences from botany to medicine are engaged in biochemical research. Today the main focus of pure biochemistry is in understanding how biological molecules give rise to the processes that occur within living cells, which in turn relates greatly to the study and understanding of whole organisms. Among the vast number of different biomolecules, many are complex and large molecules (called biopolymers), which are composed of similar repeating subunits (called monomers). Each class of polymeric biomolecule has a different set of subunit types.[1] For example, a protein is a polymer whose subunits are selected from a set of 20 or more amino acids. Biochemistry studies the chemical properties of important biological molecules, like proteins, and in particular the chemistry of enzyme-catalyzed reactions. The biochemistry of cell metabolism and the endocrine system has been extensively described. Other areas of biochemistry include the genetic code (DNA, RNA), protein synthesis, cell membrane transport, and signal transduction.

History
It once was generally believed that life and its materials had some essential property or substance distinct from any found in non-living matter, and it was thought that only living beings could produce the molecules of life. Then, in 1828, Friedrich Whler published a paper on the synthesis of urea, proving that organic compounds can be created artificially.[2] [3] The dawn of biochemistry may have been the discovery of the first enzyme, diastase (today called amylase), in 1833 by Anselme Payen. Eduard Buchner contributed the first demonstration of a complex biochemical process outside of a cell in 1896: alcoholic fermentation in cell extracts of yeast. Although the term biochemistry seems to have been first used in 1882, it is generally accepted that the formal coinage of biochemistry occurred in 1903 by Carl Neuberg, a German chemist. Previous to this time, this area would have been referred to as physiological chemistry. Since then, biochemistry has advanced, especially since the mid-20th century, with the development of new techniques such as chromatography, X-ray diffraction, dual polarisation interferometry, NMR spectroscopy, radioisotopic labeling, electron microscopy, and molecular dynamics simulations. These techniques allowed for the discovery and detailed analysis of many molecules and metabolic pathways of the cell, such as glycolysis and the Krebs cycle (citric acid cycle). Another significant historic event in biochemistry is the discovery of the gene and its role in the transfer of information in the cell. This part of biochemistry is often called molecular biology. In the 1950s, James D. Watson, Francis Crick, Rosalind Franklin, and Maurice Wilkins were instrumental in solving DNA structure and suggesting its relationship with genetic transfer of information. In 1958, George Beadle and Edward Tatum received the Nobel Prize for work in fungi showing that one gene produces one enzyme. In 1988, Colin Pitchfork was the first person

Biochemistry convicted of murder with DNA evidence, which led to growth of forensic science. More recently, Andrew Z. Fire and Craig C. Mello received the 2006 Nobel Prize for discovering the role of RNA interference (RNAi), in the silencing of gene expression.

Starting materials: the chemical elements of life

Around two dozen of the 94 naturally-occurring chemical elements are essential to various kinds of biological life. Most rare elements on Earth are not needed by life (exceptions being selenium and iodine), while a few common ones (aluminum and titanium) are not used. Most organisms share element needs, but there are a few differences between plants and animals. For example ocean algae use bromine but land plants and animals seem to need none. All animals require sodium, but some plants do not. Plants need boron and silicon, but animals may not (or may need ultra-small amounts). Just six elementscarbon, hydrogen, nitrogen, oxygen, calcium, and phosphorusmake up almost 99% of the mass of a human body (see composition of the human body for a complete list). In addition to the six major elements that compose most of the human body, humans require smaller amounts of possibly 18 more.[4]

Biomolecules
The four main classes of molecules in biochemistry are carbohydrates, lipids, proteins, and nucleic acids. Many biological molecules are polymers: in this terminology, monomers are relatively small micromolecules that are linked together to create large macromolecules, which are known as polymers. When monomers are linked together to synthesize a biological polymer, they undergo a process called dehydration synthesis.

Carbohydrates
Carbohydrates are made from monomers called monosaccharides. Some of these monosaccharides include glucose (C6H12O6), fructose (C6H12O6), and deoxyribose (C5H10O4). When two monosaccharides undergo dehydration synthesis, water is produced, as two hydrogen atoms and one oxygen atom are lost from the two monosaccharides' hydroxyl group.

A molecule of sucrose (glucose + fructose), a disaccharide.

Lipids
Lipids are usually made from one molecule of glycerol combined with other molecules. In triglycerides, the main group of bulk lipids, there is one molecule of glycerol and three fatty acids. Fatty acids are considered the monomer in that case, and may be saturated (no double bonds in the carbon chain) or unsaturated (one or more double bonds in the carbon chain). Lipids, especially phospholipids, are also used in various pharmaceutical products, either as co-solubilisers (e.g., in parenteral infusions) or else as drug carrier components (e.g., in a liposome or transfersome).

A triglyceride with a glycerol molecule on the left and three fatty acids coming off it.

Biochemistry

Proteins
Proteins are very large molecules macro-biopolymers made from monomers called amino acids. There are 20 standard amino acids, each containing a carboxyl group, an amino group, and a side-chain (known as an "R" group). The "R" group is what makes each amino acid different, and the properties of the side-chains greatly influence the overall three-dimensional conformation of a protein. When amino acids combine, they form a special bond called a peptide bond through dehydration synthesis, and become a polypeptide, or protein. In order to determine whether two proteins are related, or in other words to decide whether they are homologous or not, scientists use sequence-comparison methods. Methods like Sequence Alignments and Structural Alignments are powerful tools that help scientists identify homologies between related molecules.

The general structure of an -amino acid, with the amino group on the left and the carboxyl group on the right.

The relevance of finding homologies among proteins goes beyond forming an evolutionary pattern of protein families. By finding how similar two protein sequences are, we acquire knowledge about their structure and therefore their function.

Nucleic acids
Nucleic acids are the molecules that make up DNA, an extremely important substance that all cellular organisms use to store their genetic information. The most common nucleic acids are deoxyribonucleic acid and ribonucleic acid. Their monomers are called nucleotides. The most common nucleotides are adenine, cytosine, guanine, thymine, and uracil. Adenine binds with thymine and uracil; Thymine binds only with adenine; and cytosine and guanine can bind only with each other.

Carbohydrates
The function of carbohydrates includes energy storage and providing structure. Sugars are carbohydrates, but not all carbohydrates are sugars. There are more carbohydrates on Earth than any other known type of biomolecule; they are used to store energy and genetic information, as well as play important roles in cell to cell interactions and communications.

The structure of deoxyribonucleic acid (DNA), the picture shows the monomers being put together.

Monosaccharides
The simplest type of carbohydrate is a monosaccharide, which among other properties contains carbon, hydrogen, and oxygen, mostly in a ratio of 1:2:1 (generalized formula CnH2nOn, where n is at least 3). Glucose, one of the most important carbohydrates, is an example of a monosaccharide. So is fructose, the sugar commonly associated with the sweet taste of fruits.[5] [a] Some carbohydrates (especially after
Glucose

Biochemistry condensation to oligo- and polysaccharides) contain less carbon relative to H and O, which still are present in 2:1 (H:O) ratio. Monosaccharides can be grouped into aldoses (having an aldehyde group at the end of the chain, e.g. glucose) and ketoses (having a keto group in their chain; e.g. fructose). Both aldoses and ketoses occur in an equilibrium (starting with chain lengths of C4) cyclic forms. These are generated by bond formation between one of the hydroxyl groups of the sugar chain with the carbon of the aldehyde or keto group to form a hemiacetal bond. This leads to saturated five-membered (in furanoses) or six-membered (in pyranoses) heterocyclic rings containing one O as heteroatom.

Disaccharides
Two monosaccharides can be joined together using dehydration synthesis, in which a hydrogen atom is removed from the end of one molecule and a hydroxyl group (OH) is removed from the other; the remaining residues are then attached at the sites from which the atoms were removed. The HOH or H2O is then released as a molecule of water, hence the term dehydration. The new molecule, consisting of Sucrose: ordinary table sugar and probably the two monosaccharides, is called a disaccharide and is conjoined most familiar carbohydrate. together by a glycosidic or ether bond. The reverse reaction can also occur, using a molecule of water to split up a disaccharide and break the glycosidic bond; this is termed hydrolysis. The most well-known disaccharide is sucrose, ordinary sugar (in scientific contexts, called table sugar or cane sugar to differentiate it from other sugars). Sucrose consists of a glucose molecule and a fructose molecule joined together. Another important disaccharide is lactose, consisting of a glucose molecule and a galactose molecule. As most humans age, the production of lactase, the enzyme that hydrolyzes lactose back into glucose and galactose, typically decreases. This results in lactase deficiency, also called lactose intolerance. Sugar polymers are characterised by having reducing or non-reducing ends. A reducing end of a carbohydrate is a carbon atom that can be in equilibrium with the open-chain aldehyde or keto form. If the joining of monomers takes place at such a carbon atom, the free hydroxy group of the pyranose or furanose form is exchanged with an OH-side-chain of another sugar, yielding a full acetal. This prevents opening of the chain to the aldehyde or keto form and renders the modified residue non-reducing. Lactose contains a reducing end at its glucose moiety, whereas the galactose moiety form a full acetal with the C4-OH group of glucose. Saccharose does not have a reducing end because of full acetal formation between the aldehyde carbon of glucose (C1) and the keto carbon of fructose (C2).

Oligosaccharides and polysaccharides

When a few (around three to six) monosaccharides are joined together, it is called an oligosaccharide (oligo- meaning "few"). These molecules tend to be used as markers and signals, as well as having some other uses. Many monosaccharides joined together make a polysaccharide. They can be joined together in one long linear chain, or they may be branched. Two of the most common polysaccharides are cellulose and glycogen, both consisting of repeating glucose monomers.

Cellulose as polymer of -D-glucose

Cellulose is made by plants and is an important structural component of their cell walls. Humans can neither manufacture nor digest it. Glycogen, on the other hand, is an animal carbohydrate; humans and other animals use it as a form of energy storage.

Biochemistry

Use of carbohydrates as an energy source

Glucose is the major energy source in most life forms. For instance, polysaccharides are broken down into their monomers (glycogen phosphorylase removes glucose residues from glycogen). Disaccharides like lactose or sucrose are cleaved into their two component monosaccharides. Glycolysis (anaerobic) Glucose is mainly metabolized by a very important ten-step pathway called glycolysis, the net result of which is to break down one molecule of glucose into two molecules of pyruvate; this also produces a net two molecules of ATP, the energy currency of cells, along with two reducing equivalents in the form of converting NAD+ to NADH. This does not require oxygen; if no oxygen is available (or the cell cannot use oxygen), the NAD is restored by converting the pyruvate to lactate (lactic acid) (e.g., in humans) or to ethanol plus carbon dioxide (e.g., in yeast). Other monosaccharides like galactose and fructose can be converted into intermediates of the glycolytic pathway. Aerobic In aerobic cells with sufficient oxygen, as in most human cells, the pyruvate is further metabolized. It is irreversibly converted to acetyl-CoA, giving off one carbon atom as the waste product carbon dioxide, generating another reducing equivalent as NADH. The two molecules acetyl-CoA (from one molecule of glucose) then enter the citric acid cycle, producing two more molecules of ATP, six more NADH molecules and two reduced (ubi)quinones (via FADH2 as enzyme-bound cofactor), and releasing the remaining carbon atoms as carbon dioxide. The produced NADH and quinol molecules then feed into the enzyme complexes of the respiratory chain, an electron transport system transferring the electrons ultimately to oxygen and conserving the released energy in the form of a proton gradient over a membrane (inner mitochondrial membrane in eukaryotes). Thus, oxygen is reduced to water and the original electron acceptors NAD+ and quinone are regenerated. This is why humans breathe in oxygen and breathe out carbon dioxide. The energy released from transferring the electrons from high-energy states in NADH and quinol is conserved first as proton gradient and converted to ATP via ATP synthase. This generates an additional 28 molecules of ATP (24 from the 8 NADH + 4 from the 2 quinols), totaling to 32 molecules of ATP conserved per degraded glucose (two from glycolysis + two from the citrate cycle). It is clear that using oxygen to completely oxidize glucose provides an organism with far more energy than any oxygen-independent metabolic feature, and this is thought to be the reason why complex life appeared only after Earth's atmosphere accumulated large amounts of oxygen. Gluconeogenesis In vertebrates, vigorously contracting skeletal muscles (during weightlifting or sprinting, for example) do not receive enough oxygen to meet the energy demand, and so they shift to anaerobic metabolism, converting glucose to lactate. The liver regenerates the glucose, using a process called gluconeogenesis. This process is not quite the opposite of glycolysis, and actually requires three times the amount of energy gained from glycolysis (six molecules of ATP are used, compared to the two gained in glycolysis). Analogous to the above reactions, the glucose produced can then undergo glycolysis in tissues that need energy, be stored as glycogen (or starch in plants), or be converted to other monosaccharides or joined into di- or oligosaccharides. The combined pathways of glycolysis during exercise, lactate's crossing via the bloodstream to the liver, subsequent gluconeogenesis and release of glucose into the bloodstream is called the Cori cycle.

Biochemistry

Proteins
Like carbohydrates, some proteins perform largely structural roles. For instance, movements of the proteins actin and myosin ultimately are responsible for the contraction of skeletal muscle. One property many proteins have is that they specifically bind to a certain molecule or class of moleculesthey may be extremely selective in what they bind. Antibodies are an example of proteins that attach to one specific type of molecule. In fact, the enzyme-linked immunosorbent assay (ELISA), which uses antibodies, is currently one of the most sensitive tests modern medicine uses to detect various biomolecules. Probably the most important proteins, however, are the enzymes. These molecules recognize specific reactant A schematic of hemoglobin. The red and molecules called substrates; they then catalyze the reaction between them. By blue ribbons represent the protein globin; lowering the activation energy, the enzyme speeds up that reaction by a rate the green structures are the heme groups. of 1011 or more: a reaction that would normally take over 3,000 years to complete spontaneously might take less than a second with an enzyme. The enzyme itself is not used up in the process, and is free to catalyze the same reaction with a new set of substrates. Using various modifiers, the activity of the enzyme can be regulated, enabling control of the biochemistry of the cell as a whole. In essence, proteins are chains of amino acids. An amino acid consists of a carbon atom bound to four groups. One is an amino group, NH2, and one is a carboxylic acid group, COOH (although these exist as NH3+ and COO under physiologic conditions). The third is a simple hydrogen atom. The fourth is commonly denoted "R" and is different for each amino acid. There are twenty standard amino acids. Some of these have functions by themselves or in a modified form; for instance, glutamate functions as an important neurotransmitter. Amino acids can be joined together via a peptide bond. In this dehydration synthesis, a water molecule is removed and the peptide bond connects the nitrogen of one amino acid's amino group to the carbon of the other's Generic amino acids (1) in neutral form, (2) as they exist physiologically, and (3) joined carboxylic acid group. The resulting together as a dipeptide. molecule is called a dipeptide, and short stretches of amino acids (usually, fewer than thirty) are called peptides or polypeptides. Longer stretches merit the title proteins. As an example, the important blood serum protein albumin contains 585 amino acid residues. The structure of proteins is traditionally described in a hierarchy of four levels. The primary structure of a protein simply consists of its linear sequence of amino acids; for instance, "alanine-glycine-tryptophan-serine-glutamate-asparagine-glycine-lysine-". Secondary structure is concerned with local morphology (morphology being the study of structure). Some combinations of amino acids will tend to curl up in a coil called an -helix or into a sheet called a -sheet; some -helixes can be seen in the hemoglobin schematic above. Tertiary structure is the entire three-dimensional shape of the protein. This shape is determined by the sequence of amino acids. In fact, a single change can change the entire structure. The alpha chain of hemoglobin contains 146 amino acid residues; substitution of the glutamate residue at position 6 with a valine residue changes the behavior of hemoglobin so much that it results in sickle-cell disease. Finally, quaternary structure is concerned with the structure of a protein with multiple peptide subunits, like hemoglobin with its four subunits. Not all proteins have more than one subunit.

Biochemistry Ingested proteins are usually broken up into single amino acids or dipeptides in the small intestine, and then absorbed. They can then be joined together to make new proteins. Intermediate products of glycolysis, the citric acid cycle, and the pentose phosphate pathway can be used to make all twenty amino acids, and most bacteria and plants possess all the necessary enzymes to synthesize them. Humans and other mammals, however, can synthesize only half of them. They cannot synthesize isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine. These are the essential amino acids, since it is essential to ingest them. Mammals do possess the enzymes to synthesize alanine, asparagine, aspartate, cysteine, glutamate, glutamine, glycine, proline, serine, and tyrosine, the nonessential amino acids. While they can synthesize arginine and histidine, they cannot produce it in sufficient amounts for young, growing animals, and so these are often considered essential amino acids. If the amino group is removed from an amino acid, it leaves behind a carbon skeleton called an -keto acid. Enzymes called transaminases can easily transfer the amino group from one amino acid (making it an -keto acid) to another -keto acid (making it an amino acid). This is important in the biosynthesis of amino acids, as for many of the pathways, intermediates from other biochemical pathways are converted to the -keto acid skeleton, and then an amino group is added, often via transamination. The amino acids may then be linked together to make a protein. A similar process is used to break down proteins. It is first hydrolyzed into its component amino acids. Free ammonia (NH3), existing as the ammonium ion (NH4+) in blood, is toxic to life forms. A suitable method for excreting it must therefore exist. Different strategies have evolved in different animals, depending on the animals' needs. Unicellular organisms, of course, simply release the ammonia into the environment. Likewise, bony fish can release the ammonia into the water where it is quickly diluted. In general, mammals convert the ammonia into urea, via the urea cycle.

Lipids
The term lipid comprises a diverse range of molecules and to some extent is a catchall for relatively water-insoluble or nonpolar compounds of biological origin, including waxes, fatty acids, fatty-acid derived phospholipids, sphingolipids, glycolipids, and terpenoids (e.g., retinoids and steroids). Some lipids are linear aliphatic molecules, while others have ring structures. Some are aromatic, while others are not. Some are flexible, while others are rigid. Most lipids have some polar character in addition to being largely nonpolar. In general, the bulk of their structure is nonpolar or hydrophobic ("water-fearing"), meaning that it does not interact well with polar solvents like water. Another part of their structure is polar or hydrophilic ("water-loving") and will tend to associate with polar solvents like water. This makes them amphiphilic molecules (having both hydrophobic and hydrophilic portions). In the case of cholesterol, the polar group is a mere -OH (hydroxyl or alcohol). In the case of phospholipids, the polar groups are considerably larger and more polar, as described below. Lipids are an integral part of our daily diet. Most oils and milk products that we use for cooking and eating like butter, cheese, ghee etc., are composed of fats. Vegetable oils are rich in various polyunsaturated fatty acids (PUFA). Lipid-containing foods undergo digestion within the body and are broken into fatty acids and glycerol, which are the final degradation products of fats and lipids.

Nucleic acids
A nucleic acid is a complex, high-molecular-weight biochemical macromolecule composed of nucleotide chains that convey genetic information. The most common nucleic acids are deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). Nucleic acids are found in all living cells and viruses. Aside from the genetic material of the cell, nucleic acids often play a role as second messengers, as well as forming the base molecule for adenosine triphosphate, the primary energy-carrier molecule found in all living organisms. Nucleic acid, so called because of its prevalence in cellular nuclei, is the generic name of the family of biopolymers. The monomers are called nucleotides, and each consists of three components: a nitrogenous heterocyclic base (either

Biochemistry a purine or a pyrimidine), a pentose sugar, and a phosphate group. Different nucleic acid types differ in the specific sugar found in their chain (e.g., DNA or deoxyribonucleic acid contains 2-deoxyriboses). Also, the nitrogenous bases possible in the two nucleic acids are different: adenine, cytosine, and guanine occur in both RNA and DNA, while thymine occurs only in DNA and uracil occurs in RNA.

Relationship to other "molecular-scale" biological sciences

Researchers in biochemistry use specific techniques native to biochemistry, but increasingly combine these with techniques and ideas from genetics, molecular biology and biophysics. There has never been a hard-line between these disciplines in terms of content and technique. Today, the terms molecular biology and biochemistry are nearly interchangeable. The following figure is a schematic that depicts one possible view of the relationship between the fields: Biochemistry is the study of the chemical substances and vital processes occurring in living organisms. Biochemists focus heavily on the role, function, and structure of biomolecules. The study of the chemistry behind biological processes and the synthesis of biologically active molecules are examples of biochemistry.

Schematic relationship between biochemistry, genetics, and molecular biology

Genetics is the study of the effect of genetic differences on organisms. Often this can be inferred by the absence of a normal component (e.g., one gene). The study of "mutants" organisms with a changed gene that leads to the organism being different with respect to the so-called "wild type" or normal phenotype. Genetic interactions (epistasis) can often confound simple interpretations of such "knock-out" or "knock-in" studies. Molecular biology is the study of molecular underpinnings of the process of replication, transcription and translation of the genetic material. The central dogma of molecular biology where genetic material is transcribed into RNA and then translated into protein, despite being an oversimplified picture of molecular biology, still provides a good starting point for understanding the field. This picture, however, is undergoing revision in light of emerging novel roles for RNA. Chemical Biology seeks to develop new tools based on small molecules that allow minimal perturbation of biological systems while providing detailed information about their function. Further, chemical biology employs biological systems to create non-natural hybrids between biomolecules and synthetic devices (for example emptied viral capsids that can deliver gene therapy or drug molecules).

Biochemistry

Notes
a. It should be noted that fructose is not the only sugar found in fruits. Glucose and sucrose are also found in varying quantities in various fruits, and indeed sometimes exceed the fructose present. For example, 32% of the edible portion of date is glucose, compared with 23.70% fructose and 8.20% sucrose. However, peaches contain more sucrose (6.66%) than they do fructose (0.93%) or glucose (1.47%).[6]

References
[1] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [2] Whler, F. (1828). "Ueber knstliche Bildung des Harnstoffs". Ann. Phys. Chem. 12: 253256. [3] Kauffman, G. B. and Chooljian, S.H. (2001). "Friedrich Whler (18001882), on the Bicentennial of His Birth". The Chemical Educator 6 (2): 121133. doi:10.1007/s00897010444a. [4] Ultratrace minerals. Authors: Nielsen, Forrest H. USDA, ARS Source: Modern nutrition in health and disease / editors, Maurice E. Shils ... et al.. Baltimore : Williams & Wilkins, c1999., p. 283-303. Issue Date: 1999 URI: (http:/ / hdl. handle. net/ 10113/ 46493) [5] Whiting, G.C (1970). "Sugars". In A.C. Hulme. The Biochemistry of Fruits and their Products. Volume 1. London & New York: Academic Press. pp.1=31 [6] Whiting, G.C. (1970), p.5

Further reading
Hunter, Graeme K. (2000). Vital Forces: The Discovery of the Molecular Basis of Life. San Diego: Academic Press. ISBN0-12-361810-X. OCLC162129355 191848148 44187710.

External links
The Virtual Library of Biochemistry and Cell Biology (http://www.biochemweb.org/) Biochemistry, 5th ed. (http://www.ncbi.nlm.nih.gov/books/bv.fcgi?call=bv.View..ShowTOC&rid=stryer. TOC&depth=2) Full text of Berg, Tymoczko, and Stryer, courtesy of NCBI. Biochemistry, 2nd ed. (http://www.web.virginia.edu/Heidi/home.htm) Full text of Garrett and Grisham. Biochemistry Animation (http://www.1lec.com/Biochemistry/) (Narrated Flash animations.) SystemsX.ch - The Swiss Initiative in Systems Biology (http://www.systemsX.ch/) Biochemistry Online Resources (http://www.icademic.org/97445/Biochemistry/) Lists of Biochemistry departments, websites, journals, books and reviews, employment opportunities and events. biochemical families: prot nucl carb (glpr, alco, glys) lipd (fata/i, phld, strd, gllp, eico) amac/i ncbs/i ttpy/i

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Cells
The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life.[1] Organisms can be classified as unicellular (consisting of a single cell; including most bacteria) or multicellular (including plants and animals). Humans contain about 100 trillion cells; a typical cell size is 10m and a typical cell mass is 1nanogram. The human cell extrema are: largest - anterior horn in the spinal cord (135 vs. 120 for the ovum), longest - pseudounipolar cells which reach from extremities, including the toes to the lower brain stem, and smallest - granule cells in the cerebellum, at 4.[2] The largest known cells are unfertilised ostrich egg cells, which weigh 3.3 pounds.[3] [4]

Allium cells in different phases of the cell cycle

The cell was discovered by Robert Hooke in 1665. In 1835, before the final cell theory was developed, Jan Evangelista Purkyn observed small "granules" while looking at the plant tissue through a microscope. The cell Cells in culture, stained for keratin (red) and theory, first developed in 1839 by DNA (green) Matthias Jakob Schleiden and Theodor Schwann, states that all organisms are composed of one or more cells, that all cells come from preexisting cells, that vital functions of an organism occur within cells, and that all cells contain the hereditary information necessary for regulating cell functions and for transmitting information to the next generation of cells.[5] The word cell comes from the Latin cellula, meaning "a small room". The

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descriptive term for the smallest living biological structure was coined by Robert Hooke in a book he published in 1665 when he compared the cork cells he saw through his microscope to the small rooms monks lived in.[6]

Drawing of the structure of cork as it appeared under the microscope to Robert Hooke from Micrographia, which is the origin of the word "cell" being used to describe the smallest unit of a living organism

Anatomy
There are two types of cells: eukaryotic and prokaryotic. Prokaryotic cells are usually independent, while eukaryotic cells are often found in multicellular organisms.

The cells of eukaryotes (left) and prokaryotes (right)

Table 1: Comparison of features of prokaryotic and eukaryotic cells

Prokaryotes Typical organisms Typical size Type of nucleus DNA RNA-/protein-synthesis bacteria, archaea ~ 110 m protists, fungi, plants, animals ~ 10100 m (sperm cells, apart from the tail, are smaller) Eukaryotes

nucleoid region; no real nucleus real nucleus with double membrane circular (usually) coupled in cytoplasm linear molecules (chromosomes) with histone proteins RNA-synthesis inside the nucleus protein synthesis in cytoplasm 60S+40S

Ribosomes

50S+30S

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very few structures flagella made of flagellin none none usually single cells highly structured by endomembranes and a cytoskeleton flagella and cilia containing microtubules; lamellipodia and filopodia containing actin one to several thousand (though some lack mitochondria) in algae and plants single cells, colonies, higher multicellular organisms with specialized cells

Cytoplasmatic structure Cell movement Mitochondria Chloroplasts Organization Cell division

Binary fission (simple division) Mitosis (fission or budding) Meiosis

Prokaryotic cells
The prokaryote cell is simpler, and therefore smaller, than a eukaryote cell, lacking a nucleus and most of the other organelles of eukaryotes. There are two kinds of prokaryotes: bacteria and archaea; these share a similar structure. Nuclear material of prokaryotic cell consist of a single chromosome that is in direct contact with cytoplasm. Here, the undefined nuclear region in the cytoplasm is called nucleoid. A prokaryotic cell architectural regions: has three

On the outside, flagella and pili project from the cell's surface. These are structures (not present in all prokaryotes) made of proteins that facilitate movement and communication between cells;

Diagram of a typical prokaryotic cell

Enclosing the cell is the cell envelope generally consisting of a cell wall covering a plasma membrane though some bacteria also have a further covering layer called a capsule. The envelope gives rigidity to the cell and separates the interior of the cell from its environment, serving as a protective filter. Though most prokaryotes have a cell wall, there are exceptions such as Mycoplasma (bacteria) and Thermoplasma (archaea). The cell wall consists of peptidoglycan in bacteria, and acts as an additional barrier against exterior forces. It also prevents the cell from expanding and finally bursting (cytolysis) from osmotic pressure against a hypotonic environment. Some eukaryote cells (plant cells and fungi cells) also have a cell wall; Inside the cell is the cytoplasmic region that contains the cell genome (DNA) and ribosomes and various sorts of inclusions. A prokaryotic chromosome is usually a circular molecule (an exception is that of the bacterium Borrelia burgdorferi, which causes Lyme disease). Though not forming a nucleus, the DNA is condensed in a nucleoid. Prokaryotes can carry extrachromosomal DNA elements called plasmids, which are usually circular. Plasmids enable additional functions, such as antibiotic resistance.

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Eukaryotic cells
Plants, animals, fungi, slime moulds, protozoa, & algae are all Eukaryotic. These cells are about 15 times wider than a typical prokaryote and can be as much as 1000 times greater in volume. The major difference between prokaryotes and eukaryotes is that eukaryotic cells contain membrane-bound compartments in which specific metabolic activities take place. Most important among these is a cell nucleus, a membrane-delineated compartment that houses the eukaryotic cell's DNA. This nucleus gives the eukaryote its name, which means "true nucleus." Other differences include: The plasma membrane resembles that of prokaryotes in function, with minor differences in the setup. Cell walls may or may not be present. The eukaryotic DNA is organized in one or more linear molecules, called chromosomes, which are associated with histone proteins. All chromosomal DNA is stored in the cell nucleus, separated from the cytoplasm by a membrane. Some eukaryotic organelles such as mitochondria also contain some DNA. Many eukaryotic cells are ciliated with primary cilia. Primary cilia play important roles in chemosensation, mechanosensation, and thermosensation. Cilia may thus be "viewed as sensory cellular antennae that coordinate a large number of cellular signaling pathways, sometimes coupling the signaling to ciliary motility or alternatively to cell division and differentiation."[7] Eukaryotes can move using motile cilia or flagella. The flagella are more complex than those of prokaryotes.

Structure of a typical animal cell

Structure of a typical plant cell

Table 2: Comparison of structures between animal and plant cells

Typical animal cell Organelles Nucleus Nucleolus (within nucleus) Rough endoplasmic reticulum (ER) Smooth ER Ribosomes Cytoskeleton Golgi apparatus Cytoplasm Mitochondria Vesicles Lysosomes Centrosome Centrioles Typical plant cell Nucleus Nucleolus (within nucleus) Rough ER Smooth ER Ribosomes Cytoskeleton Golgi apparatus (dictiosomes) Cytoplasm Mitochondria Plastids and its derivatives Vacuole(s) Cell wall

Cells

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Subcellular components
All cells, whether prokaryotic or eukaryotic, have a membrane that envelops the cell, separates its interior from its environment, regulates what moves in and out (selectively permeable), and maintains the electric potential of the cell. Inside the membrane, a salty cytoplasm takes up most of the cell volume. All cells possess DNA, the hereditary material of genes, and RNA, containing the information necessary to build various proteins such as enzymes, the cell's primary machinery. There are also other kinds of biomolecules in cells. This article lists these primary components of the cell, then briefly describe their function.

Membrane
The cytoplasm of a cell is surrounded by a cell membrane or plasma membrane. The plasma membrane in plants and prokaryotes is usually covered by a cell wall. This membrane serves to separate and protect a cell from its surrounding environment and is made mostly from a double layer of lipids (hydrophobic fat-like molecules) and hydrophilic phosphorus molecules. Hence, the layer is called a phospholipid bilayer. It may also be called a fluid mosaic membrane. Embedded within this membrane is a variety of protein molecules that act as channels and pumps that move different molecules into and out of the cell. The membrane is said to be 'semi-permeable', in that it can either let a substance (molecule or ion) pass through freely, pass through to a limited extent or not pass through at all. Cell surface membranes also contain receptor proteins that allow cells to detect external signaling molecules such as hormones.

Cytoskeleton
The cytoskeleton acts to organize and maintain the cell's shape; anchors organelles in place; helps during endocytosis, the uptake of external materials by a cell, and cytokinesis, the separation of daughter cells after cell division; and moves parts of the cell in processes of growth and mobility. The eukaryotic cytoskeleton is composed of microfilaments, intermediate filaments and microtubules. There is a great number of proteins associated with them, each controlling a cell's structure by directing, bundling, and aligning filaments. The prokaryotic cytoskeleton is less well-studied but is involved in the maintenance of cell shape, polarity and cytokinesis.[8]

Genetic material

Bovine Pulmonary Artery Endothelial cell: nuclei stained blue, mitochondria stained red, and F-actin, an important component in microfilaments, stained green. Cell imaged on a fluorescent microscope.

Two different kinds of genetic material exist: deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). Most organisms use DNA for their long-term information storage, but some viruses (e.g., retroviruses) have RNA as their genetic material. The biological information contained in an organism is encoded in its DNA or RNA sequence. RNA is also used for information transport (e.g., mRNA) and enzymatic functions (e.g., ribosomal RNA) in organisms that use DNA for the genetic code itself. Transfer RNA (tRNA) molecules are used to add amino acids during protein translation. Prokaryotic genetic material is organized in a simple circular DNA molecule (the bacterial chromosome) in the nucleoid region of the cytoplasm. Eukaryotic genetic material is divided into different, linear molecules called chromosomes inside a discrete nucleus, usually with additional genetic material in some organelles like mitochondria and chloroplasts (see endosymbiotic theory).

Cells A human cell has genetic material contained in the cell nucleus (the nuclear genome) and in the mitochondria (the mitochondrial genome). In humans the nuclear genome is divided into 23 pairs of linear DNA molecules called chromosomes. The mitochondrial genome is a circular DNA molecule distinct from the nuclear DNA. Although the mitochondrial DNA is very small compared to nuclear chromosomes, it codes for 13 proteins involved in mitochondrial energy production and specific tRNAs. Foreign genetic material (most commonly DNA) can also be artificially introduced into the cell by a process called transfection. This can be transient, if the DNA is not inserted into the cell's genome, or stable, if it is. Certain viruses also insert their genetic material into the genome.

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Organelles
The human body contains many different organs, such as the heart, lung, and kidney, with each organ performing a different function. Cells also have a set of "little organs," called organelles, that are adapted and/or specialized for carrying out one or more vital functions. Both eukaryotic and prokaryotic cells have organelles but organelles in eukaryotes are generally more complex and may be membrane bound. There are several types of organelles in a cell. Some (such as the nucleus and golgi apparatus) are typically solitary, while others (such as mitochondria, peroxisomes and lysosomes) can be numerous (hundreds to thousands). The cytosol is the gelatinous fluid that fills the cell and surrounds the organelles.
Cell nucleus eukaryotes only - a cell's information center The cell nucleus is the most conspicuous organelle found in a eukaryotic cell. It houses the cell's chromosomes, and is the place where almost all DNA replication and RNA synthesis (transcription) occur. The nucleus is spherical and separated from the cytoplasm by a double membrane called the nuclear envelope. The nuclear envelope isolates and protects a cell's DNA from various molecules that could accidentally damage its structure or interfere with its processing. During processing, DNA is transcribed, or copied into a special RNA, called messenger RNA (mRNA). This mRNA is then transported out of the nucleus, where it is translated into a specific protein molecule. The nucleolus is a specialized region within the nucleus where ribosome subunits are assembled. In prokaryotes, DNA processing takes place in the cytoplasm.

Diagram of a cell nucleus Mitochondria and Chloroplasts eukaryotes only - the power generators Mitochondria are self-replicating organelles that occur in various numbers, shapes, and sizes in the cytoplasm of all eukaryotic cells. Mitochondria play a critical role in generating energy in the eukaryotic cell. Mitochondria generate the cell's energy by oxidative phosphorylation, using oxygen to release energy stored in cellular nutrients (typically pertaining to glucose) to generate ATP. Mitochondria multiply by splitting in two. Respiration occurs in the cell mitochondria. Endoplasmic reticulum eukaryotes only The endoplasmic reticulum (ER) is the transport network for molecules targeted for certain modifications and specific destinations, as compared to molecules that float freely in the cytoplasm. The ER has two forms: the rough ER, which has ribosomes on its surface and secretes proteins into the cytoplasm, and the smooth ER, which lacks them. Smooth ER plays a role in calcium sequestration and release.

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Golgi apparatus eukaryotes only The primary function of the Golgi apparatus is to process and package the macromolecules such as proteins and lipids that are synthesized by the cell.

Diagram of an endomembrane system

Ribosomes The ribosome is a large complex of RNA and protein molecules. They each consist of two subunits, and act as an assembly line where RNA from the nucleus is used to synthesise proteins from amino acids. Ribosomes can be found either floating freely or bound to a membrane (the rough endoplasmatic reticulum in [9] eukaryotes, or the cell membrane in prokaryotes). Lysosomes and Peroxisomes eukaryotes only Lysosomes contain digestive enzymes (acid hydrolases). They digest excess or worn-out organelles, food particles, and engulfed viruses or bacteria. Peroxisomes have enzymes that rid the cell of toxic peroxides. The cell could not house these destructive enzymes if they were not contained in a membrane-bound system. Centrosome the cytoskeleton organiser The centrosome produces the microtubules of a cell a key component of the cytoskeleton. It directs the transport through the ER and the Golgi apparatus. Centrosomes are composed of two centrioles, which separate during cell division and help in the formation of the mitotic spindle. A single centrosome is present in the animal cells. They are also found in some fungi and algae cells. Vacuoles Vacuoles store food and waste. Some vacuoles store extra water. They are often described as liquid filled space and are surrounded by a membrane. Some cells, most notably Amoeba, have contractile vacuoles, which can pump water out of the cell if there is too much water. The vacuoles of eukaryotic cells are usually larger in those of plants than animals.

Structures outside the cell wall

Capsule
A gelatinous capsule is present in some bacteria outside the cell wall. The capsule may be polysaccharide as in pneumococci, meningococci or polypeptide as Bacillus anthracis or hyaluronic acid as in streptococci. Capsules are not marked by ordinary stain and can be detected by special stain.

Flagella
Flagella are the organelles of cellular mobility. They arise from cytoplasm and extrude through the cell wall. They are long and thick thread-like appendages, protein in nature. Are most commonly found in bacteria cells but are found in animal cells as well.

Cells

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Fimbriae (pili)
They are short and thin hair like filaments, formed of protein called pilin (antigenic). Fimbriae are responsible for attachment of bacteria to specific receptors of human cell (adherence). There are special types of pili called (sex pili) involved in conjunction.

Functions
Growth and metabolism
Between successive cell divisions, cells grow through the functioning of cellular metabolism. Cell metabolism is the process by which individual cells process nutrient molecules. Metabolism has two distinct divisions: catabolism, in which the cell breaks down complex molecules to produce energy and reducing power, and anabolism, in which the cell uses energy and reducing power to construct complex molecules and perform other biological functions. Complex sugars consumed by the organism can be broken down into a less chemically complex sugar molecule called glucose. Once inside the cell, glucose is broken down to make adenosine triphosphate (ATP), a form of energy, through two different pathways. The first pathway, glycolysis, requires no oxygen and is referred to as anaerobic metabolism. Each reaction is designed to produce some hydrogen ions that can then be used to make energy packets (ATP). In prokaryotes, glycolysis is the only method used for converting energy. The second pathway, called the Krebs cycle, or citric acid cycle, occurs inside the mitochondria and can generate enough ATP to run all the cell functions.

Cells

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Creation
Cell division involves a single cell (called a mother cell) dividing into two daughter cells. This leads to growth in multicellular organisms (the growth of tissue) and to procreation (vegetative reproduction) in unicellular organisms. Prokaryotic cells divide by binary fission. Eukaryotic cells usually undergo a process of nuclear division, called mitosis, followed by division of the cell, called cytokinesis. A diploid cell may also undergo meiosis to produce haploid cells, usually four. Haploid cells serve as gametes in multicellular organisms, fusing to form new diploid cells. DNA replication, or the process of duplicating a cell's genome, is required every time a cell divides. Replication, like all cellular activities, requires specialized proteins for carrying out the job.

Protein synthesis
Cells are capable of synthesizing new proteins, which are essential for the modulation and maintenance of cellular activities. This process involves the formation of new protein molecules from amino acid building blocks based on information encoded in DNA/RNA. Protein synthesis generally consists of two major steps: transcription and translation. Transcription is the process where genetic information in DNA is used to produce a complementary RNA strand. This RNA strand is then processed to give messenger RNA (mRNA), which is free to migrate through the cell. mRNA molecules bind to protein-RNA complexes called ribosomes located in the cytosol, where they are translated into polypeptide sequences. The ribosome mediates the formation of a polypeptide sequence based on the mRNA sequence. The mRNA sequence directly relates to the polypeptide sequence by binding to transfer RNA (tRNA) adapter molecules in binding pockets within the ribosome. The new polypeptide then folds into a functional three-dimensional protein molecule.

Movement or motility

Cells can move during many processes: such as wound healing, the immune response and cancer metastasis. For wound healing to occur, white blood cells and cells that ingest bacteria move to the wound site to kill the microorganisms that cause infection. At the same time fibroblasts (connective tissue cells) move there to remodel damaged structures. In the case of tumor development, cells from a primary tumor move away and spread to other parts of the body. Cell motility involves many receptors, crosslinking, bundling, binding, adhesion, motor and other proteins.[10] The process is divided into three steps protrusion of the leading edge of the cell, adhesion of the leading edge and de-adhesion at the cell body and rear, and cytoskeletal contraction to pull the cell forward. Each step is driven by physical forces generated by unique segments of the cytoskeleton.[11] [12]

An overview of protein synthesis.Within the nucleus of the cell (light blue), genes (DNA, dark blue) are transcribed into RNA. This RNA is then subject to post-transcriptional modification and control, resulting in a mature mRNA (red) that is then transported out of the nucleus and into the cytoplasm (peach), where it undergoes translation into a protein. mRNA is translated by ribosomes (purple) that match the three-base codons of the mRNA to the three-base anti-codons of the appropriate tRNA. Newly synthesized proteins (black) are often further modified, such as by binding to an effector molecule (orange), to become fully active.

Cells

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Evolution
The origin of cells has to do with the origin of life, which began the history of life on Earth.

Origin of the first cell

Further information: Abiogenesis There are several theories about the origin of small molecules that could lead to life in an early Earth. One is that they came from meteorites (see Murchison meteorite). Another is that they were created at deep-sea vents. A third is that they were synthesized by lightning in a reducing atmosphere (see MillerUrey experiment); although it is not clear if Earth had such an atmosphere. There are essentially no experimental data defining what the first self-replicating forms were. RNA is generally assumed the earliest self-replicating molecule, as it is capable of both storing genetic information and catalyzing chemical reactions (see RNA world hypothesis). But some other entity with the potential to self-replicate could have preceded RNA, like clay or peptide nucleic acid.[13] Cells emerged at least 4.04.3 billion years ago. The current belief is that these cells were heterotrophs. An important characteristic of cells is the cell membrane, composed of a bilayer of lipids. The early cell membranes were probably more simple and permeable than modern ones, with only a single fatty acid chain per lipid. Lipids are known to spontaneously form bilayered vesicles in water, and could have preceded RNA, but the first cell membranes could also have been produced by catalytic RNA, or even have required structural proteins before they could form.[14]

Origin of eukaryotic cells

The eukaryotic cell seems to have evolved from a symbiotic community of prokaryotic cells. DNA-bearing organelles like the mitochondria and the chloroplasts are almost certainly what remains of ancient symbiotic oxygen-breathing proteobacteria and cyanobacteria, respectively, where the rest of the cell appears derived from an ancestral archaean prokaryote cellan idea called the endosymbiotic theory. There is still considerable debate about whether organelles like the hydrogenosome predated the origin of mitochondria, or viceversa: see the hydrogen hypothesis for the origin of eukaryotic cells. Sex, as the stereotyped choreography of meiosis and syngamy that persists in nearly all extant eukaryotes, may have played a role in the transition from prokaryotes to eukaryotes. An 'origin of sex as vaccination' theory suggests that the eukaryote genome accreted from prokaryan parasite genomes in numerous rounds of lateral gene transfer. Sex-as-syngamy (fusion sex) arose when infected hosts began swapping nuclearized genomes containing co-evolved, vertically transmitted symbionts that conveyed protection against horizontal infection by more virulent symbionts.[15]

History
16321723: Antonie van Leeuwenhoek teaches himself to grind lenses, builds a microscope and draws protozoa, such as Vorticella from rain water, and bacteria from his own mouth. 1665: Robert Hooke discovers cells in cork, then in living plant tissue using an early microscope.[6] 1839: Theodor Schwann and Matthias Jakob Schleiden elucidate the principle that plants and animals are made of cells, concluding that cells are a common unit of structure and development, and thus founding the cell theory. The belief that life forms can occur spontaneously (generatio spontanea) is contradicted by Louis Pasteur (18221895) (although Francesco Redi had performed an experiment in 1668 that suggested the same conclusion). 1855: Rudolf Virchow states that cells always emerge from cell divisions (omnis cellula ex cellula). 1931: Ernst Ruska builds first transmission electron microscope (TEM) at the University of Berlin. By 1935, he has built an EM with twice the resolution of a light microscope, revealing previously unresolvable organelles.

Cells 1953: Watson and Crick made their first announcement on the double-helix structure for DNA on February 28. 1981: Lynn Margulis published Symbiosis in Cell Evolution detailing the endosymbiotic theory.

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References
[1] Cell Movements and the Shaping of the Vertebrate Body (http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Search& db=books& doptcmdl=GenBookHL& term=Cell+ Movements+ and+ the+ Shaping+ of+ the+ Vertebrate+ Body+ AND+ mboc4[book]+ AND+ 374635[uid]& rid=mboc4. section. 3919) in Chapter 21 of Molecular Biology of the Cell (http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Search& db=books& doptcmdl=GenBookHL& term=cell+ biology+ AND+ mboc4[book]+ AND+ 373693[uid]& rid=mboc4) fourth edition, edited by Bruce Alberts (2002) published by Garland Science. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos. It is also common to describe small molecules such as amino acids as " molecular building blocks (http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Search& db=books& doptcmdl=GenBookHL& term="all+ cells"+ AND+ mboc4[book]+ AND+ 372023[uid]& rid=mboc4. section. 4#23)". [2] Integrative Biology 131 - Lecture 03: Skeletal System (https:/ / www. youtube. com/ watch?v=EvrWHa1PLUQ) on YouTube first 12 minutes of the lecture covers cells (by Marian Diamond). [3] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [4] Mitzi Perdue. "Facts about Birds and Eggs" (http:/ / www. eggscape. com/ birds. htm). . Retrieved 2010-04-15. [5] Maton, Anthea; Hopkins, Jean Johnson, Susan LaHart, David Quon Warner, Maryanna Wright, Jill D (1997). Cells Building Blocks of Life. New Jersey: Prentice Hall. ISBN0-13-423476-6. [6] "... I could exceedingly plainly perceive it to be all perforated and porous, much like a Honey-comb, but that the pores of it were not regular [..] these pores, or cells, [..] were indeed the first microscopical pores I ever saw, and perhaps, that were ever seen, for I had not met with any Writer or Person, that had made any mention of them before this. . ." Hooke describing his observations on a thin slice of cork. Robert Hooke (http:/ / www. ucmp. berkeley. edu/ history/ hooke. html) [7] Satir, P; Christensen, ST; Sren T. Christensen (2008-03-26). "Structure and function of mammalian cilia" (http:/ / www. springerlink. com/ content/ x5051hq648t3152q/ ). Histochemistry and Cell Biology (Springer Berlin / Heidelberg) 129 (6): 687693. doi:10.1007/s00418-008-0416-9. PMC2386530. PMID18365235. 1432-119X. . Retrieved 2009-09-12. [8] Michie K, Lwe J (2006). "Dynamic filaments of the bacterial cytoskeleton". Annu Rev Biochem 75: 46792. doi:10.1146/annurev.biochem.75.103004.142452. PMID16756499. [9] Mntret JF, Schaletzky J, Clemons WM, et al., CW; Akey (December 2007). "Ribosome binding of a single copy of the SecY complex: implications for protein translocation". Mol. Cell 28 (6): 108392. doi:10.1016/j.molcel.2007.10.034. PMID18158904. [10] Revathi Ananthakrishnan1 *, Allen Ehrlicher2 . "The Forces Behind Cell Movement" (http:/ / www. biolsci. org/ v03p0303. htm). Biolsci.org. . Retrieved 2009-04-17. [11] Alberts B, Johnson A, Lewis J. et al. Molecular Biology of the Cell, 4e. Garland Science. 2002 [12] Ananthakrishnan R, Ehrlicher A. The Forces Behind Cell Movement. Int J Biol Sci 2007; 3:303317. http:/ / www. biolsci. org/ v03p0303. htm [13] Orgel LE (1998). "The origin of life--a review of facts and speculations". Trends Biochem Sci 23 (12): 4915. doi:10.1016/S0968-0004(98)01300-0. PMID9868373. [14] Griffiths G (December 2007). "Cell evolution and the problem of membrane topology". Nature reviews. Molecular cell biology 8 (12): 101824. doi:10.1038/nrm2287. PMID17971839. [15] Sterrer W (2002). "On the origin of sex as vaccination". Journal of Theoretical Biology 216: 387396. doi:10.1006/jtbi.2002.3008. PMID12151256.

This article incorporates public domain material from the NCBI document "Science Primer" (http://www. ncbi.nlm.nih.gov/About/primer/index.html).

External links
Inside the Cell (http://publications.nigms.nih.gov/insidethecell/) Virtual Cell's Educational Animations (http://vcell.ndsu.nodak.edu/animations/) The Inner Life of A Cell (http://www.studiodaily.com/main/searchlist/6850.html), a flash video showing what happens inside of a cell. Daniel Reda of Singularity University narrates (beginning at 22:24) (http://www. youtube.com/watch?v=It83JKAxejM) The Virtual Cell (http://www.ibiblio.org/virtualcell/tour/cell/cell.htm) Cells Alive! (http://www.cellsalive.com/) Journal of Cell Biology (http://www.jcb.org/)

Cells The Biology Project > Cell Biology (http://www.biology.arizona.edu/cell_bio/cell_bio.html) Centre of the Cell online (http://www.centreofthecell.org/) The Image & Video Library of The American Society for Cell Biology (http://cellimages.ascb.org/), a collection of peer-reviewed still images, video clips and digital books that illustrate the structure, function and biology of the cell. HighMag Blog (http://highmagblog.blogspot.com/), still images of cells from recent research articles. New Microscope Produces Dazzling 3D Movies of Live Cells (http://www.hhmi.org/news/betzig20110304. html), March 4, 2011 - Howard Hughes Medical Institute. WormWeb.org: Interactive Visualization of the C. elegans Cell lineage (http://wormweb.org/celllineage) Visualize the entire cell lineage tree of the nematode C. elegans

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Textbooks
Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell (http://www. ncbi.nlm.nih.gov/books/bv.fcgi?rid=mboc4.TOC&depth=2) (4th ed.). Garland. ISBN0815332181. Lodish H, Berk A, Matsudaira P, Kaiser CA, Krieger M, Scott MP, Zipurksy SL, Darnell J (2004). Molecular Cell Biology (http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=mcb.TOC) (5th ed.). WH Freeman: New York, NY. ISBN978-0716743668. Cooper GM (2000). The cell: a molecular approach (http://www.ncbi.nlm.nih.gov/books/bv. fcgi?rid=cooper.TOC&depth=2) (2nd ed.). Washington, D.C: ASM Press. ISBN0-87893-102-3.

Water
Water is a chemical substance with the chemical formula H2O. A water molecule contains one oxygen and two hydrogen atoms connected by covalent bonds. Water is a liquid at ambient conditions, but it often co-exists on Earth with its solid state, ice, and gaseous state (water vapor or steam). Water also exists in a liquid crystal state near hydrophilic surfaces.[1] [2] Under nomenclature used to name chemical compounds, Dihydrogen monoxide is the scientific name for water, though it is almost never used.[3]

Water in three states: liquid, solid (ice), and (invisible) water vapor in the air. Clouds are accumulations of water droplets, condensed from vapor-saturated air.

Water covers 70.9% of the Earth's surface,[4] and is vital for all known forms of life.[5] On Earth, 96.5% of the planet's water is found mostly in oceans; 1.7% in groundwater; 1.7% in glaciers and the ice caps of Antarctica and Greenland; a small fraction in other large water bodies, and 0.001% in the air as vapor, clouds (formed of solid and liquid water particles suspended in air), and precipitation.[6] [7] Only 2.5% of the Earth's water is freshwater, and 98.8% of that water is in ice and groundwater. Less than 0.3% of all freshwater is in rivers, lakes, and the atmosphere, and an even smaller amount of the Earth's freshwater (0.003%) is contained within biological bodies and manufactured products.[6] Water on Earth moves continually through the hydrological cycle of evaporation and transpiration (evapotranspiration), condensation, precipitation, and runoff, usually reaching the sea. Evaporation and transpiration contribute to the precipitation over land.

Water Safe drinking water is essential to humans and other lifeforms. Access to safe drinking water has improved over the last decades in almost every part of the world, but approximately one billion people still lack access to safe water and over 2.5 billion lack access to adequate sanitation.[8] There is a clear correlation between access to safe water and GDP per capita.[9] However, some observers have estimated that by 2025 more than half of the world population will be facing water-based vulnerability.[10] A recent report (November 2009) suggests that by 2030, in some developing regions of the world, water demand will exceed supply by 50%.[11] Water plays an important role in the world economy, as it functions as a solvent for a wide variety of chemical substances and facilitates industrial cooling and transportation. Approximately 70% of the fresh water used by humans goes to agriculture.[12]

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Chemical and physical properties

Water is the chemical substance with chemical formula H2O: one molecule of water has two hydrogen atoms covalently bonded to a single oxygen atom. Water appears in nature in all three common states of matter and may take many different forms on Earth: water vapor and clouds in the sky; seawater and icebergs in the polar oceans; glaciers and rivers in the mountains; and the liquid in aquifers in the ground. At high temperatures and pressures, such as in the interior of giant planets, it is argued that water exists as ionic water in which the molecules break down into a soup of hydrogen and oxygen ions, and at even higher pressures as superionic water in which the oxygen crystallises but the hydrogen ions float around freely within the oxygen lattice.[13] The major chemical and physical properties of water are: Water is a liquid at standard temperature and pressure. It is tasteless and odorless. The intrinsic colour of water and ice is a very slight blue hue, although both appear colorless in small quantities. Water vapour is essentially invisible as a gas.[14] Water is transparent in the visible electromagnetic spectrum. Thus aquatic plants can live in water because sunlight can reach them. Infrared light is strongly absorbed by the hydrogen-oxygen or OH bonds.

Model of hydrogen bonds (1) between molecules of water

Since the water molecule is not linear and the oxygen atom has a higher electronegativity than hydrogen atoms, it carries a slight negative charge, whereas the hydrogen atoms are slightly positive. As a result, water is a polar molecule with an electrical dipole moment. Water also can form an unusually large number of intermolecular hydrogen bonds (four) for a molecule of its size. These factors lead to strong attractive forces between molecules of water, giving rise to water's high surface tension[15] and capillary forces. The capillary action refers to the tendency of water to move up a narrow tube against the force of gravity. This property is relied upon by all vascular plants, such as trees.[16]

Impact from a water drop causes an upward "rebound" jet surrounded by circular capillary waves.

Water Water is a good solvent and is often referred to as the universal solvent. Substances that dissolve in water, e.g., salts, sugars, acids, alkalis, and some gases especially oxygen, carbon dioxide (carbonation) are known as hydrophilic (water-loving) substances, while those that do not mix well with water (e.g., fats and oils), are known as hydrophobic (water-fearing) substances. All the major components in cells (proteins, DNA and polysaccharides) are also dissolved in water. Pure water has a low electrical conductivity, but this increases significantly with the dissolution of a small amount of ionic material such as sodium chloride. The boiling point of water (and all other liquids) is dependent on the barometric pressure. For example, on the top of Mt. Everest water boils at 68 C (154F), compared to 100 C (212F) at sea level. Conversely, water deep in the ocean near geothermal vents can reach temperatures of hundreds of degrees and remain liquid. At 4181.3 J/(kgK), water has the second highest specific heat capacity of any known substance (after ammonia), as well as a high heat of vaporization (40.65 kJmol1), both of which are a result of the extensive hydrogen bonding between its molecules. These two unusual properties allow water to moderate Earth's climate by buffering large fluctuations in temperature. The maximum density of water occurs at 3.98 C (39.16F).[17] It has the anomalous property of becoming less dense, not more, when it is cooled down to its solid form, ice. It expands to occupy 9% greater volume in this solid state, which accounts for the fact of ice floating on liquid water, as in icebergs. Its density is 1,000kg/m3 liquid (4C), weighs 62.4lb/ft.3 (917kg/m3, solid). It weighs 8.3454lb/gal. (US, liquid).[18]

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Snowflakes by Wilson Bentley, 1902

Dew drops adhering to a spider web

Capillary action of water compared to mercury

Water Water is miscible with many liquids, such as ethanol, in all proportions, forming a single homogeneous liquid. On the other hand, water and most oils are immiscible, usually forming layers according to increasing density from the top. As a gas, water vapor is completely miscible with air. Water forms an azeotrope with many other solvents. Water can be split by electrolysis into hydrogen and oxygen. As an oxide of hydrogen, water is formed when hydrogen or hydrogen-containing compounds burn or react with oxygen or oxygen-containing compounds. Water is not a fuel, it is an end-product of the combustion of hydrogen. The energy required to split water into hydrogen and oxygen by electrolysis or any other means is greater than the energy that can be collected when the hydrogen and oxygen recombine.[19]

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ADR label for transporting goods dangerously reactive with water

Elements which are more electropositive than hydrogen such as lithium, sodium, calcium, potassium and caesium displace hydrogen from water, forming hydroxides. Being a flammable gas, the hydrogen given off is dangerous and the reaction of water with the more electropositive of these elements may be violently explosive.

Taste and odor

Water can dissolve many different substances, giving it varying tastes and odors. Humans and other animals have developed senses that enable them to evaluate the potability of water by avoiding water that is too salty or putrid. The taste of spring water and mineral water, often advertised in marketing of consumer products, derives from the minerals dissolved in it. However, pure H2O is tasteless and odorless. The advertised purity of spring and mineral water refers to absence of toxins, pollutants and microbes, not the absence of naturally occurring minerals.

Distribution in nature
In the universe
Much of the universe's water is produced as a byproduct of star formation. When stars are born, their birth is accompanied by a strong outward wind of gas and dust. When this outflow of material eventually impacts the surrounding gas, the shock waves that are created compress and heat the gas. The water observed is quickly produced in this warm dense gas.[20] On 22 July 2011, a report described the discovery of a gigantic cloud of water vapor, containing "140 trillion times more water than all of Earth's oceans combined," around a quasar located 12 billion light years from Earth. According to the researchers, the "discovery shows that water has been prevalent in the universe for nearly its entire existence."[21] [22] Water has been detected in interstellar clouds within our galaxy, the Milky Way. Water probably exists in abundance in other galaxies, too, because its components, hydrogen and oxygen, are among the most abundant elements in the universe. Interstellar clouds eventually condense into solar nebulae and solar systems such as ours. Water vapor is present in Atmosphere of Mercury: 3.4%, and large amounts of water in Mercury's exosphere[23] Atmosphere of Venus: 0.002% Earth's atmosphere: ~0.40% over full atmosphere, typically 14% at surface Atmosphere of Mars: 0.03% Atmosphere of Jupiter: 0.0004%

Water Atmosphere of Saturn in ices only Enceladus (moon of Saturn): 91% exoplanets known as HD 189733 b[24] and HD 209458 b.[25] Liquid water is present on Earth: 71% of surface Europa: 100km deep subsurface ocean Strong evidence suggests that liquid water is present just under the surface of Saturn's moon Enceladus. Water ice is present on Earth mainly as ice sheets polar ice caps on Mars Moon Titan Europa Saturn's rings[26] Enceladus Pluto and Charon[26]

25

Comets and comet source populations (Kuiper belt and Oort cloud objects). Water ice may be present on Ceres and Tethys. Water and other volatiles probably comprise much of the internal structures of Uranus and Neptune and the water in the deeper layers may be in the form of ionic water in which the molecules break down into a soup of hydrogen and oxygen ions, and deeper down as superionic water in which the oxygen crystallises but the hydrogen ions float around freely within the oxygen lattice.[13] Some of the Moon's minerals contain water molecules. For instance, in 2008 a laboratory device which ejects and identifies particles found small amounts of the compound in the inside of volcanic rock brought from Moon to Earth by the Apollo 15 crew in 1971.[27] NASA reported the detection of water molecules by NASA's Moon Mineralogy Mapper aboard the Indian Space Research Organization's Chandrayaan-1 spacecraft in September 2009.[28]

Water and habitable zone

The existence of liquid water, and to a lesser extent its gaseous and solid forms, on Earth are vital to the existence of life on Earth as we know it. The Earth is located in the habitable zone of the solar system; if it were slightly closer to or farther from the Sun (about 5%, or about 8 million kilometers), the conditions which allow the three forms to be present simultaneously would be far less likely to exist.[29] [30] Earth's gravity allows it to hold an atmosphere. Water vapor and carbon dioxide in the atmosphere provide a temperature buffer (greenhouse effect) which helps maintain a relatively steady surface temperature. If Earth were smaller, a thinner atmosphere would allow temperature extremes, thus preventing the accumulation of water except in polar ice caps (as on Mars). The surface temperature of Earth has been relatively constant through geologic time despite varying levels of incoming solar radiation (insolation), indicating that a dynamic process governs Earth's temperature via a combination of greenhouse gases and surface or atmospheric albedo. This proposal is known as the Gaia hypothesis. The state of water on a planet depends on ambient pressure, which is determined by the planet's gravity. If a planet is sufficiently massive, the water on it may be solid even at high temperatures, because of the high pressure caused by gravity, as it was observed on exoplanets Gliese 436 b[31] and GJ 1214 b.[32] There are various theories about origin of water on Earth.

Water

26

On Earth
Hydrology is the study of the movement, distribution, and quality of water throughout the Earth. The study of the distribution of water is hydrography. The study of the distribution and movement of groundwater is hydrogeology, of glaciers is glaciology, of inland waters is limnology and distribution of oceans is oceanography. Ecological processes with hydrology are in focus of ecohydrology. The collective mass of water found on, under, and over the surface of a planet is called the hydrosphere. Earth's approximate water volume (the total water supply of the world) is 1,338,000,000km3 (321,000,000mi3).[6] Liquid water is found in bodies of water, such as an ocean, sea, lake, river, stream, canal, pond, or puddle. The majority of water on Earth is sea water. Water is also present in the atmosphere in solid, liquid, and vapor states. It also exists as groundwater in aquifers. Water is important in many geological oceans contain 96.5% of the Earth's water. The processes. Groundwater is present in Antarctic ice sheet, which contains 61% of all fresh water on Earth, is visible at the bottom. most rocks, and the pressure of this Condensed atmospheric water can be seen as groundwater affects patterns of clouds, contributing to the Earth's albedo. faulting. Water in the mantle is responsible for the melt that produces volcanoes at subduction zones. On the surface of the Earth, water is important in both chemical and physical weathering processes. Water and, to a lesser but still significant extent, ice, are also responsible for a large amount of sediment transport that occurs on the surface of the earth. Deposition of transported sediment forms many types of sedimentary rocks, which make up the geologic record of Earth history.
Water covers 71% of the Earth's surface; the

A graphical distribution of the locations of water on Earth.

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Water cycle
The water cycle (known scientifically as the hydrologic cycle) refers to the continuous exchange of water within the hydrosphere, between the atmosphere, soil water, surface water, groundwater, and plants. Water moves perpetually through each of these regions in the water cycle consisting of following transfer processes: evaporation from oceans and other water bodies into the air and transpiration from land plants and animals into air. precipitation, from water vapor condensing from the air and falling to earth or ocean. runoff from the land usually reaching the sea. Most water vapor over the oceans returns to the oceans, but winds carry water vapor over land at the same rate as runoff into the sea, about 47Tt per year. Over land, evaporation and transpiration contribute another 72Tt per year. Precipitation, at a rate of 119 Tt per year over land, has several forms: most commonly rain, snow, and hail, with some contribution from fog and dew.[33] Condensed water in the air may also refract sunlight to produce rainbows. Water runoff often collects over watersheds flowing into rivers. A mathematical model used to simulate river or stream flow and calculate water quality parameters is hydrological transport model. Some of water is diverted to irrigation for agriculture. Rivers and seas offer opportunity for travel and commerce. Through erosion, runoff shapes the environment creating river valleys and deltas which provide rich soil and level ground for the establishment of population centers. A flood occurs when an area of land, usually low-lying, is covered with water. It is when a river overflows its banks or flood from the sea. A drought is an extended period of months or years when a region notes a deficiency in its water supply. This occurs when a region receives consistently below average precipitation.

Water cycle

Fresh water storage

The Bay of Fundy at high tide (left) and low tide (right)

Water Some runoff water is trapped for periods of time, for example in lakes. At high altitude, during winter, and in the far north and south, snow collects in ice caps, snow pack and glaciers. Water also infiltrates the ground and goes into aquifers. This groundwater later flows back to the surface in springs, or more spectacularly in hot springs and geysers. Groundwater is also extracted artificially in wells. This water storage is important, since clean, fresh water is essential to human and other land-based life. In many parts of the world, it is in short supply.

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Sea water
Sea water contains about 3.5% salt on average, plus smaller amounts of other substances. The physical properties of sea water differ from fresh water in some important respects. It freezes at a lower temperature (about 1.9 C) and its density increases with decreasing temperature to the freezing point, instead of reaching maximum density at a temperature above freezing. The salinity of water in major seas varies from about 0.7% in the Baltic Sea to 4.0% in the Red Sea.

Tides
Tides are the cyclic rising and falling of local sea levels caused by the tidal forces of the Moon and the Sun acting on the oceans. Tides cause changes in the depth of the marine and estuarine water bodies and produce oscillating currents known as tidal streams. The changing tide produced at a given location is the result of the changing positions of the Moon and Sun relative to the Earth coupled with the effects of Earth rotation and the local bathymetry. The strip of seashore that is submerged at high tide and exposed at low tide, the intertidal zone, is an important ecological product of ocean tides.

Effects on life
From a biological standpoint, water has many distinct properties that are critical for the proliferation of life that set it apart from other substances. It carries out this role by allowing organic compounds to react in ways that ultimately allow replication. All known forms of life depend on water. Water is vital both as a solvent in which many of the body's solutes dissolve and as an essential part of many metabolic processes within the body. Metabolism is the sum total of anabolism and catabolism. In anabolism, water is removed from molecules (through energy requiring enzymatic chemical reactions) in order to An oasis is an isolated water source with grow larger molecules (e.g. starches, triglycerides and proteins for vegetation in a desert storage of fuels and information). In catabolism, water is used to break bonds in order to generate smaller molecules (e.g. glucose, fatty acids and amino acids to be used for fuels for energy use or other purposes). Without water, these particular metabolic processes could not exist. Water is fundamental to photosynthesis and respiration. Photosynthetic cells use the sun's energy to split off water's hydrogen from oxygen. Hydrogen is combined with CO2 (absorbed from air or water) to form glucose and release oxygen. All living cells use such fuels and oxidize the hydrogen and carbon to capture the sun's energy and reform water and CO2 in the process (cellular respiration).

Water

29

Water is also central to acid-base neutrality and enzyme function. An acid, a hydrogen ion (H+, that is, a proton) donor, can be neutralized by a base, a proton acceptor such as hydroxide ion (OH) to form water. Water is considered to be neutral, with a pH (the negative log of the hydrogen ion concentration) of 7. Acids have pH values less than 7 while bases have values greater than 7.

Overview of photosynthesis and respiration. Water (at right), together with carbon dioxide (CO2), form oxygen and organic compounds (at left), which can be respired to water and (CO2).

Some of the biodiversity of a coral reef

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Aquatic life forms

Earth surface waters are filled with life. The earliest life forms appeared in water; nearly all fish live exclusively in water, and there are many types of marine mammals, such as dolphins and whales. Some kinds of animals, such as amphibians, spend portions of their lives in water and portions on land. Plants such as kelp and algae grow in the water and are the basis for some underwater ecosystems. Plankton is generally the foundation of the ocean food chain. Aquatic vertebrates must obtain oxygen to survive, and they do so in various ways. Fish have gills instead of lungs, although some species of fish, such as the lungfish, have both. Marine mammals, such as dolphins, whales, otters, and seals need to surface periodically to breathe air. Some amphibians are able to absorb oxygen through their skin. Invertebrates exhibit a wide range of modifications to survive in poorly oxygenated waters including breathing tubes (see insect and mollusc siphons) and gills (Carcinus). However as invertebrate life evolved in an aquatic habitat most have little or no specialisation for respiration in water.
Some marine diatoms a key phytoplankton group

Effects on human civilization

Civilization has historically flourished around rivers and major waterways; Mesopotamia, the so-called cradle of civilization, was situated between the major rivers Tigris and Euphrates; the ancient society of the Egyptians depended entirely upon the Nile. Large metropolises like Rotterdam, London, Montreal, Paris, New York City, Buenos Aires, Shanghai, Tokyo, Chicago, and Hong Kong owe their success in part to their easy accessibility via water and the resultant expansion of trade. Islands with safe water ports, like Singapore, have flourished for the same reason. In places such as North Africa and the Middle East, where water is more scarce, access to clean drinking water was and is a major factor in human development.

Water fountain

Health and pollution

Water fit for human consumption is called drinking water or potable water. Water that is not potable may be made potable by filtration or distillation, or by a range of other methods. Water that is not fit for drinking but is not harmful for humans when used for swimming or bathing is called by various names other than potable or drinking water, and is sometimes called safe water, or "safe for bathing". Chlorine is a skin and mucous membrane irritant that is used to make water safe for bathing or drinking. Its use is highly technical and is usually monitored by government regulations Environmental Science Program, Iowa State (typically 1 part per million (ppm) for drinking water, and 12 ppm of University student sampling water. chlorine not yet reacted with impurities for bathing water). Water for bathing may be maintained in satisfactory microbiological condition using chemical disinfectants such as chlorine or ozone or by the use of ultraviolet light.

Water In the USA, non-potable forms of wastewater generated by humans may be referred to as greywater, which is treatable and thus easily able to be made potable again, and blackwater, which generally contains sewage and other forms of waste which require further treatment in order to be made reusable. Greywater composes 5080% of residential wastewater generated by a household's sanitation equipment (sinks, showers and kitchen runoff, but not toilets, which generate blackwater.) These terms may have different meanings in other countries and cultures. This natural resource is becoming scarcer in certain places, and its availability is a major social and economic concern. Currently, about a billion people around the world routinely drink unhealthy water. Most countries accepted the goal of halving by 2015 the number of people worldwide who do not have access to safe water and sanitation during the 2003 G8 Evian summit.[34] Even if this difficult goal is met, it will still leave more than an estimated half a billion people without access to safe drinking water and over a billion without access to adequate sanitation. Poor water quality and bad sanitation are deadly; some five million deaths a year are caused by polluted drinking water. The World Health Organization estimates that safe water could prevent 1.4 million child deaths from diarrhea each year.[35] Water, however, is not a finite resource, but rather re-circulated as potable water in precipitation in quantities many degrees of magnitude higher than human consumption. Therefore, it is the relatively small quantity of water in reserve in the earth (about 1% of our drinking water supply, which is replenished in aquifers around every 1 to 10 years), that is a non-renewable resource, and it is, rather, the distribution of potable and irrigation water which is scarce, rather than the actual amount of it that exists on the earth. Water-poor countries use importation of goods as the primary method of importing water (to leave enough for local human consumption), since the manufacturing process uses around 10 to 100 times products' masses in water. In the developing world, 90% of all wastewater still goes untreated into local rivers and streams.[36] Some 50 countries, with roughly a third of the worlds population, also suffer from medium or high water stress, and 17 of these extract more water annually than is recharged through their natural water cycles.[37] The strain not only affects surface freshwater bodies like rivers and lakes, but it also degrades groundwater resources.

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Human uses
Further information: Water supply Agriculture The most important use of water in agriculture is for irrigation, which is a key component to produce enough food. Irrigation takes up to 90% of water withdrawn in some developing countries[38] and significant proportions in more economically developed countries (United States, 30% of freshwater usage is for irrigation).[39] It takes around 3,000 litres of water, converted from liquid to vapour, to produce enough food to satisfy one person's daily dietary need. This is a considerable amount, when compared to that required for drinking, which is between two and five litres. To produce food for the 6.5 billion or so Irrigation of field crops people who inhabit the planet today requires the water that would fill a canal ten metres deep, 100 metres wide and 7.1 million kilometres long that's enough to circle the globe 180 times. Fifty years ago, the common perception was that water was an infinite resource. At this time, there were fewer than half the current number of people on the planet. People were not as wealthy as today, consumed fewer calories and ate less meat, so less water was needed to produce their food. They required a third of the volume of water we presently take from rivers. Today, the competition for the fixed amount of water resources is much more intense, giving rise to the concept of peak water.[40] This is because there are now nearly seven billion people on the planet, their consumption of water-thirsty meat and vegetables is rising, and there is increasing competition for water from industry, urbanisation and biofuel crops. In future, even more water will be needed to produce food because the

Water Earth's population is forecast to rise to 9 billion by 2050.[41] An additional 2.5 or 3 billion people, choosing to eat fewer cereals and more meat and vegetables could add an additional five million kilometres to the virtual canal mentioned above. An assessment of water management in agriculture was conducted in 2007 by the International Water Management Institute in Sri Lanka to see if the world had sufficient water to provide food for its growing population.[42] It assessed the current availability of water for agriculture on a global scale and mapped out locations suffering from water scarcity. It found that a fifth of the world's people, more than 1.2 billion, live in areas of physical water scarcity, where there is not enough water to meet all demands. A further 1.6 billion people live in areas experiencing economic water scarcity, where the lack of investment in water or insufficient human capacity make it impossible for authorities to satisfy the demand for water. The report found that it would be possible to produce the food required in future, but that continuation of today's food production and environmental trends would lead to crises in many parts of the world. To avoid a global water crisis, farmers will have to strive to increase productivity to meet growing demands for food, while industry and cities find ways to use water more efficiently.[43] As a scientific standard On 7 April 1795, the gram was defined in France to be equal to "the absolute weight of a volume of pure water equal to a cube of one hundredth of a meter, and to the temperature of the melting ice."[44] For practical purposes though, a metallic reference standard was required, one thousand times more massive, the kilogram. Work was therefore commissioned to determine precisely the mass of one liter of water. In spite of the fact that the decreed definition of the gram specified water at 0C a highly reproducible temperature the scientists chose to redefine the standard and to perform their measurements at the temperature of highest water density, which was measured at the time as 4 C (39F).[45] The Kelvin temperature scale of the SI system is based on the triple point of water, defined as exactly 273.16K or 0.01C. The scale is an absolute temperature scale with the same increment as the Celsius temperature scale, which was originally defined according the boiling point (set to 100C) and melting point (set to 0C) of water. Natural water consists mainly of the isotopes hydrogen-1 and oxygen-16, but there is also small quantity of heavier isotopes such as hydrogen-2 (deuterium). The amount of deuterium oxides or heavy water is very small, but it still affects the properties of water. Water from rivers and lakes tends to contain less deuterium than seawater. Therefore, standard water is defined in the Vienna Standard Mean Ocean Water specification. For drinking The human body contains from 55% to 78% water, depending on body size.[46] To function properly, the body requires between one and seven liters of water per day to avoid dehydration; the precise amount depends on the level of activity, temperature, humidity, and other factors. Most of this is ingested through foods or beverages other than drinking straight water. It is not clear how much water intake is needed by healthy people, though most advocates agree that approximately 2 liters (6 to 7 glasses) of water daily is the minimum to maintain proper hydration.[47] Medical literature favors a lower consumption, typically A young girl drinking bottled water 1 liter of water for an average male, excluding extra requirements due to fluid loss from exercise or warm weather.[48] For those who have healthy kidneys, it is rather difficult to drink too much water, but (especially in warm humid weather and while exercising) it is

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Water

33

dangerous to drink too little. People can drink far more water than necessary while exercising, however, putting them at risk of water intoxication (hyperhydration), which can be fatal.[49] [50] The popular claim that "a person should consume eight glasses of water per day" seems to have no real basis in science.[51] Similar misconceptions concerning the effect of water on weight loss and constipation have also been dispelled.[52]

Water quality: fraction of population using improved water sources by country

An original recommendation for water intake in 1945 by the Food and Nutrition Board of the United States National Research Council read: "An ordinary standard for diverse persons is 1 milliliter for each calorie of food. Most of this quantity is contained in prepared foods."[53] The latest dietary reference intake report by the United States National Research Council in general recommended (including food sources): 3.7 liters for men and 2.7 liters of water total for women.[54] Specifically, pregnant and breastfeeding women need additional fluids to stay hydrated. The Institute of Medicine (U.S.) recommends that, on average, men consume 3.0 liters and women 2.2 liters; pregnant women should increase intake to 2.4 liters (10 cups) and breastfeeding women should get 3 liters (12 cups), since an especially large amount Hazard symbol for non-potable water of fluid is lost during nursing.[55] Also noted is that normally, about 20% of water intake comes from food, while the rest comes from drinking water and beverages (caffeinated included). Water is excreted from the body in multiple forms; through urine and feces, through sweating, and by exhalation of water vapor in the breath. With physical exertion and heat exposure, water loss will increase and daily fluid needs may increase as well. Humans require water with few impurities. Common impurities include metal salts and oxides, including copper, iron, calcium and lead,[56] and/or harmful bacteria, such as Vibrio. Some solutes are acceptable and even desirable for taste enhancement and to provide needed electrolytes.[57] The single largest (by volume) freshwater resource suitable for drinking is Lake Baikal in Siberia.[58] Washing The propensity of water to form solutions and emulsions is useful in various washing processes. Many industrial processes rely on reactions using chemicals dissolved in water, suspension of solids in water slurries or using water to dissolve and extract substances. Washing is also an important component of several aspects of personal body hygiene. Transportation The use of water for transportation of materials through rivers and canals as well as the international shipping lanes is an important part of the world economy.

Water Chemical uses Water is widely used in chemical reactions as a solvent or reactant and less commonly as a solute or catalyst. In inorganic reactions, water is a common solvent, dissolving many ionic compounds. In organic reactions, it is not usually used as a reaction solvent, because it does not dissolve the reactants well and is amphoteric (acidic and basic) and nucleophilic. Nevertheless, these properties are sometimes desirable. Also, acceleration of Diels-Alder reactions by water has been observed. Supercritical water has recently been a topic of research. Oxygen-saturated supercritical water combusts organic pollutants efficiently. Heat exchange Water and steam are used as heat transfer fluids in diverse heat exchange systems, due to its availability and high heat capacity, both as a coolant and for heating. Cool water may even be naturally available from a lake or the sea. Condensing steam is a particularly efficient heating fluid because of the large heat of vaporization. A disadvantage is that water and steam are somewhat corrosive. In almost all electric power stations, water is the coolant, which vaporizes and drives steam turbines to drive generators. In the U.S., cooling power plants is the largest use of water.[39] In the nuclear power industry, water can also be used as a neutron moderator. In most nuclear reactors, water is both a coolant and a moderator. This provides something of a passive safety measure, as removing the water from the reactor also slows the nuclear reaction down however other methods are favored for stopping a reaction and it is preferred to keep the nuclear core covered with water so as to ensure adequate cooling. Fire extinction Water has a high heat of vaporization and is relatively inert, which makes it a good fire extinguishing fluid. The evaporation of water carries heat away from the fire. It is dangerous to use water on fires involving oils and organic solvents, because many organic materials float on water and the water tends to spread the burning liquid. Use of water in fire fighting should also take into account the hazards of a steam explosion, which may occur when water is used on very hot fires in confined spaces, and of a hydrogen explosion, when substances which react with water, such as certain metals or hot carbon such as coal, charcoal, coke graphite, decompose the water, producing water gas.

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Water is used for fighting wildfires.

The power of such explosions was seen in the Chernobyl disaster, although the water involved did not come from fire-fighting at that time but the reactor's own water cooling system. A steam explosion occurred when the extreme overheating of the core caused water to flash into steam. A hydrogen explosion may have occurred as a result of reaction between steam and hot zirconium.

Water Recreation Humans use water for many recreational purposes, as well as for exercising and for sports. Some of these include swimming, waterskiing, boating, surfing and diving. In addition, some sports, like ice hockey and ice skating, are played on ice. Lakesides, beaches and waterparks are popular places for people to go to relax and enjoy recreation. Many find the sound and appearance of flowing water to be calming, and fountains and other water features are popular decorations. Some keep fish and other life in aquariums or ponds for show, fun, and companionship. Humans also use water for snow sports i.e. skiing, sledding, snowmobiling or snowboarding, which requires the water to be frozen.

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Grand Anse Beach, St. George's, Grenada, West Indies, often reported as one of the top 10 beaches in the world.

Water industry The water industry provides drinking water and wastewater services (including sewage treatment) to households and industry. Water supply facilities include water wells cisterns for rainwater harvesting, water supply network, water purification facilities, water tanks, water towers, water pipes including old aqueducts. Atmospheric water generators are in development. Drinking water is often collected at springs, extracted from artificial borings (wells) in the ground, or pumped from lakes and rivers. Building more wells in adequate places is thus a possible way to produce more water, assuming the aquifers can supply an adequate flow. Other water sources include rainwater collection. Water may require purification for human consumption. This may involve removal of undissolved substances, dissolved substances and harmful microbes. Popular methods are filtering with sand which only removes undissolved material, while chlorination and boiling kill harmful microbes. Distillation does all three functions. More advanced techniques exist, such as reverse osmosis. Desalination of abundant seawater is a more expensive solution used in coastal arid climates. The distribution of drinking water is done through municipal water systems, tanker delivery or as bottled water. Governments in many countries have programs to distribute water to the needy at no charge. Reducing usage by using drinking (potable) water only for human consumption is another option. In some cities such as Hong Kong, sea water is extensively used for flushing toilets citywide in order to conserve fresh water resources. Polluting water may be the biggest single misuse of water; to the extent that a pollutant limits other uses of the water, it becomes a waste of the resource, regardless of benefits to the polluter. Like other types of

A water-carrier in India, 1882. In many places where running water is not available, water has to be transported by people.

A manual water pump in China

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36 pollution, this does not enter standard accounting of market costs, being conceived as externalities for which the market cannot account. Thus other people pay the price of water pollution, while the private firms' profits are not redistributed to the local population victim of this pollution. Pharmaceuticals consumed by humans often end up in the waterways and can have detrimental effects on aquatic life if they bioaccumulate and if they are not biodegradable.
Water purification facility

Wastewater facilities are storm sewers and wastewater treatment plants. Another way to remove pollution from surface runoff water is bioswale.

Industrial applications Water is used in power generation. Hydroelectricity is electricity obtained from hydropower. Hydroelectric power comes from water driving a water turbine connected to a generator. Hydroelectricity is a low-cost, non-polluting, renewable energy source. The energy is supplied by the motion of water. Typically a dam is constructed on a river, creating an artificial lake behind it. Water flowing out of the lake is forced through turbines that turn generators.

Three Gorges Dam is the largest hydro-electric power station. Pressurized water is used in water blasting and water jet cutters. Also, very high pressure water guns are used for precise cutting. It works very well, is relatively safe, and is not harmful to the environment. It is also used in the cooling of machinery to prevent overheating, or prevent saw blades from overheating. Water is also used in many industrial processes and machines, such as the steam turbine and heat exchanger, in addition to its use as a chemical solvent. Discharge of untreated water from industrial uses is pollution. Pollution includes discharged solutes (chemical pollution) and discharged coolant water (thermal pollution). Industry requires pure water for many applications and utilizes a variety of purification techniques both in water supply and discharge.

Water Food processing Water plays many critical roles within the field of food science. It is important for a food scientist to understand the roles that water plays within food processing to ensure the success of their products. Solutes such as salts and sugars found in water affect the physical properties of water. The boiling and freezing points of water are affected by solutes, as well as air pressure, which is in turn affected by altitude. Water boils at lower temperatures with the lower air pressure which occurs at higher elevations. One mole of sucrose (sugar) per kilogram of water raises the boiling point of water by 0.51 C, and one Water can be used to cook foods such as noodles. mole of salt per kg raises the boiling point by 1.02 C; similarly, increasing the number of dissolved particles lowers water's freezing [59] point. Solutes in water also affect water activity which affects many chemical reactions and the growth of microbes in food.[60] Water activity can be described as a ratio of the vapor pressure of water in a solution to the vapor pressure of pure water.[59] Solutes in water lower water activity. This is important to know because most bacterial growth ceases at low levels of water activity.[60] Not only does microbial growth affect the safety of food but also the preservation and shelf life of food. Water hardness is also a critical factor in food processing. It can dramatically affect the quality of a product as well as playing a role in sanitation. Water hardness is classified based on the amounts of removable calcium carbonate salt it contains per gallon. Water hardness is measured in grains; 0.064 g calcium carbonate is equivalent to one grain of hardness.[59] Water is classified as soft if it contains 1 to 4 grains, medium if it contains 5 to 10 grains and hard if it contains 11 to 20 grains. [59] The hardness of water may be altered or treated by using a chemical ion exchange system. The hardness of water also affects its pH balance which plays a critical role in food processing. For example, hard water prevents successful production of clear beverages. Water hardness also affects sanitation; with increasing hardness, there is a loss of effectiveness for its use as a sanitizer.[59] Boiling, steaming, and simmering are popular cooking methods that often require immersing food in water or its gaseous state, steam. Water is also used for dishwashing.

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Water law, water politics and water crisis

Water politics is politics affected by water and water resources. For this reason, water is a strategic resource in the globe and an important element in many political conflicts. It causes health impacts and damage to biodiversity. 1.6 billion people have gained access to a safe water source since 1990.[61] The proportion of people in developing countries with access to safe water is calculated to have improved from 30% in 1970[62] to 71% in 1990, 79% in 2000 and 84% in 2004. This trend is projected to continue.[8] To halve, by 2015, the proportion of people without sustainable access to safe drinking water is one of the Millennium Development Goals. This goal is projected to be reached.

An estimate of the share of people in developing countries with access to potable water 19702000

A 2006 United Nations report stated that "there is enough water for everyone", but that access to it is hampered by mismanagement and corruption.[63] In addition, global initiatives to improve the efficiency of aid delivery, such as the Paris Declaration on Aid Effectiveness, have not been taken up by water sector donors as effectively as they have in education and health, potentially leaving multiple donors working on overlapping projects and recipient governments without empowerment to act.[64] The authors of the 2007 Comprehensive Assessment of Water Management in Agriculture cited poor governance as one reason for some forms of water scarcity. Water governance is the set of formal and informal processes through which decisions related to water management are made. Good water governance is primarily about knowing what processes work best in a particular physical and socioeconomic context. Mistakes have sometimes been made by trying to apply 'blueprints' that work in the developed world to developing world locations and contexts. The Mekong river is one example; a review by the International Water Management Institute of policies in six countries that rely on the Mekong river for water found that thorough and transparent cost-benefit analyses and environmental impact assessments were rarely undertaken. They also discovered that Cambodia's draft water law was much more complex than it needed to be.[65] The UN World Water Development Report (WWDR, 2003) from the World Water Assessment Program indicates that, in the next 20 years, the quantity of water available to everyone is predicted to decrease by 30%. 40% of the world's inhabitants currently have insufficient fresh water for minimal hygiene. More than 2.2 million people died in 2000 from waterborne diseases (related to the consumption of contaminated water) or drought. In 2004, the UK charity WaterAid reported that a child dies every 15 seconds from easily preventable water-related diseases; often this means lack of sewage disposal; see toilet. Organizations concerned with water protection include International Water Association (IWA), WaterAid, Water 1st, American Water Resources Association [66]. The International Water Management Institute undertakes projects with the aim of using effective water management to reduce poverty. Water related conventions are United Nations Convention to Combat Desertification (UNCCD), International Convention for the Prevention of Pollution from Ships, United Nations Convention on the Law of the Sea and Ramsar Convention. World Day for Water takes place on 22 March and World Ocean Day on 8 June.

Water Water used in the production of a good or service is virtual water.

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In culture
Religion
Water is considered a purifier in most religions. Major faiths that incorporate ritual washing (ablution) include Christianity, Islam, Hinduism, Rastafari movement, Shinto, Taoism, Judaism, and Wicca. Immersion (or aspersion or affusion) of a person in water is a central sacrament of Christianity (where it is called baptism); it is also a part of the practice of other religions, including Judaism (mikvah) and Sikhism (Amrit Sanskar). In addition, a ritual bath in pure water is performed for the dead in many religions including Judaism and Islam. In Islam, the five daily prayers can be done in most cases (see Tayammum) after completing washing certain parts of the body using clean water (wudu). In Shinto, water is used in almost all rituals to cleanse a person or an area (e.g., in the ritual of misogi). Water is mentioned numerous times in the Bible, for example: "The earth was formed out of water and by water" (NIV). In the Qur'an it is stated that "Living things are made of water" and it is often used to describe paradise.

Philosophy
The Ancient Greek philosopher Empedocles held that water is one of the four classical elements along with fire, earth and air, and was regarded as the ylem, or basic substance of the universe. Water was considered cold and moist. In the theory of the four bodily humors, water was associated with phlegm. The classical element of Water was also one of the five elements in traditional Chinese philosophy, along with earth, fire, wood, and metal. Water is also taken as a role model in some parts of traditional and popular Asian philosophy. James Legge's 1891 translation of the Dao De Jing states "The highest excellence is like (that of) water. The excellence of water appears in its benefiting all things, and in its occupying, without striving (to the contrary), the low place which all men dislike. Hence (its way) is near to (that of) the Tao" and "There is nothing in the world more soft and weak than water, and yet for attacking things that are firm and strong there is nothing that can take precedence of itfor there is nothing (so effectual) for which it can be changed."[67]

Literature
Water is used in literature as a symbol of purification. Examples include the critical importance of a river in As I Lay Dying by William Faulkner and the drowning of Ophelia in Hamlet. Sherlock Holmes held that "From a drop of water, a logician could infer the possibility of an Atlantic or a Niagara without having seen or heard of one or the other."[68]

References
[1] Henniker, J. C. (1949). "The Depth of the Surface Zone of a Liquid". Reviews of Modern Physics (Reviews of Modern Physics) 21 (2): 322341. doi:10.1103/RevModPhys.21.322. [2] Pollack, Gerald. "Water Science" (http:/ / faculty. washington. edu/ ghp/ researcthemes/ water-science). University of Washington, Pollack Laboratory. . Retrieved 2011-02-05. "Water has three phases gas, liquid, and solid; but recent findings from our laboratory imply the presence of a surprisingly extensive fourth phase that occurs at interfaces." [3] Bramer, Scott. "Chemical Nomenclature" (http:/ / science. widener. edu/ svb/ pset/ nomen_b. html). Widener University, Department of Chemistry. . Retrieved 20 September 2011. [4] "CIA- The world fact book" (https:/ / www. cia. gov/ library/ publications/ the-world-factbook/ geos/ xx. html#Geo). Central Intelligence Agency. . Retrieved 2008-12-20. [5] "United Nations" (http:/ / www. un. org/ waterforlifedecade/ background. html). Un.org. 2005-03-22. . Retrieved 2010-07-25. [6] Gleick, P.H., ed (1993). Water in Crisis: A Guide to the World's Freshwater Resources. Oxford University Press. p.13, Table 2.1 "Water reserves on the earth". [7] Water Vapor in the Climate System (http:/ / www. agu. org/ sci_soc/ mockler. html), Special Report, [AGU], December 1995 (linked 4/2007). Vital Water (http:/ / www. unep. org/ dewa/ assessments/ ecosystems/ water/ vitalwater/ ) UNEP.

Water
[8] "MDG Report 2008" (http:/ / mdgs. un. org/ unsd/ mdg/ Resources/ Static/ Products/ Progress2008/ MDG_Report_2008_En. pdf#page=44). . Retrieved 2010-07-25. [9] "Public Services" (http:/ / www. gapminder. org/ videos/ gapcasts/ gapcast-9-public-services/ ), Gapminder video [10] Kulshreshtha, S.N (1998). "A Global Outlook for Water Resources to the Year 2025". Water Resources Management 12 (3): 167184. doi:10.1023/A:1007957229865. [11] "Charting Our Water Future: Economic frameworks to inform decision-making" (http:/ / www. mckinsey. com/ App_Media/ Reports/ Water/ Charting_Our_Water_Future_Full_Report_001. pdf) (PDF). . Retrieved 2010-07-25. [12] Baroni, L.; Cenci, L.; Tettamanti, M.; Berati, M. (2007). "Evaluating the environmental impact of various dietary patterns combined with different food production systems". European Journal of Clinical Nutrition 61 (2): 279286. doi:10.1038/sj.ejcn.1602522. PMID17035955. [13] Weird water lurking inside giant planets (http:/ / www. newscientist. com/ article/ mg20727764. 500-weird-water-lurking-inside-giant-planets. html), New Scientist,01 September 2010, Magazine issue 2776. [14] Braun, Charles L.; Sergei N. Smirnov (1993). "Why is water blue?" (http:/ / www. dartmouth. edu/ ~etrnsfer/ water. htm). J. Chem. Educ. 70 (8): 612. doi:10.1021/ed070p612. . [15] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [16] Capillary Action Liquid, Water, Force, and Surface JRank Articles (http:/ / science. jrank. org/ pages/ 1182/ Capillary-Action. html). Science.jrank.org. Retrieved on 2011-11-03. [17] Kotz, J. C., Treichel, P., & Weaver, G. C. (2005). Chemistry & Chemical Reactivity. Thomson Brooks/Cole. ISBN053439597X. [18] Online Conversion Density (http:/ / www. kidsnewsroom. org/ elmer/ infocentral/ conversions/ density. htm), Kidsnewsroom.org/elmer/infocentral/conversions [19] Ball, Philip (14 September 2007). "Burning water and other myths" (http:/ / www. nature. com/ news/ 2007/ 070910/ full/ 070910-13. html). Nature News. . Retrieved 2007-09-14. [20] Melnick, Gary, Harvard-Smithsonian Center for Astrophysics and Neufeld, David, Johns Hopkins University quoted in: "Discover of Water Vapor Near Orion Nebula Suggests Possible Origin of H20 in Solar System (sic)" (http:/ / www. news. harvard. edu/ gazette/ 1998/ 04. 23/ DiscoverofWater. html). The Harvard University Gazette. April 23, 1998. . "Space Cloud Holds Enough Water to Fill Earth's Oceans 1 Million Times" (http:/ / www. jhu. edu/ news_info/ news/ home98/ apr98/ clouds. html). Headlines@Hopkins, JHU. April 9, 1998. . "Water, Water Everywhere: Radio telescope finds water is common in universe" (http:/ / news. harvard. edu/ gazette/ 1999/ 02. 25/ telescope. html). The Harvard University Gazette. February 25, 1999. .(linked 4/2007) [21] Clavin, Whitney; Buis, Alan (22 July 2011). "Astronomers Find Largest, Most Distant Reservoir of Water" (http:/ / www. nasa. gov/ topics/ universe/ features/ universe20110722. html). NASA. . Retrieved 2011-07-25. [22] Staff (22 July 2011). "Astronomers Find Largest, Oldest Mass of Water in Universe" (http:/ / www. space. com/ 12400-universe-biggest-oldest-cloud-water. html). Space.com. . Retrieved 2011-07-23. [23] "MESSENGER Scientists 'Astonished' to Find Water in Mercury's Thin Atmosphere" (http:/ / www. planetary. org/ news/ 2008/ 0703_MESSENGER_Scientists_Astonished_to. html). Planetary Society. 2008-07-03. . Retrieved 2008-07-05. [24] Water Found on Distant Planet (http:/ / www. time. com/ time/ health/ article/ 0,8599,1642811,00. html) July 12, 2007 By Laura Blue, Time [25] Water Found in Extrasolar Planet's Atmosphere (http:/ / www. space. com/ scienceastronomy/ 070410_water_exoplanet. html) Space.com [26] Sparrow, Giles (2006). The Solar System. Thunder Bay Press. ISBN1592235794. [27] Versteckt in Glasperlen: Auf dem Mond gibt es Wasser Wissenschaft [[Der Spiegel (http:/ / www. spiegel. de/ wissenschaft/ weltall/ 0,1518,564911,00. html)] Nachrichten] [28] Water Molecules Found on the Moon (http:/ / science. nasa. gov/ headlines/ y2009/ 24sep_moonwater. htm), NASA, September 24, 2009 [29] Ehlers, E.; Krafft, T, ed (2001). "J. C. I. Dooge. "Integrated Management of Water Resources"". Understanding the Earth System: compartments, processes, and interactions. Springer. p.116. [30] "Habitable Zone" (http:/ / www. daviddarling. info/ encyclopedia/ H/ habzone. html). The Encyclopedia of Astrobiology, Astronomy and Spaceflight. . [31] Shiga, David (6 May 2007). "Strange alien world made of "hot ice"" (http:/ / space. newscientist. com/ article/ dn11864-strange-alien-world-made-of-hot-ice-and-steam. html). New Scientist. . Retrieved 2010-03-28. [32] Aguilar, David A. (16 December 2009). "Astronomers Find Super-Earth Using Amateur, Off-the-Shelf Technology" (http:/ / www. cfa. harvard. edu/ news/ 2009/ pr200924. html). Harvard-Smithsonian Center for Astrophysics. . Retrieved 2010-03-28. [33] Gleick, P.H., ed (1993). Water in Crisis: A Guide to the World's Freshwater Resources. Oxford University Press. p.15, Table 2.3. [34] "G8 "Action plan" decided upon at the 2003 Evian summit" (http:/ / www. g8. fr/ evian/ english/ navigation/ 2003_g8_summit/ summit_documents/ water_-_a_g8_action_plan. html). G8.fr. 2003-06-02. . Retrieved 2010-07-25. [35] "World Health Organization. Safe Water and Global Health" (http:/ / www. who. int/ features/ qa/ 70/ en/ ). Who.int. 2008-06-25. . Retrieved 2010-07-25. [36] UNEP International Environment (2002). Environmentally Sound Technology for Wastewater and Stormwater Management: An International Source Book. IWA Publishing. ISBN1843390086. OCLC49204666. [37] Ravindranath, Nijavalli H.; Jayant A. Sathaye (2002). Climate Change and Developing Countries. Springer. ISBN1402001045. OCLC231965991. [38] "WBCSD Water Facts & Trends" (http:/ / www. wbcsd. org/ includes/ getTarget. asp?type=d& id=MTYyNTA). . Retrieved 2010-07-25. [39] Water Use in the United States (http:/ / nationalatlas. gov/ articles/ water/ a_wateruse. html), National Atlas.gov

40

Water
[40] Gleick, P.H. (2010). "Peak Water" (http:/ / www. pacinst. org/ press_center/ press_releases/ peak_water_pnas. pdf). Proceedings National Academy of Science (National Academy of Science) 107 (125): 1115511162. . Retrieved 2011-10-11. [41] United Nations Press Release POP/952, 13 March 2007. World population will increase by 2.5 billion by 2050 (http:/ / www. un. org/ News/ Press/ docs/ 2007/ pop952. doc. htm) [42] Molden, D. (Ed). Water for food, Water for life: A Comprehensive Assessment of Water Management in Agriculture. Earthscan/IWMI, 2007. [43] Chartres, C. and Varma, S. Out of water. From Abundance to Scarcity and How to Solve the Worlds Water Problems FT Press (USA), 2010 [44] Dcret relatif aux poids et aux mesures. 18 germinal an 3 (7 avril 1795) (http:/ / smdsi. quartier-rural. org/ histoire/ 18germ_3. htm). Decree relating to the weights and measurements (in French). quartier-rural.org [45] here L'Histoire Du Mtre, La Dtermination De L'Unit De Poids (http:/ / histoire. du. metre. free. fr/ fr/ index. htm). histoire.du.metre.free.fr [46] Re: What percentage of the human body is composed of water? (http:/ / www. madsci. org/ posts/ archives/ 2000-05/ 958588306. An. r. html) Jeffrey Utz, M.D., The MadSci Network [47] "Healthy Water Living" (http:/ / www. bbc. co. uk/ health/ healthy_living/ nutrition/ drinks_water. shtml). BBC. . Retrieved 2007-02-01. [48] Rhoades RA, Tanner GA (2003). Medical Physiology (2nd ed.). Baltimore: Lippincott Williams & Wilkins. ISBN0781719364. OCLC50554808. [49] Noakes TD, Goodwin N, Rayner BL, et al. (1985). "Water intoxication: a possible complication during endurance exercise" (http:/ / journals. lww. com/ acsm-msse/ Abstract/ 1985/ 06000/ Water_intoxication__a_possible_complication_during. 12. aspx). Med Sci Sports Exerc 17 (3): 370375. PMID4021781. . [50] Noakes TD, Goodwin N, Rayner BL, Branken T, Taylor RK (2005). "Water intoxication: a possible complication during endurance exercise, 1985". Wilderness Environ Med 16 (4): 2217. PMID16366205. [51] "Drink at least eight glasses of water a day." Really? Is there scientific evidence for "8 8"? (http:/ / ajpregu. physiology. org/ cgi/ content/ full/ 283/ 5/ R993) by Heinz Valdin, Department of Physiology, Dartmouth Medical School, Lebanon, New Hampshire [52] Drinking Water How Much? (http:/ / www. factsmart. org/ h2o/ h2o. htm), Factsmart.org web site and references within [53] Food and Nutrition Board, National Academy of Sciences. Recommended Dietary Allowances.. National Research Council, Reprint and Circular Series, No. 122. 1945. pp.318. [54] Dietary Reference Intakes: Water, Potassium, Sodium, Chloride, and Sulfate (http:/ / www. iom. edu/ report. asp?id=18495), Food and Nutrition Board [55] "Water: How much should you drink every day?" (http:/ / www. mayoclinic. com/ health/ water/ NU00283). Mayoclinic.com. . Retrieved 2010-07-25. [56] "Conquering Chemistry" 4th Ed. Published 2008 [57] Maton, Anthea; Jean Hopkins, Charles William McLaughlin, Susan Johnson, Maryanna Quon Warner, David LaHart, Jill D. Wright (1993). Human Biology and Health. Englewood Cliffs, New Jersey, USA: Prentice Hall. ISBN0-13-981176-1. OCLC32308337. [58] Unesco (2006). Water: a shared responsibility (http:/ / books. google. com/ ?id=1cV8tziHZ5sC& pg=PA125). Berghahn Books. p.125. ISBN1845451775. . [59] Vaclavik, Vickie A. and Christian, Elizabeth W (2007). Essentials of Food Science (http:/ / books. google. com/ ?id=iCCsvwZrguUC& printsec=frontcover). Springer. ISBN0387699392. . [60] DeMan, John M (1999). Principles of Food Chemistry (http:/ / books. google. com/ ?id=kDYJ7a1HbD0C& pg=PA434). Springer. ISBN083421234X. . [61] The Millennium Development Goals Report (http:/ / mdgs. un. org/ unsd/ mdg/ Resources/ Static/ Products/ Progress2008/ MDG_Report_2008_En. pdf#page=44), United Nations, 2008 [62] Lomborg, Bjrn (2001). The Skeptical Environmentalist (http:/ / www. lomborg. com/ dyn/ files/ basic_items/ 69-file/ skeptenvironChap1. pdf). Cambridge University Press. p.22. ISBN0521010683. . [63] UNESCO, (2006), Water, a shared responsibility. The United Nations World Water Development Report 2 (http:/ / unesdoc. unesco. org/ images/ 0014/ 001444/ 144409E. pdf). [64] Welle, Katharina; Evans, Barbara; Tucker, Josephine and Nicol, Alan (2008) Is water lagging behind on Aid Effectiveness? (http:/ / www. odi. org. uk/ resources/ download/ 1894. pdf) [65] Water governance (http:/ / www. iwmi. cgiar. org/ Publications/ Water_Issue_Briefs/ index. aspx), Water Issue Brief, Issue 5, 2010, IWMI [66] http:/ / www. awra. org/ [67] "Internet Sacred Text Archive Home" (http:/ / www. sacred-texts. com/ tao/ taote. htm). Sacred-texts.com. . Retrieved 2010-07-25. [68] Arthur Conan Doyle, A Study in Scarlet, Chapter 2, "The Science of Deduction"

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Water

42

Further reading
Jones, OA., JN Lester and N Voulvoulis, Pharmaceuticals: a threat to drinking water? TRENDS in Biotechnology 23(4): 163, 2005 Franks, F (Ed), Water, A comprehensive treatise, Plenum Press, New York, 19721982 Water in Crisis (http:/ / www. oup. com/ us/ catalog/ general/ subject/ EarthSciences/ Oceanography/ ?view=usa& ci=9780195076288) Gleick,PH., (editor), The World's Water: The Biennial Report on Freshwater Resources. Island Press, Washington, D.C. (published every two years, beginning in 1998.) The World's Water, Island Press (http://www. worldwater.org) Postel,S., Last Oasis: Facing Water Scarcity. W.W. Norton and Company, New York. 1992 Reisner,M., Cadillac Desert: The American West and Its Disappearing Water. Penguin Books, New York. 1986. Debenedetti,PG., and HE Stanley, "Supercooled and Glassy Water", Physics Today 56 (6), p.4046 (2003). Downloadable PDF (1.9 MB) (http://polymer.bu.edu/hes/articles/ds03.pdf) Journal of Contemporary Water Resources and Education (http://ucowr.org/updates/index.html) United Nations World Water Development Report. Produced every three years. UN World Water Development Report (http://www.unesco.org/water/wwap/wwdr/)

External links
OECD Water statistics (http://stats.oecd.org/wbos/Index.aspx?DataSetCode=ENV_WAT) The World's Water Data Page (http://www.worldwater.org) FAO Comprehensive Water Database, AQUASTAT (http://www.fao.org/nr/water/aquastat/main/index.stm) The Water Conflict Chronology: Water Conflict Database (http://worldwater.org/conflict.html) US Geological Survey Water for Schools information (http://ga.water.usgs.gov/edu/) Portal to The World Bank's strategy, work and associated publications on water resources (http://water. worldbank.org/water/)

43

Structural Biochemistry

44

Nucleic acids
Nucleic acid
Nucleic acids are biological molecules essential for life, and include DNA (deoxyribonucleic acid) and RNA (ribonucleic acid). Together with proteins, nucleic acids make up the most important macromolecules; each is found in abundance in all living things, where they function in encoding, transmitting and expressing genetic information. Nucleic acids were first discovered by Friedrich Miescher in 1871.[1] Experimental studies of nucleic acids constitute a major part of modern biological and medical research, and form a foundation for genome and forensic science, as well as the biotechnology and pharmaceutical industries.[2] [3] [4]

Occurrence and nomenclature[5]

The term nucleic acid is the overall name for DNA and RNA, members of a family of biopolymers,[6] and is synonymous with polynucleotide. Nucleic acids were named for their initial discovery within the nucleus, and for the presence of phosphate groups (related to phosphoric acid). Although first discovered within the nucleus of eukaryotic cells, nucleic acids are now known to be found in all life forms, including within bacteria, archaea, mitochondria, chloroplasts, viruses and viroids. All living cells and organelles contain both DNA and RNA, while viruses contain either DNA or RNA, but usually not both.[7] The basic component of biological nucleic acids is the nucleotide, each of which contains a pentose sugar (ribose or deoxyribose), a phosphate group, and a nucleobase. Nucleic acids are also generated within the laboratory, through the use of enzymes[8] (DNA and RNA polymerases) and by solid-phase chemical synthesis. The chemical methods also enable the generation of altered nucleic acids that are not found in nature,[9] for example peptide nucleic acids.

Molecular composition and size[10]

Nucleic acids can vary in size, but are generally very large molecules. Indeed, DNA molecules are probably the largest individual molecules known. Well-studied biological nucleic acid molecules range in size from 21 nucleotides (small interfering RNA) to large chromosomes (human chromosome 1 is a single molecule that contains 247 million base pairs[11] ). In most cases, naturally occurring DNA molecules are double-stranded and RNA molecules are single-stranded. There are numerous exceptions, howeversome viruses have genomes made of double-stranded RNA and other viruses have single-stranded DNA genomes, and, in some circumstances, nucleic acid structures with three or four strands can form. Nucleic acids are linear polymers (chains) of nucleotides. Each nucleotide consists of three components: a purine or pyrimidine nucleobase (sometimes termed nitrogenous base or simply base), a pentose sugar, and a phosphate group. The substructure consisting of a nucleobase plus sugar is termed a nucleoside. Nucleic acid types differ in the structure of the sugar in their nucleotides - DNA contains 2'-deoxyribose while RNA contains ribose (where the only difference is the presence of a hydroxyl group). Also, the nucleobases found in the two nucleic acid types are different: adenine, cytosine, and guanine are found in both RNA and DNA, while thymine occurs in DNA and uracil occurs in RNA. The sugars and phosphates in nucleic acids are connected to each other in an alternating chain (sugar-phosphate backbone) through phosphodiester linkages.[10] In conventional nomenclature, the carbons to which the phosphate groups attach are the 3'-end and the 5'-end carbons of the sugar. This gives nucleic acids directionality, and the ends

Nucleic acid of nucleic acid molecules are referred to as 5'-end and 3'-end. The nucleobases are joined to the sugars via an N-glycosidic linkage involving a nucleobase ring nitrogen (N-1 for pyrimidines and N-9 for purines) and the 1' carbon of the pentose sugar ring. Non-standard nucleosides are also found in both RNA and DNA and usually arise from modification of the standard nucleosides within the DNA molecule or the primary (initial) RNA transcript. Transfer RNA (tRNA) molecules contain a particularly large number of modified nucleosides.[12]

45

Topology
Double-stranded nucleic acids are made up of complementary sequences, in which extensive Watson-Crick base pairing results in the a highly repeated and quite uniform double-helical three-dimensional structure.[13] In contrast, single-stranded RNA and DNA molecules are not constrained to a regular double helix, and can adopt highly complex three-dimensional structures that are based on short stretches of intramolecular base-paired sequences that include both Watson-Crick and noncanonical base pairs, as well as a wide range of complex tertiary interactions.[14] Nucleic acid molecules are usually unbranched, and may occur as linear and circular molecules. For example, bacterial chromosomes, plasmids, mitochondrial DNA and chloroplast DNA are usually circular double-stranded DNA molecules, while chromosomes of the eukaryotic nucleus are usually linear double-stranded DNA molecules.[7] Most RNA molecules are linear, single-stranded molecules, but both circular and branched molecules can result from RNA splicing reactions.[5]

Nucleic acid sequences

One DNA or RNA molecule differs from another primarily in the sequence of nucleotides. Nucleotide sequences are of great importance in biology, since they carry the ultimate instructions that encode all biological molecules, molecular assemblies, subcellular and cellular structures, organs and organisms, and directly enable cognition, memory and behavior (See: Genetics). Enormous efforts have gone into the development of experimental methods to determine the nucleotide sequence of biological DNA and RNA molecules,[15] [16] and today hundreds of millions of nucleotides are sequenced daily at genome centers and smaller laboratories worldwide.

Types of nucleic acids

Deoxyribonucleic acid
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms. The main role of DNA molecules is the long-term storage of information and DNA is often compared to a set of blueprints, since it contains the instructions needed to construct other components of cells, such as proteins and RNA molecules. The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in regulating the use of this genetic information.

Ribonucleic acid
Ribonucleic acid (RNA) functions in converting genetic information from genes into the amino acid sequences of proteins. The three universal types of RNA include transfer RNA (tRNA), messenger RNA (mRNA), and ribosomal RNA (rRNA). Messenger RNA acts to carry genetic sequence information between DNA and ribosomes, directing protein synthesis. Ribosomal RNA is a major component of the ribosome, and catalyzes peptide bond formation. Transfer RNA serves as the carrier molecule for amino acids to be used in protein synthesis, and is responsible for decoding the mRNA. In addition, many other classes of RNA are now known.

Nucleic acid

46

Artificial nucleic acid analogs

Artificial nucleic acid analogs have been designed and synthesized by chemists, and include peptide nucleic acid, morpholino- and locked nucleic acid, as well as glycol nucleic acid and threose nucleic acid. Each of these is distinguished from naturally-occurring DNA or RNA by changes to the backbone of the molecule.

References
[1] Dahm, R (Jan 2008). "Discovering DNA: Friedrich Miescher and the early years of nucleic acid research". Human genetics 122 (6): 56581. doi:10.1007/s00439-007-0433-0. ISSN0340-6717. PMID17901982. [2] International Human Genome Sequencing Consortium (2001). "Initial sequencing and analysis of the human genome." (http:/ / www. nature. com/ nature/ journal/ v409/ n6822/ pdf/ 409860a0. pdf) (PDF). Nature 409 (6822): 860921. doi:10.1038/35057062. PMID11237011. . [3] Venter, JC, et al. (2001). "The sequence of the human genome." (http:/ / www. sciencemag. org/ cgi/ reprint/ 291/ 5507/ 1304. pdf) (PDF). Science 291 (5507): 13041351. doi:10.1126/science.1058040. PMID11181995. . [4] Budowle B, van Daal A (April 2009). "Extracting evidence from forensic DNA analyses: future molecular biology directions". BioTechniques 46 (5): 33940, 34250. doi:10.2144/000113136. PMID19480629. [5] Alberts, Bruce (2008). Molecular biology of the cell. New York: Garland Science. ISBN0-8153-4105-9. [6] Elson D (1965). "Metabolism of nucleic acids (macromolecular DNA and RNA)". Annu. Rev. Biochem. 34: 44986. doi:10.1146/annurev.bi.34.070165.002313. PMID14321176. [7] Brock, Thomas D.; Madigan, Michael T. (2009). Brock biology of microorganisms. Pearson / Benjamin Cummings. ISBN0-321-53615-0. [8] Mullis, Kary B. The Polymerase Chain Reaction (Nobel Lecture). 1993. (retrieved December 1, 2010) http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1993/ mullis-lecture. html [9] Verma S, Eckstein F (1998). "Modified oligonucleotides: synthesis and strategy for users". Annu. Rev. Biochem. 67: 99134. doi:10.1146/annurev.biochem.67.1.99. PMID9759484. [10] Stryer, Lubert; Berg, Jeremy Mark; Tymoczko, John L. (2007). Biochemistry. San Francisco: W.H. Freeman. ISBN0-7167-6766-X. [11] Gregory SG, Barlow KF, McLay KE, et al. (May 2006). "The DNA sequence and biological annotation of human chromosome 1". Nature 441 (7091): 31521. doi:10.1038/nature04727. PMID16710414. [12] Rich A, RajBhandary UL (1976). "Transfer RNA: molecular structure, sequence, and properties". Annu. Rev. Biochem. 45: 80560. doi:10.1146/annurev.bi.45.070176.004105. PMID60910. [13] Watson JD, Crick FH (April 1953). "Molecular structure of nucleic acids; a structure for deoxyribose nucleic acid". Nature 171 (4356): 7378. Bibcode1953Natur.171..737W. doi:10.1038/171737a0. PMID13054692. [14] Ferr-D'Amar AR, Doudna JA (1999). "RNA folds: insights from recent crystal structures". Annu Rev Biophys Biomol Struct 28: 5773. doi:10.1146/annurev.biophys.28.1.57. PMID10410795. [15] Gilbert, Walter G. 1980. DNA Sequencing and Gene Structure (Nobel Lecture) http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1980/ gilbert-lecture. html [16] Sanger, Frederick. 1980. Determination of Nucleotide Sequences in DNA (Nobel Lecture) http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1980/ sanger-lecture. html

Further reading
Wolfram Saenger, Principles of Nucleic Acid Structure, 1984, Springer-Verlag New York Inc. Bruce Alberts, Alexander Johnson, Julian Lewis, Martin Raff, Keith Roberts, and Peter Walter Molecular Biology of the Cell, 2007, ISBN 978-0-8153-4105-5. Fourth edition is available online through the NCBI Bookshelf: link (http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=mboc4) Jeremy M Berg, John L Tymoczko, and Lubert Stryer, Biochemistry 5th edition, 2002, W H Freeman. Available online through the NCBI Bookshelf: link (http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=stryer)

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External links
Interview with Aaron Klug, Nobel Laureate for structural elucidation of biologically important nucleic-acid protein complexes (http://www.vega.org.uk/video/programme/122) provided by the Vega Science Trust. Nucleic Acids Research (Journal) (http://nar.oxfordjournals.org/)

RNA
Ribonucleic acid (English pronunciation: /rab.njukle.k sd/), or RNA, is one of the three major macromolecules (along with DNA and proteins) that are essential for all known forms of life. Like DNA, RNA is made up of a long chain of components called nucleotides. Each nucleotide consists of a nucleobase (sometimes called a nitrogenous base), a ribose sugar, and a phosphate group. The sequence of nucleotides allows RNA to encode genetic information. All cellular organisms use messenger RNA (mRNA) to carry the genetic information that directs the synthesis of proteins. In addition, some viruses use RNA instead of DNA as their genetic material; perhaps a reflection of the suggested key role of RNA in the evolutionary history of life on Earth.[1] [2] Like proteins, some RNA molecules play an active role in cells by catalyzing biological reactions, controlling gene expression, or sensing and communicating responses to cellular signals. One of these active processes is protein synthesis, a universal function whereby mRNA molecules direct the assembly of proteins on ribosomes. This process uses transfer RNA (tRNA) molecules to deliver amino acids to the ribosome, where ribosomal RNA (rRNA) links amino acids together to form proteins. In 2011, it was demonstrated that the methylation of mRNA has a critical role in human energy homeostasis. This opens up the field of RNA epigenetics.[3]

The chemical structure of RNA is very similar to that of DNA, with two differences: (a) RNA contains the sugar ribose, while DNA contains the slightly different sugar deoxyribose (a type of ribose that lacks one oxygen atom), and (b) RNA has the nucleobase uracil while DNA contains thymine. Uracil and thymine have similar base-pairing properties. Unlike DNA, most RNA molecules are single-stranded. Single-stranded RNA molecules adopt very complex three-dimensional structures, since they are not restricted to the repetitive double-helical form of double-stranded DNA. RNA is made within living cells by RNA polymerases, enzymes that act to copy a DNA or RNA template into a new RNA strand through processes known as transcription or RNA replication, respectively.

A hairpin loop from a pre-mRNA. Highlighted are the nucleobases (green) and the ribose-phosphate backbone (blue).

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Comparison with DNA

RNA and DNA are both nucleic acids, but differ in three main ways. First, unlike double-stranded DNA, RNA is a single-stranded molecule in many of its biological roles and has a much shorter chain of nucleotides. Second, while DNA contains deoxyribose, RNA contains ribose (in deoxyribose there is no hydroxyl group attached to the pentose ring in the 2' position). These hydroxyl groups make RNA less stable than DNA because it is more prone to hydrolysis. Third, the complementary base to adenine is not thymine, as it is in DNA, but rather uracil, which is an unmethylated form of thymine.[4] Like DNA, most biologically active RNAs, including mRNA, tRNA, rRNA, snRNAs, and other non-coding RNAs, contain Three-dimensional representation of the 50S self-complementary sequences that allow parts of the RNA to fold and ribosomal subunit. RNA is in ochre, protein in blue. The active site is in the middle (red). pair with itself to form double helices. Analysis of these RNAs has revealed that they are highly structured. Unlike DNA, their structures do not consist of long double helices but rather collections of short helices packed together into structures akin to proteins. In this fashion, RNAs can achieve chemical catalysis, like enzymes.[5] For instance, determination of the structure of the ribosomean enzyme that catalyzes peptide bond formationrevealed that its active site is composed entirely of RNA.[6]

Structure
Each nucleotide in RNA contains a ribose sugar, with carbons numbered 1' through 5'. A base is attached to the 1' position, in general, adenine (A), cytosine (C), guanine (G), or uracil (U). Adenine and guanine are purines, cytosine, and uracil are pyrimidines. A phosphate group is attached to the 3' position of one ribose and the 5' position of the next. The phosphate groups have a negative charge each at physiological pH, making RNA a charged molecule (polyanion). The bases may form hydrogen bonds between cytosine and guanine, between adenine and uracil and between guanine and uracil.[7] However, other interactions are possible, such as a group of adenine bases binding to each other in a bulge,[8] or the GNRA tetraloop that has a guanineadenine base-pair.[7]

Watson-Crick base pairs in a siRNA (hydrogen atoms are not shown)

RNA

49 An important structural feature of RNA that distinguishes it from DNA is the presence of a hydroxyl group at the 2' position of the ribose sugar. The presence of this functional group causes the helix to adopt the A-form geometry rather than the B-form most commonly observed in DNA.[9] This results in a very deep and narrow major groove and a shallow and wide minor groove.[10] A second consequence of the presence of the 2'-hydroxyl group is that in conformationally flexible regions of an RNA molecule (that is, not involved in formation of a double helix), it can chemically attack the adjacent phosphodiester bond to cleave the backbone.[11]
Chemical structure of RNA

RNA is transcribed with only four bases (adenine, cytosine, guanine and uracil),[12] but these bases and attached sugars can be modified in numerous ways as the RNAs mature. Pseudouridine (), in which the linkage between uracil and ribose is changed from a CN bond to a CC bond, and ribothymidine (T) are found in various places (the most notable ones being in the TC loop of tRNA).[13] Another notable modified base is hypoxanthine, a deaminated adenine base whose nucleoside is called inosine (I). Inosine plays a key role in the wobble hypothesis of the genetic code.[14]

Secondary structure of a telomerase RNA.

There are nearly 100 other naturally occurring modified nucleosides,[15] of which pseudouridine and nucleosides with 2'-O-methylribose are the most common.[16] The specific roles of many of these modifications in RNA are not fully understood. However, it is notable that, in ribosomal RNA, many of the post-transcriptional modifications occur in highly functional regions, such as the peptidyl transferase center and the subunit interface, implying that they are important for normal function.[17] The functional form of single stranded RNA molecules, just like proteins, frequently requires a specific tertiary structure. The scaffold for this structure is provided by secondary structural elements that are hydrogen bonds within the molecule. This leads to several recognizable "domains" of secondary structure like hairpin loops, bulges, and internal loops.[18] Since RNA is charged, metal ions such as Mg2+ are needed to stabilise many secondary and tertiary structures.[19]

Synthesis
Synthesis of RNA is usually catalyzed by an enzymeRNA polymeraseusing DNA as a template, a process known as transcription. Initiation of transcription begins with the binding of the enzyme to a promoter sequence in the DNA (usually found "upstream" of a gene). The DNA double helix is unwound by the helicase activity of the enzyme. The enzyme then progresses along the template strand in the 3 to 5 direction, synthesizing a complementary RNA molecule with elongation occurring in the 5 to 3 direction. The DNA sequence also dictates where termination of RNA synthesis will occur.[20] RNAs are often modified by enzymes after transcription. For example, a poly(A) tail and a 5' cap are added to eukaryotic pre-mRNA and introns are removed by the spliceosome. There are also a number of RNA-dependent RNA polymerases that use RNA as their template for synthesis of a new strand of RNA. For instance, a number of RNA viruses (such as poliovirus) use this type of enzyme to replicate their genetic material.[21] Also, RNA-dependent RNA polymerase is part of the RNA interference pathway in many

RNA organisms.[22]

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Types of RNA
Overview
Messenger RNA (mRNA) is the RNA that carries information from DNA to the ribosome, the sites of protein synthesis (translation) in the cell. The coding sequence of the mRNA determines the amino acid sequence in the protein that is produced.[23] Many RNAs do not code for protein however (about 97% of the transcriptional output is non-protein-coding in eukaryotes [24] [25] [26] [27] ). These so-called non-coding RNAs ("ncRNA") can be encoded by their own genes (RNA genes), but can also derive from mRNA introns.[28] The most prominent examples of non-coding RNAs are transfer RNA (tRNA) and ribosomal RNA (rRNA), both of which are involved in the process of translation.[4] There are also non-coding RNAs involved in gene regulation, RNA processing and other roles. Certain RNAs are able to catalyse chemical reactions such as cutting and ligating other RNA molecules,[29] and the catalysis of peptide bond formation in the ribosome;[6] these are known as ribozymes.

In translation
Messenger RNA (mRNA) carries information about a protein sequence Structure of a hammerhead ribozyme, a ribozyme to the ribosomes, the protein synthesis factories in the cell. It is coded that cuts RNA so that every three nucleotides (a codon) correspond to one amino acid. In eukaryotic cells, once precursor mRNA (pre-mRNA) has been transcribed from DNA, it is processed to mature mRNA. This removes its intronsnon-coding sections of the pre-mRNA. The mRNA is then exported from the nucleus to the cytoplasm, where it is bound to ribosomes and translated into its corresponding protein form with the help of tRNA. In prokaryotic cells, which do not have nucleus and cytoplasm compartments, mRNA can bind to ribosomes while it is being transcribed from DNA. After a certain amount of time the message degrades into its component nucleotides with the assistance of ribonucleases.[23] Transfer RNA (tRNA) is a small RNA chain of about 80 nucleotides that transfers a specific amino acid to a growing polypeptide chain at the ribosomal site of protein synthesis during translation. It has sites for amino acid attachment and an anticodon region for codon recognition that binds to a specific sequence on the messenger RNA chain through hydrogen bonding.[28] Ribosomal RNA (rRNA) is the catalytic component of the ribosomes. Eukaryotic ribosomes contain four different rRNA molecules: 18S, 5.8S, 28S and 5S rRNA. Three of the rRNA molecules are synthesized in the nucleolus, and one is synthesized elsewhere. In the cytoplasm, ribosomal RNA and protein combine to form a nucleoprotein called a ribosome. The ribosome binds mRNA and carries out protein synthesis. Several ribosomes may be attached to a single mRNA at any time.[23] Nearly all the RNA found in a typical eukaryotic cell is rRNA. Transfer-messenger RNA (tmRNA) is found in many bacteria and plastids. It tags proteins encoded by mRNAs that lack stop codons for degradation and prevents the ribosome from stalling.[30]

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51

Regulatory RNAs
Several types of RNA can downregulate gene expression by being complementary to a part of an mRNA or a gene's DNA. MicroRNAs (miRNA; 21-22nt) are found in eukaryotes and act through RNA interference (RNAi), where an effector complex of miRNA and enzymes can cleave complementary mRNA, block the mRNA from being translated, or accelerate its degradation.[31] [32] While small interfering RNAs (siRNA; 20-25nt) are often produced by breakdown of viral RNA, there are also endogenous sources of siRNAs.[33] [34] siRNAs act through RNA interference in a fashion similar to miRNAs. Some miRNAs and siRNAs can cause genes they target to be methylated, thereby decreasing or increasing transcription of those genes.[35] [36] [37] Animals have Piwi-interacting RNAs (piRNA; 29-30nt) which are active in germline cells and are thought to be a defense against transposons and play a role in gametogenesis.[38] [39] Many prokaryotes have CRISPR RNAs, a regulatory system similar to RNA interference.[40] Antisense RNAs are widespread; most downregulate a gene, but a few are activators of transcription.[41] One way antisense RNA can act is by binding to an mRNA, forming double-stranded RNA that is enzymatically degraded.[42] There are many long noncoding RNAs that regulate genes in eukaryotes,[43] one such RNA is Xist, which coats one X chromosome in female mammals and inactivates it.[44] An mRNA may contain regulatory elements itself, such as riboswitches, in the 5' untranslated region or 3' untranslated region; these cis-regulatory elements regulate the activity of that mRNA.[45] The untranslated regions can also contain elements that regulate other genes.[46]

In RNA processing
Many RNAs are involved in modifying other RNAs. Introns are spliced out of pre-mRNA by spliceosomes, which contain several small nuclear RNAs (snRNA),[4] or the introns can be ribozymes that are spliced by themselves.[47] RNA can also be altered by having its nucleotides modified to other nucleotides than A, C, G and U. In eukaryotes, modifications of RNA nucleotides are generally directed by small nucleolar RNAs (snoRNA; 60-300nt),[28] found in the Uridine to pseudouridine is a common RNA modification. nucleolus and cajal bodies. snoRNAs associate with enzymes and guide them to a spot on an RNA by basepairing to that RNA. These enzymes then perform the nucleotide modification. rRNAs and tRNAs are extensively modified, but snRNAs and mRNAs can also be the target of base modification.[48] [49] RNA can also be methylated.[50] [51]

RNA genomes
Like DNA, RNA can carry genetic information. RNA viruses have genomes composed of RNA which encodes a number of proteins. The viral genome is replicated by some of those proteins, while other proteins protect the genome as the virus particle moves to a new host cell. Viroids are another group of pathogens, but they consist only of RNA, do not encode any protein and are replicated by a host plant cell's polymerase.[52]

In reverse transcription
Reverse transcribing viruses replicate their genomes by reverse transcribing DNA copies from their RNA; these DNA copies are then transcribed to new RNA. Retrotransposons also spread by copying DNA and RNA from one another,[53] and telomerase contains an RNA that is used as template for building the ends of eukaryotic chromosomes.[54]

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Double-stranded RNA
Double-stranded RNA (dsRNA) is RNA with two complementary strands, similar to the DNA found in all cells. dsRNA forms the genetic material of some viruses (double-stranded RNA viruses). Double-stranded RNA such as viral RNA or siRNA can trigger RNA interference in eukaryotes, as well as interferon response in vertebrates.[55] [56]
[57]

Key discoveries in RNA biology

Further information: History of RNA biology Research on RNA has led to many important biological discoveries and numerous Nobel Prizes. Nucleic acids were discovered in 1868 by Friedrich Miescher, who called the material 'nuclein' since it was found in the nucleus.[58] It was later discovered that prokaryotic cells, which do not have a nucleus, also contain nucleic acids. The role of RNA in protein synthesis was suspected already in 1939.[59] Severo Ochoa won the 1959 Nobel Prize in Medicine (shared with Arthur Kornberg) after he discovered an enzyme that can synthesize RNA in the laboratory.[60] Ironically, the enzyme discovered by Ochoa (polynucleotide phosphorylase) was later shown to be responsible for RNA degradation, not RNA synthesis. The sequence of the 77 nucleotides of a yeast tRNA was found by Robert W. Holley in 1965,[61] winning Holley the 1968 Nobel Prize in Medicine (shared with Har Gobind Khorana and Marshall Nirenberg). In 1967, Carl Woese hypothesized that RNA might be catalytic and suggested that the earliest forms of life (self-replicating molecules) could have relied on RNA both to carry genetic information and to catalyze biochemical reactionsan RNA world.[62] [63] During the early 1970s, retroviruses and reverse transcriptase were discovered, showing for the first time that enzymes could copy RNA into DNA (the opposite of the usual route for transmission of genetic information). For this work, David Baltimore, Renato Dulbecco and Howard Temin were awarded a Nobel Prize in 1975. In 1976, Walter Fiers and his team determined the first complete nucleotide sequence of an RNA virus genome, that of bacteriophage MS2.[64] In 1977, introns and RNA splicing were discovered in both mammalian viruses and in cellular genes, resulting in a 1993 Nobel to Philip Sharp and Richard Roberts. Catalytic RNA molecules (ribozymes) were discovered in the early 1980s, leading to a 1989 Nobel award to Thomas Cech and Sidney Altman. In 1990 it was found in petunia that introduced genes can silence similar genes of the plant's own, now known to be a result of RNA interference.[65] [66] At about the same time, 22 nt long RNAs, now called microRNAs, were found to have a role in the development of C. elegans.[67] Studies on RNA interference gleaned a Nobel Prize for Andrew Fire and Craig Mello in 2006, and another Nobel was awarded for studies on transcription of RNA to Roger Kornberg in the same year. The discovery of gene regulatory RNAs has led to attempts to develop drugs made of RNA, such as siRNA, to silence genes.[68]

References
[1] Yarus, Michael (2011). Getting Past the RNA World: The Initial Darwinian Ancestor. Cold Spring Harb Perspect Biol April 2011;3:a003590 First published online April 21, 2010 doi:10.1101/cshperspect.a003590. http:/ / cshperspectives. cshlp. org/ content/ 3/ 4/ a003590. full [2] Cech TR. (2011). The RNA Worlds in Context. Source: Department of Chemistry and Biochemistry, Howard Hughes Medical Institute, University of Colorado, Boulder, Colorado 80309-0215. Cold Spring Harb Perspect Biol. 2011 Feb 16. pii: cshperspect.a006742v1. doi: 10.1101/cshperspect.a006742. [Epub ahead of print] http:/ / cshperspectives. cshlp. org/ content/ early/ 2011/ 02/ 14/ cshperspect. a006742. full. pdf+ html [3] http:/ / www. physorg. com/ news/ 2011-10-links-common-rna-modification-obesity. html [4] Berg JM, Tymoczko JL, Stryer L (2002). Biochemistry (5th ed.). WH Freeman and Company. pp.11819, 781808. ISBN0-7167-4684-0. OCLC179705944 48055706 59502128. [5] Higgs PG (2000). "RNA secondary structure: physical and computational aspects". Quarterly Reviews of Biophysics 33: 199253. doi:10.1017/S0033583500003620. PMID11191843.

RNA
[6] Nissen P, Hansen J, Ban N, Moore PB, Steitz TA (2000). "The structural basis of ribosome activity in peptide bond synthesis". Science 289 (5481): 92030. doi:10.1126/science.289.5481.920. PMID10937990. [7] Lee JC, Gutell RR (2004). "Diversity of base-pair conformations and their occurrence in rRNA structure and RNA structural motifs". J. Mol. Biol. 344 (5): 122549. doi:10.1016/j.jmb.2004.09.072. PMID15561141. [8] Barciszewski J, Frederic B, Clark C (1999). RNA biochemistry and biotechnology. Springer. pp.7387. ISBN0792358627. OCLC52403776. [9] Salazar M, Fedoroff OY, Miller JM, Ribeiro NS, Reid BR (1992). "The DNA strand in DNAoRNA hybrid duplexes is neither B-form nor A-form in solution". Biochemistry 32 (16): 420715. doi:10.1021/bi00067a007. PMID7682844. [10] Hermann T, Patel DJ (2000). "RNA bulges as architectural and recognition motifs". Structure 8 (3): R47R54. doi:10.1016/S0969-2126(00)00110-6. PMID10745015. [11] Mikkola S, Nurmi K, Yousefi-Salakdeh E, Strmberg R, Lnnberg H (1999). "The mechanism of the metal ion promoted cleavage of RNA phosphodiester bonds involves a general acid catalysis by the metal aquo ion on the departure of the leaving group". Perkin transactions 2 (8): 161926. doi:10.1039/a903691a. [12] Jankowski JAZ, Polak JM (1996). Clinical gene analysis and manipulation: Tools, techniques and troubleshooting. Cambridge University Press. pp.14. ISBN0521478960. OCLC33838261. [13] Yu Q, Morrow CD (2001). "Identification of critical elements in the tRNA acceptor stem and TC loop necessary for human immunodeficiency virus type 1 infectivity". J Virol. 75 (10): 49026. doi:10.1128/JVI.75.10.4902-4906.2001. PMID11312362. [14] Elliott MS, Trewyn RW (1983). "Inosine biosynthesis in transfer RNA by an enzymatic insertion of hypoxanthine". J. Biol. Chem. 259 (4): 240710. PMID6365911. [15] Sll D, RajBhandary U (1995). TRNA: Structure, biosynthesis, and function. ASM Press. pp.165. ISBN155581073X. OCLC183036381 30663724. [16] Kiss T (2001). "Small nucleolar RNA-guided post-transcriptional modification of cellular RNAs". The EMBO Journal 20: 361722. doi:10.1093/emboj/20.14.3617. PMC125535. PMID11447102. [17] King TH, Liu B, McCully RR, Fournier MJ (2002). "Ribosome structure and activity are altered in cells lacking snoRNPs that form pseudouridines in the peptidyl transferase center". Molecular Cell 11 (2): 42535. doi:10.1016/S1097-2765(03)00040-6. PMID12620230. [18] Mathews DH, Disney MD, Childs JL, Schroeder SJ, Zuker M, Turner DH (2004). "Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA secondary structure". Proc. Natl. Acad. Sci. USA 101 (19): 728792. Bibcode2004PNAS..101.7287M. doi:10.1073/pnas.0401799101. PMC409911. PMID15123812. [19] Tan ZJ, Chen SJ (2008). "Salt dependence of nucleic acid hairpin stability". Biophys. J. 95 (2): 73852. doi:10.1529/biophysj.108.131524. PMC2440479. PMID18424500. [20] Nudler E, Gottesman ME (2002). "Transcription termination and anti-termination in E. coli". Genes to Cells 7: 75568. doi:10.1046/j.1365-2443.2002.00563.x. PMID12167155. [21] Jeffrey L Hansen, Alexander M Long, Steve C Schultz (1997). "Structure of the RNA-dependent RNA polymerase of poliovirus". Structure 5 (8): 110922. doi:10.1016/S0969-2126(97)00261-X. PMID9309225. [22] Ahlquist P (2002). "RNA-Dependent RNA Polymerases, Viruses, and RNA Silencing". Science 296 (5571): 127073. doi:10.1126/science.1069132. PMID12016304. [23] Cooper GC, Hausman RE (2004). The Cell: A Molecular Approach (3rd ed.). Sinauer. pp.26176, 297, 33944. ISBN0-87893-214-3. OCLC174924833 52121379 52359301 56050609. [24] Mattick JS, Gagen MJ (1 September 2001). "The evolution of controlled multitasked gene networks: the role of introns and other noncoding RNAs in the development of complex organisms" (http:/ / mbe. oxfordjournals. org/ cgi/ pmidlookup?view=long& pmid=11504843). Mol. Biol. Evol. 18 (9): 161130. PMID11504843. . [25] Mattick, JS (2001). "Noncoding RNAs: the architects of eukaryotic complexity" (http:/ / emboreports. npgjournals. com/ cgi/ content/ full/ 2/ 11/ 986). EMBO Reports 2 (11): 98691. doi:10.1093/embo-reports/kve230. PMC1084129. PMID11713189. . [26] Mattick JS (October 2003). "Challenging the dogma: the hidden layer of non-protein-coding RNAs in complex organisms" (http:/ / www. imb-jena. de/ jcb/ journal_club/ mattick2003. pdf). BioEssays : News and Reviews in Molecular, Cellular and Developmental Biology 25 (10): 9309. doi:10.1002/bies.10332. PMID14505360. . [27] Mattick JS (October 2004). "The hidden genetic program of complex organisms" (http:/ / www. sciam. com/ article. cfm?articleID=00045BB6-5D49-1150-902F83414B7F4945). Scientific American 291 (4): 607. doi:10.1038/scientificamerican1004-60. PMID15487671. . [28] Wirta W (2006). Mining the transcriptome methods and applications (http:/ / kth. diva-portal. org/ smash/ get/ diva2:10803/ FULLTEXT01). Stockholm: School of Biotechnology, Royal Institute of Technology. ISBN91-7178-436-5. OCLC185406288. . [29] Rossi, JJ (2004). "Ribozyme diagnostics comes of age". Chemistry & Biology 11 (7): 89495. doi:10.1016/j.chembiol.2004.07.002. PMID15271347. [30] Gueneau de Novoa P, Williams KP (2004). "The tmRNA website: reductive evolution of tmRNA in plastids and other endosymbionts". Nucleic Acids Res. 32 (Database issue): D1048. doi:10.1093/nar/gkh102. PMC308836. PMID14681369. [31] Wu L, Belasco JG (January 2008). "Let me count the ways: mechanisms of gene regulation by miRNAs and siRNAs". Mol. Cell 29 (1): 17. doi:10.1016/j.molcel.2007.12.010. PMID18206964. [32] Matzke MA, Matzke AJM (2004). "Planting the seeds of a new paradigm". PLoS Biology 2 (5): e133. doi:10.1371/journal.pbio.0020133. PMC406394. PMID15138502.

53

RNA
[33] Vazquez F, Vaucheret H, Rajagopalan R, Lepers C, Gasciolli V, Mallory AC, Hilbert J, Bartel DP, Crt P (2004). "Endogenous trans-acting siRNAs regulate the accumulation of Arabidopsis mRNAs". Molecular Cell 16 (1): 6979. doi:10.1016/j.molcel.2004.09.028. PMID15469823. [34] Watanabe T, Totoki Y, Toyoda A, et al. (May 2008). "Endogenous siRNAs from naturally formed dsRNAs regulate transcripts in mouse oocytes". Nature 453 (7194): 53943. doi:10.1038/nature06908. PMID18404146. [35] Sontheimer EJ, Carthew RW (July 2005). "Silence from within: endogenous siRNAs and miRNAs". Cell 122 (1): 912. doi:10.1016/j.cell.2005.06.030. PMID16009127. [36] Doran G (2007). "RNAi Is one suffix sufficient?" (http:/ / libpubmedia. co. uk/ RNAiJ-Issues/ Issue-5/ Doran. htm). Journal of RNAi and Gene Silencing 3 (1): 21719. . [37] Pushparaj PN, Aarthi JJ, Kumar SD, Manikandan J (2008). "RNAi and RNAa - The Yin and Yang of RNAome". Bioinformation 2 (6): 2357. PMC2258431. PMID18317570. [38] Horwich MD, Li C Matranga C, Vagin V, Farley G, Wang P, Zamore PD (2007). "The Drosophila RNA methyltransferase, DmHen1, modifies germline piRNAs and single-stranded siRNAs in RISC". Current Biology 17: 126572. doi:10.1016/j.cub.2007.06.030. PMID17604629. [39] Girard A, Sachidanandam R, Hannon GJ, Carmell MA (2006). "A germline-specific class of small RNAs binds mammalian Piwi proteins". Nature 442 (7099): 199202. doi:10.1038/nature04917. PMID16751776. [40] Horvath P, Barrangou R (2010). "CRISPR/Cas, the Immune System of Bacteria and Archaea" (http:/ / www. sciencemag. org/ cgi/ content/ abstract/ 327/ 5962/ 167). Science 327 (5962): 16770. doi:10.1126/science.1179555. PMID20056882. . [41] Wagner EG, Altuvia S, Romby P (2002). "Antisense RNAs in bacteria and their genetic elements". Adv Genet. 46: 36198. doi:10.1016/S0065-2660(02)46013-0. PMID11931231. [42] Gilbert SF (2003). Developmental Biology (7th ed.). Sinauer. pp.1013. ISBN0878932585. OCLC154656422 154663147 174530692 177000492 177316159 51544170 54743254 59197768 61404850 66754122. [43] Amaral PP, Mattick JS (October 2008). "Noncoding RNA in development". Mammalian genome : official journal of the International Mammalian Genome Society 19 (78): 45492. doi:10.1007/s00335-008-9136-7. PMID18839252. [44] Heard E, Mongelard F, Arnaud D, Chureau C, Vourc'h C, Avner P (1999). "Human XIST yeast artificial chromosome transgenes show partial X inactivation center function in mouse embryonic stem cells". Proc. Natl. Acad. Sci. USA 96 (12): 684146. doi:10.1073/pnas.96.12.6841. PMC22003. PMID10359800. [45] Batey RT (2006). "Structures of regulatory elements in mRNAs". Curr. Opin. Struct. Biol. 16 (3): 299306. doi:10.1016/j.sbi.2006.05.001. PMID16707260. [46] Scotto L, Assoian RK (June 1993). "A GC-rich domain with bifunctional effects on mRNA and protein levels: implications for control of transforming growth factor beta 1 expression" (http:/ / mcb. asm. org/ cgi/ pmidlookup?view=long& pmid=8497272). Mol. Cell. Biol. 13 (6): 358897. PMC359828. PMID8497272. . [47] Steitz TA, Steitz JA (1993). "A general two-metal-ion mechanism for catalytic RNA". Proc. Natl. Acad. Sci. U.S.A. 90 (14): 6498502. doi:10.1073/pnas.90.14.6498. PMC46959. PMID8341661. [48] Xie J, Zhang M, Zhou T, Hua X, Tang L, Wu W (2007). "Sno/scaRNAbase: a curated database for small nucleolar RNAs and cajal body-specific RNAs". Nucleic Acids Res. 35 (Database issue): D1837. doi:10.1093/nar/gkl873. PMC1669756. PMID17099227. [49] Omer AD, Ziesche S, Decatur WA, Fournier MJ, Dennis PP (2003). "RNA-modifying machines in archaea". Molecular Microbiology 48 (3): 61729. doi:10.1046/j.1365-2958.2003.03483.x. PMID12694609. [50] Cavaill J, Nicoloso M, Bachellerie JP (1996). "Targeted ribose methylation of RNA in vivo directed by tailored antisense RNA guides". Nature 383 (6602): 7325. doi:10.1038/383732a0. PMID8878486. [51] Kiss-Lszl Z, Henry Y, Bachellerie JP, Caizergues-Ferrer M, Kiss T (1996). "Site-specific ribose methylation of preribosomal RNA: a novel function for small nucleolar RNAs". Cell 85 (7): 107788. doi:10.1016/S0092-8674(00)81308-2. PMID8674114. [52] Dars JA, Elena SF, Flores R (2006). "Viroids: an Ariadne's thread into the RNA labyrinth". EMBO Rep. 7 (6): 5938. doi:10.1038/sj.embor.7400706. PMC1479586. PMID16741503. [53] Kalendar R, Vicient CM, Peleg O, Anamthawat-Jonsson K, Bolshoy A, Schulman AH (2004). "Large retrotransposon derivatives: abundant, conserved but nonautonomous retroelements of barley and related genomes". Genetics 166 (3): D339. doi:10.1534/genetics.166.3.1437. PMC1470764. PMID15082561. [54] Podlevsky JD, Bley CJ, Omana RV, Qi X, Chen JJ (2008). "The telomerase database". Nucleic Acids Res. 36 (Database issue): D33943. doi:10.1093/nar/gkm700. PMC2238860. PMID18073191. [55] Blevins T et al. (2006). "Four plant Dicers mediate viral small RNA biogenesis and DNA virus induced silencing". Nucleic Acids Res 34 (21): 623346. doi:10.1093/nar/gkl886. PMC1669714. PMID17090584. [56] Jana S, Chakraborty C, Nandi S, Deb JK (2004). "RNA interference: potential therapeutic targets". Appl. Microbiol. Biotechnol. 65 (6): 64957. doi:10.1007/s00253-004-1732-1. PMID15372214. [57] Schultz U, Kaspers B, Staeheli P (2004). "The interferon system of non-mammalian vertebrates". Dev. Comp. Immunol. 28 (5): 499508. doi:10.1016/j.dci.2003.09.009. PMID15062646. [58] Dahm R (2005). "Friedrich Miescher and the discovery of DNA". Developmental Biology 278 (2): 27488. doi:10.1016/j.ydbio.2004.11.028. PMID15680349. [59] Caspersson T, Schultz J (1939). "Pentose nucleotides in the cytoplasm of growing tissues". Nature 143 (3623): 6023. doi:10.1038/143602c0.

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RNA
[60] Ochoa S (1959). "Enzymatic synthesis of ribonucleic acid" (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1959/ ochoa-lecture. pdf). Nobel Lecture. . [61] Holley RW et al. (1965). "Structure of a ribonucleic acid". Science 147 (3664): 146265. doi:10.1126/science.147.3664.1462. PMID14263761. [62] Siebert S (2006). "Common sequence structure properties and stable regions in RNA secondary structures" (http:/ / deposit. ddb. de/ cgi-bin/ dokserv?idn=982323891& dok_var=d1& dok_ext=pdf& filename=982323891. pdf). Dissertation, Albert-Ludwigs-Universitt, Freiburg im Breisgau. pp. 1. . [63] Szathmry E (1999). "The origin of the genetic code: amino acids as cofactors in an RNA world". Trends Genet. 15 (6): 2239. doi:10.1016/S0168-9525(99)01730-8. PMID10354582. [64] Fiers W et al. (1976). "Complete nucleotide-sequence of bacteriophage MS2-RNA: primary and secondary structure of replicase gene". Nature 260 (5551): 5007. Bibcode1976Natur.260..500F. doi:10.1038/260500a0. PMID1264203. [65] Napoli C, Lemieux C, Jorgensen R (1990). "Introduction of a chimeric chalcone synthase gene into petunia results in reversible co-suppression of homologous genes in trans". Plant Cell 2 (4): 27989. doi:10.1105/tpc.2.4.279. PMC159885. PMID12354959. [66] Dafny-Yelin M, Chung SM, Frankman EL, Tzfira T (December 2007). "pSAT RNA interference vectors: a modular series for multiple gene down-regulation in plants". Plant Physiol. 145 (4): 127281. doi:10.1104/pp.107.106062. PMC2151715. PMID17766396. [67] Ruvkun G (2001). "Glimpses of a tiny RNA world". Science 294 (5543): 79799. doi:10.1126/science.1066315. PMID11679654. [68] Fichou Y, Frec C (2006). "The potential of oligonucleotides for therapeutic applications". Trends in Biotechnology 24 (12): 56370. doi:10.1016/j.tibtech.2006.10.003. PMID17045686.

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External links
RNA World website (http://www.imb-jena.de/RNA.html) Link collection (structures, sequences, tools, journals) Nucleic Acid Database (http://ndbserver.rutgers.edu/atlas/xray/) Images of DNA, RNA and complexes. EteRNA (http://eterna.cmu.edu/content/EteRNA) a game forming RNA by pairing bases.

DNA
Deoxyribonucleic acid (English pronunciation: /diksirab.njukle.k sd/( listen); DNA) is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms (with the exception of RNA viruses). The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in regulating the use of this genetic information. Along with RNA and proteins, DNA is one of the three major macromolecules that are essential for all known forms of life. DNA consists of two long polymers of simple units called nucleotides, with backbones made of sugars and phosphate groups joined by ester

The structure of the DNA double helix. The atoms in the structure are colour coded by element and the detailed structure of two base pairs is shown in the bottom right.

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bonds. These two strands run in opposite directions to each other and are therefore anti-parallel. Attached to each sugar is one of four types of molecules called nucleobases (informally, bases). It is the sequence of these four nucleobases along the backbone that encodes information. This information is read using the genetic code, which specifies the sequence of the amino acids within proteins. The code is read by copying stretches of DNA into the related nucleic acid RNA in a process called transcription. Within cells DNA is organized into long structures called chromosomes. During cell division these chromosomes are duplicated in the process of DNA replication, providing each cell its own complete set of chromosomes. Eukaryotic organisms (animals, plants, fungi, and protists) store most of their DNA inside the cell nucleus and some of their DNA in organelles, such as mitochondria or chloroplasts.[1] In contrast, prokaryotes (bacteria and archaea) store their DNA only in the cytoplasm. Within the chromosomes, chromatin proteins such as histones compact and organize DNA. These compact structures guide the interactions between DNA and other proteins, helping control which parts of the DNA are transcribed.
The structure of part of a DNA double helix

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Properties
DNA is a long polymer made from repeating units called nucleotides.[2] [3] [4] As first discovered by James D. Watson and Francis Crick, the structure of DNA of all species comprises two helical chains each coiled round the same axis, and each with a pitch of 34ngstrms (3.4nanometres) and a radius of 10ngstrms (1.0nanometres).[5] According to another study, when measured in a particular solution, the DNA chain measured 22 to 26ngstrms wide (2.2 to 2.6nanometres), and one nucleotide unit measured 3.3 (0.33nm) long.[6] Although each individual repeating unit is very small, DNA polymers can be very large molecules containing millions of nucleotides. For instance, the largest human chromosome, chromosome number 1, is approximately 220 million base pairs long.[7] In living organisms DNA does not usually Chemical structure of DNA. Hydrogen bonds shown as dotted lines. exist as a single molecule, but instead as a pair of molecules that are held tightly together.[5] [8] These two long strands entwine like vines, in the shape of a double helix. The nucleotide repeats contain both the segment of the backbone of the molecule, which holds the chain together, and a nucleobase, which interacts with the other DNA strand in the helix. A nucleobase linked to a sugar is called a nucleoside and a base linked to a sugar and one or more phosphate groups is called a nucleotide. Polymers comprising multiple linked nucleotides (as in DNA) are called a polynucleotide.[9] The backbone of the DNA strand is made from alternating phosphate and sugar residues.[10] The sugar in DNA is 2-deoxyribose, which is a pentose (five-carbon) sugar. The sugars are joined together by phosphate groups that form phosphodiester bonds between the third and fifth carbon atoms of adjacent sugar rings. These asymmetric bonds mean a strand of DNA has a direction. In a double helix the direction of the nucleotides in one strand is opposite to their direction in the other strand: the strands are antiparallel. The asymmetric ends of DNA strands are called the 5 (five prime) and 3 (three prime) ends, with the 5' end having a terminal phosphate group and the 3' end a terminal hydroxyl group. One major difference between DNA and RNA is the sugar, with the 2-deoxyribose in DNA being replaced by the alternative pentose sugar ribose in RNA.[8]

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The DNA double helix is stabilized primarily by two forces: hydrogen bonds between nucleotides and base-stacking interactions among the aromatic nucleobases.[12] In the aqueous environment of the cell, the conjugated bonds of nucleotide bases align perpendicular to the axis of the DNA molecule, minimizing their interaction with the solvation shell and therefore, the Gibbs free energy. The four bases found in DNA are adenine (abbreviated A), cytosine (C), guanine (G) and thymine (T). These four bases are attached to the sugar/phosphate to form the complete nucleotide, as shown for adenosine monophosphate. The nucleobases are classified into two types: the purines, A and G, being fused five- and six-membered heterocyclic compounds, and the pyrimidines, the six-membered rings C and T.[8] A fifth pyrimidine nucleobase, uracil (U), usually takes the place of thymine in RNA and differs from thymine by lacking a methyl group on its ring. Uracil is not usually found in DNA, occurring only as a breakdown product of cytosine. In addition to RNA and DNA a large number of artificial nucleic acid analogues have also been created to study the proprieties of nucleic acids, or for use in biotechnology.[13]

A section of DNA. The bases lie horizontally between the two [11] spiraling strands. Animated version at File:DNA orbit animated.gif.

Grooves
Twin helical strands form the DNA backbone. Another double helix may be found by tracing the spaces, or grooves, between the strands. These voids are adjacent to the base pairs and may provide a binding site. As the strands are not directly opposite each other, the grooves are unequally sized. One groove, the major groove, is 22 wide and the other, the minor groove, is 12 wide.[14] The narrowness of the minor groove means that the edges of the bases are more accessible in the Major and minor grooves of DNA. Minor groove major groove. As a result, proteins like transcription factors that can is a binding site for the dye Hoechst 33258. bind to specific sequences in double-stranded DNA usually make contacts to the sides of the bases exposed in the major groove.[15] This situation varies in unusual conformations of DNA within the cell (see below), but the major and minor grooves are always named to reflect the differences in size that would be seen if the DNA is twisted back into the ordinary B form.

Base pairing
Further information: Base pair In a DNA double helix, each type of nucleobase on one strand normally interacts with just one type of nucleobase on the other strand. This is called complementary base pairing. Here, purines form hydrogen bonds to pyrimidines, with A bonding only to T, and C bonding only to G. This arrangement of two nucleotides binding together across the double helix is called a base pair. As hydrogen bonds are not covalent, they can be broken and rejoined relatively easily. The two strands of DNA in a double helix can therefore be pulled apart like a zipper, either by a mechanical force or high temperature.[16] As a result of this complementarity, all the information in the double-stranded sequence of a DNA helix is duplicated on each strand, which is vital in DNA replication. Indeed, this reversible and specific interaction between complementary base pairs is critical for all the functions of DNA in living organisms.[3]

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Top, a GC base pair with three hydrogen bonds. Bottom, an AT base pair with two hydrogen bonds. Non-covalent hydrogen bonds between the pairs are shown as dashed lines. The two types of base pairs form different numbers of hydrogen bonds, AT forming two hydrogen bonds, and GC forming three hydrogen bonds (see figures, left). DNA with high GC-content is more stable than DNA with low GC-content. Although it is often stated that this is due to the added stability of an additional hydrogen bond, this is incorrect. DNA with high GC-content is more stable due to intra-strand base stacking interactions. As noted above, most DNA molecules are actually two polymer strands, bound together in a helical fashion by noncovalent bonds; this double stranded structure (dsDNA) is maintained largely by the intrastrand base stacking interactions, which are strongest for G,C stacks. The two strands can come apart a process known as melting to form two ss DNA molecules. Melting occurs when conditions favor ssDNA; such conditions are high temperature, low salt and high pH (low pH also melts DNA, but since DNA is unstable due to acid depurination, low pH is rarely used). The stability of the dsDNA form depends not only on the GC-content (% G,C basepairs) but also on sequence (since stacking is sequence specific) and also length (longer molecules are more stable). The stability can be measured in various ways; a common way is the "melting temperature", which is the temperature at which 50% of the ds molecules are converted to ss molecules; melting temperature is dependent on ionic strength and the concentration of DNA. As a result, it is both the percentage of GC base pairs and the overall length of a DNA double helix that determine the strength of the association between the two strands of DNA. Long DNA helices with a high GC-content have stronger-interacting strands, while short helices with high AT content have weaker-interacting strands.[17] In biology, parts of the DNA double helix that need to separate easily, such as the TATAAT Pribnow box in some promoters, tend to have a high AT content, making the strands easier to pull apart.[18] In the laboratory, the strength of this interaction can be measured by finding the temperature required to break the hydrogen bonds, their melting temperature (also called Tm value). When all the base pairs in a DNA double helix melt, the strands separate and exist in solution as two entirely independent molecules. These single-stranded DNA molecules (ssDNA) have no single common shape, but some conformations are more stable than others.[19]

Sense and antisense

Further information: Sense (molecular biology) A DNA sequence is called "sense" if its sequence is the same as that of a messenger RNA copy that is translated into protein.[20] The sequence on the opposite strand is called the "antisense" sequence. Both sense and antisense sequences can exist on different parts of the same strand of DNA (i.e. both strands contain both sense and antisense sequences). In both prokaryotes and eukaryotes, antisense RNA sequences are produced, but the functions of these RNAs are not entirely clear.[21] One proposal is that antisense RNAs are involved in regulating gene expression through RNA-RNA base pairing.[22] A few DNA sequences in prokaryotes and eukaryotes, and more in plasmids and viruses, blur the distinction between sense and antisense strands by having overlapping genes.[23] In these cases, some DNA sequences do double duty, encoding one protein when read along one strand, and a second protein when read in the opposite direction along the

DNA other strand. In bacteria, this overlap may be involved in the regulation of gene transcription,[24] while in viruses, overlapping genes increase the amount of information that can be encoded within the small viral genome.[25]

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Supercoiling
Further information: DNA supercoil DNA can be twisted like a rope in a process called DNA supercoiling. With DNA in its "relaxed" state, a strand usually circles the axis of the double helix once every 10.4 base pairs, but if the DNA is twisted the strands become more tightly or more loosely wound.[26] If the DNA is twisted in the direction of the helix, this is positive supercoiling, and the bases are held more tightly together. If they are twisted in the opposite direction, this is negative supercoiling, and the bases come apart more easily. In nature, most DNA has slight negative supercoiling that is introduced by enzymes called topoisomerases.[27] These enzymes are also needed to relieve the twisting stresses introduced into DNA strands during processes such as transcription and DNA replication.[28]

Alternate DNA structures

Further information: Molecular Structure of Nucleic Acids: A Structure for Deoxyribose Nucleic Acid, Molecular models of DNA, and DNA structure DNA exists in many possible conformations that include A-DNA, B-DNA, and Z-DNA forms, although, only B-DNA and Z-DNA have been directly observed in functional organisms.[10] The conformation that DNA adopts depends on the hydration level, DNA sequence, the amount and direction of supercoiling, chemical modifications of the bases, the type and concentration of metal ions, as well as the presence of polyamines in solution.[29]

From left to right, the structures of A, B and Z DNA

The first published reports of A-DNA X-ray diffraction patterns and also B-DNA used analyses based on Patterson transforms that provided only a limited amount of structural information for oriented fibers of DNA.[30] [31] An alternate analysis was then proposed by Wilkins et al., in 1953, for the in vivo B-DNA X-ray diffraction/scattering patterns of highly hydrated DNA fibers in terms of squares of Bessel functions.[32] In the same journal, James D. Watson and Francis Crick presented their molecular modeling analysis of the DNA X-ray diffraction patterns to suggest that the structure was a double-helix.[5] Although the `B-DNA form' is most common under the conditions found in cells,[33] it is not a well-defined conformation but a family of related DNA conformations[34] that occur at the high hydration levels present in living cells. Their corresponding X-ray diffraction and scattering patterns are characteristic of molecular paracrystals with a significant degree of disorder.[35] [36] Compared to B-DNA, the A-DNA form is a wider right-handed spiral, with a shallow, wide minor groove and a narrower, deeper major groove. The A form occurs under non-physiological conditions in partially dehydrated samples of DNA, while in the cell it may be produced in hybrid pairings of DNA and RNA strands, as well as in enzyme-DNA complexes.[37] [38] Segments of DNA where the bases have been chemically modified by methylation may undergo a larger change in conformation and adopt the Z form. Here, the strands turn about the helical axis in a left-handed spiral, the opposite of the more common B form.[39] These unusual structures can be recognized by specific Z-DNA binding proteins and may be involved in the regulation of transcription.[40]

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Alternate DNA chemistry

For a number of years exobiologists have proposed the existence of a shadow biosphere, a postulated microbial biosphere of Earth that uses radically different biochemical and molecular processes than currently known life. One of the proposals was the existence of lifeforms that use arsenic instead of phosphorus in DNA. A December 2010 NASA press conference stated that the bacterium GFAJ-1, which has evolved in an arsenic-rich environment, is the first terrestrial lifeform found which may have this ability. The bacterium was found in Mono Lake, east of Yosemite National Park. GFAJ-1 is a rod-shaped extremophile bacterium in the family Halomonadaceae that, when starved of phosphorus, may be capable of incorporating the usually poisonous element arsenic in its DNA.[41] This discovery may lend weight to the long-standing idea that extraterrestrial life could have a different chemical makeup from life on Earth.[41] [42] The research was carried out by a team led by Felisa Wolfe-Simon, a geomicrobiologist and geobiochemist, a Postdoctoral Fellow of the NASA Astrobiology Institute with Arizona State University. This finding has, however, faced strong criticism from the scientific community; scientists have argued that there is no evidence that arsenic is actually incorporated into biomolecules.[42] [43] Independent conformation of this finding has also not yet been possible.

Quadruplex structures
Further information: G-quadruplex At the ends of the linear chromosomes are specialized regions of DNA called telomeres. The main function of these regions is to allow the cell to replicate chromosome ends using the enzyme telomerase, as the enzymes that normally replicate DNA cannot copy the extreme 3 ends of chromosomes.[44] These specialized chromosome caps also help protect the DNA ends, and stop the DNA repair systems in the cell from treating them as damage to be corrected.[45] In human cells, telomeres are usually lengths of single-stranded DNA containing several thousand repeats of a simple TTAGGG sequence.[46] These guanine-rich sequences may stabilize chromosome ends by forming structures of stacked sets of four-base units, rather than the usual base pairs found in other DNA molecules. Here, four guanine bases form a flat plate and these flat four-base units then stack on top of each other, to form a stable G-quadruplex structure.[48] These structures are stabilized by hydrogen bonding between the edges of the bases and chelation of a metal ion in the centre of each four-base unit.[49] Other structures can also be formed, with the central set of four bases coming from either a single strand folded around the bases, or several different parallel strands, each contributing one base to the central structure.

In addition to these stacked structures, telomeres also form large loop structures called telomere loops, or T-loops. Here, the single-stranded DNA curls around in a long circle stabilized by telomere-binding proteins.[50] At the very end of the T-loop, the single-stranded telomere DNA is held onto a region of double-stranded DNA by the telomere strand disrupting the double-helical DNA and base pairing to one of the two strands. This triple-stranded structure is called a displacement loop or D-loop.[48]

DNA quadruplex formed by telomere repeats. The looped conformation of the DNA backbone [47] is very different from the typical DNA helix.

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Single branch Multiple branches

Branched DNA can form networks containing multiple branches.

Branched DNA
Further information: Branched DNA and DNA nanotechnology In DNA fraying occurs when non-complementary regions exist at the end of an otherwise complementary double-strand of DNA. However, branched DNA can occur if a third strand of DNA is introduced and contains adjoining regions able to hybridize with the frayed regions of the pre-existing double-strand. Although the simplest example of branched DNA involves only three strands of DNA, complexes involving additional strands and multiple branches are also possible.[51] Branched DNA can be used in nanotechnology to construct geometric shapes, see the section on uses in technology below.

Vibration
DNA may carry out low-frequency collective motion as observed by the Raman spectroscopy[52] with a quasi-continuum model.[54] [55]
[53]

and analyzed

Chemical modifications

cytosine

5-methylcytosine

thymine

Structure of cytosine with and without the 5-methyl group. Deamination converts 5-methylcytosine into thymine.

Base modifications
Further information: DNA methylation The expression of genes is influenced by how the DNA is packaged in chromosomes, in a structure called chromatin. Base modifications can be involved in packaging, with regions that have low or no gene expression usually containing high levels of methylation of cytosine bases. For example, cytosine methylation, produces 5-methylcytosine, which is important for X-chromosome inactivation.[56] The average level of methylation varies between organisms the worm Caenorhabditis elegans lacks cytosine methylation, while vertebrates have higher levels, with up to 1% of their DNA containing 5-methylcytosine.[57] Despite the importance of 5-methylcytosine, it can deaminate to leave a thymine base, so methylated cytosines are particularly prone to mutations.[58] Other base modifications include adenine methylation in bacteria, the presence of 5-hydroxymethylcytosine in the brain,[59] and the glycosylation of uracil to produce the "J-base" in kinetoplastids.[60] [61]

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Damage
Further information: Mutation DNA can be damaged by many sorts of mutagens, which change the DNA sequence. Mutagens include oxidizing agents, alkylating agents and also high-energy electromagnetic radiation such as ultraviolet light and X-rays. The type of DNA damage produced depends on the type of mutagen. For example, UV light can damage DNA by producing thymine dimers, which are cross-links between pyrimidine bases.[63] On the other hand, oxidants such as free radicals or hydrogen peroxide produce multiple forms of damage, including base modifications, particularly of guanosine, and double-strand breaks.[64] A typical human cell contains about 150,000 bases that have suffered oxidative damage.[65] Of these oxidative lesions, the most dangerous are double-strand breaks, as these are difficult to repair and can produce point mutations, insertions and deletions from the DNA sequence, as well as chromosomal translocations.[66] Many mutagens fit into the space between two adjacent base pairs, this A covalent adduct between a metabolically is called intercalation. Most intercalators are aromatic and planar activated form of benzo[a]pyrene, the major molecules; examples include ethidium bromide, acridines, [62] mutagen in tobacco smoke, and DNA daunomycin, and doxorubicin. In order for an intercalator to fit between base pairs, the bases must separate, distorting the DNA strands by unwinding of the double helix. This inhibits both transcription and DNA replication, causing toxicity and mutations.[67] As a result, DNA intercalators may be carcinogens, and in the case of thalidomide, a teratogen.[68] Others such as benzo[a]pyrene diol epoxide and aflatoxin form DNA adducts which induce errors in replication.[69] Nevertheless, due to their ability to inhibit DNA transcription and replication, other similar toxins are also used in chemotherapy to inhibit rapidly growing cancer cells.[70]

Biological functions
DNA usually occurs as linear chromosomes in eukaryotes, and circular chromosomes in prokaryotes. The set of chromosomes in a cell makes up its genome; the human genome has approximately 3 billion base pairs of DNA arranged into 46 chromosomes.[71] The information carried by DNA is held in the sequence of pieces of DNA called genes. Transmission of genetic information in genes is achieved via complementary base pairing. For example, in transcription, when a cell uses the information in a gene, the DNA sequence is copied into a complementary RNA sequence through the attraction between the DNA and the correct RNA nucleotides. Usually, this RNA copy is then used to make a matching protein sequence in a process called translation, which depends on the same interaction between RNA nucleotides. In alternative fashion, a cell may simply copy its genetic information in a process called DNA replication. The details of these functions are covered in other articles; here we focus on the interactions between DNA and other molecules that mediate the function of the genome.

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Genes and genomes

Further information: Cell nucleus, Chromatin, Chromosome, Gene, Noncoding DNA Genomic DNA is tightly and orderly packed in the process called DNA condensation to fit the small available volumes of the cell. In eukaryotes, DNA is located in the cell nucleus, as well as small amounts in mitochondria and chloroplasts. In prokaryotes, the DNA is held within an irregularly shaped body in the cytoplasm called the nucleoid.[72] The genetic information in a genome is held within genes, and the complete set of this information in an organism is called its genotype. A gene is a unit of heredity and is a region of DNA that influences a particular characteristic in an organism. Genes contain an open reading frame that can be transcribed, as well as regulatory sequences such as promoters and enhancers, which control the transcription of the open reading frame. In many species, only a small fraction of the total sequence of the genome encodes protein. For example, only about 1.5% of the human genome consists of protein-coding exons, with over 50% of human DNA consisting of non-coding repetitive sequences.[73] The reasons for the presence of so much noncoding DNA in eukaryotic genomes and the extraordinary differences in genome size, or C-value, among species represent a long-standing puzzle known as the "C-value enigma".[74] However, DNA sequences that do not code protein may still encode functional non-coding RNA molecules, which are involved in the regulation of gene expression.[75] Some noncoding DNA sequences play structural roles in chromosomes. Telomeres and centromeres typically contain few genes, but are important for the function and stability of chromosomes.[45] [77] An abundant form of noncoding DNA in humans are pseudogenes, which are copies of genes that have been disabled by mutation.[78] These sequences are usually just molecular fossils, although they can occasionally serve as raw genetic material for the creation of new genes through the process of gene duplication and divergence.[79]
T7 RNA polymerase (blue) producing a mRNA [76] (green) from a DNA template (orange).

Transcription and translation

Further information: Genetic code, Transcription (genetics), Protein

biosynthesis A gene is a sequence of DNA that contains genetic information and can influence the phenotype of an organism. Within a gene, the sequence of bases along a DNA strand defines a messenger RNA sequence, which then defines one or more protein sequences. The relationship between the nucleotide sequences of genes and the amino-acid sequences of proteins is determined by the rules of translation, known collectively as the genetic code. The genetic code consists of three-letter 'words' called codons formed from a sequence of three nucleotides (e.g. ACT, CAG, TTT). In transcription, the codons of a gene are copied into messenger RNA by RNA polymerase. This RNA copy is then decoded by a ribosome that reads the RNA sequence by base-pairing the messenger RNA to transfer RNA, which carries amino acids. Since there are 4 bases in 3-letter combinations, there are 64 possible codons ( combinations). These encode the twenty standard amino acids, giving most amino acids more than one possible codon. There are also three 'stop' or 'nonsense' codons signifying the end of the coding region; these are the TAA, TGA and TAG codons.

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Replication
Further information: DNA replication Cell division is essential for an organism to grow, but, when a cell divides, it must replicate the DNA in its genome so that the two daughter cells have the same genetic information as their parent. The double-stranded structure of DNA provides a simple mechanism for DNA DNA replication. The double helix is unwound by a helicase and topoisomerase. Next, replication. Here, the two strands are one DNA polymerase produces the leading strand copy. Another DNA polymerase binds to the lagging strand. This enzyme makes discontinuous segments (called Okazaki separated and then each strand's fragments) before DNA ligase joins them together. complementary DNA sequence is recreated by an enzyme called DNA polymerase. This enzyme makes the complementary strand by finding the correct base through complementary base pairing, and bonding it onto the original strand. As DNA polymerases can only extend a DNA strand in a 5 to 3 direction, different mechanisms are used to copy the antiparallel strands of the double helix.[80] In this way, the base on the old strand dictates which base appears on the new strand, and the cell ends up with a perfect copy of its DNA.

Interactions with proteins

All the functions of DNA depend on interactions with proteins. These protein interactions can be non-specific, or the protein can bind specifically to a single DNA sequence. Enzymes can also bind to DNA and of these, the polymerases that copy the DNA base sequence in transcription and DNA replication are particularly important.

DNA-binding proteins
Further information: DNA-binding protein

Interaction of DNA (shown in orange) with histones (shown in blue). These proteins' basic amino acids bind to the acidic phosphate groups on DNA. Structural proteins that bind DNA are well-understood examples of non-specific DNA-protein interactions. Within chromosomes, DNA is held in complexes with structural proteins. These proteins organize the DNA into a compact structure called chromatin. In eukaryotes this structure involves DNA binding to a complex of small basic proteins called histones, while in prokaryotes multiple types of proteins are involved.[81] [82] The histones form a disk-shaped complex called a nucleosome, which contains two complete turns of double-stranded DNA wrapped around its surface. These non-specific interactions are formed through basic residues in the histones making ionic bonds to the acidic sugar-phosphate backbone of the DNA, and are therefore largely independent of the base sequence.[83] Chemical modifications of these basic amino acid residues include methylation, phosphorylation and acetylation.[84]

DNA These chemical changes alter the strength of the interaction between the DNA and the histones, making the DNA more or less accessible to transcription factors and changing the rate of transcription.[85] Other non-specific DNA-binding proteins in chromatin include the high-mobility group proteins, which bind to bent or distorted DNA.[86] These proteins are important in bending arrays of nucleosomes and arranging them into the larger structures that make up chromosomes.[87] A distinct group of DNA-binding proteins are the DNA-binding proteins that specifically bind single-stranded DNA. In humans, replication protein A is the best-understood member of this family and is used in processes where the double helix is separated, including DNA replication, recombination and DNA repair.[88] These binding proteins seem to stabilize single-stranded DNA and protect it from forming stem-loops or being degraded by nucleases. In contrast, other proteins have evolved to bind to particular DNA sequences. The most intensively studied of these are the various transcription factors, which are proteins that regulate transcription. Each transcription factor binds to one particular set of DNA sequences and activates or inhibits the transcription of genes that have these sequences close to their promoters. The transcription factors do this in two ways. Firstly, they can bind the RNA polymerase responsible for transcription, either directly or through other mediator proteins; this locates the polymerase at the promoter and allows it to begin transcription.[90] Alternatively, transcription factors can bind enzymes that modify the histones at the promoter; this will change the accessibility of the DNA template to the polymerase.[91] As these DNA targets can occur throughout an organism's genome, changes in The lambda repressor the activity of one type of transcription factor can affect thousands of genes.[92] helix-turn-helix transcription factor Consequently, these proteins are often the targets of the signal transduction [89] bound to its DNA target processes that control responses to environmental changes or cellular differentiation and development. The specificity of these transcription factors' interactions with DNA come from the proteins making multiple contacts to the edges of the DNA bases, allowing them to "read" the DNA sequence. Most of these base-interactions are made in the major groove, where the bases are most accessible.[15]

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DNA-modifying enzymes
Nucleases and ligases Nucleases are enzymes that cut DNA strands by catalyzing the hydrolysis of the phosphodiester bonds. Nucleases that hydrolyse nucleotides from the ends of DNA strands are called exonucleases, while endonucleases cut within strands. The most frequently used nucleases in molecular biology are the restriction endonucleases, which The restriction enzyme EcoRV (green) in a [93] cut DNA at specific sequences. For instance, the EcoRV enzyme complex with its substrate DNA shown to the left recognizes the 6-base sequence 5-GAT|ATC-3 and makes a cut at the vertical line. In nature, these enzymes protect bacteria against phage infection by digesting the phage DNA when it enters the bacterial cell, acting as part of the restriction modification system.[94] In technology, these sequence-specific nucleases are used in molecular cloning and DNA fingerprinting. Enzymes called DNA ligases can rejoin cut or broken DNA strands.[95] Ligases are particularly important in lagging strand DNA replication, as they join together the short segments of DNA produced at the replication fork into a complete copy of the DNA template. They are also used in DNA repair and genetic recombination.[95]

DNA Topoisomerases and helicases Topoisomerases are enzymes with both nuclease and ligase activity. These proteins change the amount of supercoiling in DNA. Some of these enzymes work by cutting the DNA helix and allowing one section to rotate, thereby reducing its level of supercoiling; the enzyme then seals the DNA break.[27] Other types of these enzymes are capable of cutting one DNA helix and then passing a second strand of DNA through this break, before rejoining the helix.[96] Topoisomerases are required for many processes involving DNA, such as DNA replication and transcription.[28] Helicases are proteins that are a type of molecular motor. They use the chemical energy in nucleoside triphosphates, predominantly ATP, to break hydrogen bonds between bases and unwind the DNA double helix into single strands.[97] These enzymes are essential for most processes where enzymes need to access the DNA bases. Polymerases Polymerases are enzymes that synthesize polynucleotide chains from nucleoside triphosphates. The sequence of their products are copies of existing polynucleotide chains which are called templates. These enzymes function by adding nucleotides onto the 3 hydroxyl group of the previous nucleotide in a DNA strand. As a consequence, all polymerases work in a 5 to 3 direction.[98] In the active site of these enzymes, the incoming nucleoside triphosphate base-pairs to the template: this allows polymerases to accurately synthesize the complementary strand of their template. Polymerases are classified according to the type of template that they use. In DNA replication, a DNA-dependent DNA polymerase makes a copy of a DNA sequence. Accuracy is vital in this process, so many of these polymerases have a proofreading activity. Here, the polymerase recognizes the occasional mistakes in the synthesis reaction by the lack of base pairing between the mismatched nucleotides. If a mismatch is detected, a 3 to 5 exonuclease activity is activated and the incorrect base removed.[99] In most organisms, DNA polymerases function in a large complex called the replisome that contains multiple accessory subunits, such as the DNA clamp or helicases.[100] RNA-dependent DNA polymerases are a specialized class of polymerases that copy the sequence of an RNA strand into DNA. They include reverse transcriptase, which is a viral enzyme involved in the infection of cells by retroviruses, and telomerase, which is required for the replication of telomeres.[44] [101] Telomerase is an unusual polymerase because it contains its own RNA template as part of its structure.[45] Transcription is carried out by a DNA-dependent RNA polymerase that copies the sequence of a DNA strand into RNA. To begin transcribing a gene, the RNA polymerase binds to a sequence of DNA called a promoter and separates the DNA strands. It then copies the gene sequence into a messenger RNA transcript until it reaches a region of DNA called the terminator, where it halts and detaches from the DNA. As with human DNA-dependent DNA polymerases, RNA polymerase II, the enzyme that transcribes most of the genes in the human genome, operates as part of a large protein complex with multiple regulatory and accessory subunits.[102]

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DNA

68

Genetic recombination

Structure of the Holliday junction intermediate in genetic recombination. The four separate DNA strands are coloured red, blue, green and yellow.[103] Further information: Genetic recombination A DNA helix usually does not interact with other segments of DNA, and in human cells the different chromosomes even occupy separate areas in the nucleus called "chromosome territories".[104] This physical separation of different chromosomes is important for the ability of DNA to function as a stable repository for information, as one of the few times chromosomes interact is during chromosomal crossover when they recombine. Chromosomal crossover is when two DNA helices break, swap a section and then rejoin. Recombination allows chromosomes to exchange genetic information and produces new combinations of genes, which increases the efficiency of natural selection and can be important in the rapid evolution of new proteins.[105] Genetic recombination can also be involved in DNA repair, particularly in the cell's response to double-strand breaks.[106] The most common form of chromosomal crossover is homologous recombination, where the two chromosomes involved share very similar sequences. Non-homologous recombination can be damaging to cells, as it can produce chromosomal translocations and genetic abnormalities. The recombination reaction is catalyzed by enzymes known as recombinases, such as RAD51.[107] The first step in recombination is a double-stranded break either caused by an endonuclease or damage to the DNA.[108] A series of steps catalyzed in part by the recombinase then leads to joining of the two helices by at least one Holliday junction, in which a segment of a single strand in each helix is annealed to the complementary strand in the other helix. The Holliday junction is a tetrahedral junction structure that can be moved along the pair of chromosomes, swapping one strand for another. The recombination reaction is then halted by cleavage of the junction and re-ligation of the released DNA.[109]
Recombination involves the breakage and rejoining of two chromosomes (M and F) to produce two re-arranged chromosomes (C1 and C2).

DNA

69

Evolution
Further information: RNA world hypothesis DNA contains the genetic information that allows all modern living things to function, grow and reproduce. However, it is unclear how long in the 4-billion-year history of life DNA has performed this function, as it has been proposed that the earliest forms of life may have used RNA as their genetic material.[98] [110] RNA may have acted as the central part of early cell metabolism as it can both transmit genetic information and carry out catalysis as part of ribozymes.[111] This ancient RNA world where nucleic acid would have been used for both catalysis and genetics may have influenced the evolution of the current genetic code based on four nucleotide bases. This would occur, since the number of different bases in such an organism is a trade-off between a small number of bases increasing replication accuracy and a large number of bases increasing the catalytic efficiency of ribozymes.[112] However, there is no direct evidence of ancient genetic systems, as recovery of DNA from most fossils is impossible. This is because DNA will survive in the environment for less than one million years and slowly degrades into short fragments in solution.[113] Claims for older DNA have been made, most notably a report of the isolation of a viable bacterium from a salt crystal 250 million years old,[114] but these claims are controversial.[115] [116] On August 8, 2011, a report, based on NASA studies with meteorites found on Earth, was published suggesting building blocks of DNA (adenine, guanine and related organic molecules) may have been formed extraterrestrially in outer space.[117] [118] [119]

Uses in technology
Genetic engineering
Further information: Molecular biology, nucleic acid methods and genetic engineering Methods have been developed to purify DNA from organisms, such as phenol-chloroform extraction, and to manipulate it in the laboratory, such as restriction digests and the polymerase chain reaction. Modern biology and biochemistry make intensive use of these techniques in recombinant DNA technology. Recombinant DNA is a man-made DNA sequence that has been assembled from other DNA sequences. They can be transformed into organisms in the form of plasmids or in the appropriate format, by using a viral vector.[120] The genetically modified organisms produced can be used to produce products such as recombinant proteins, used in medical research,[121] or be grown in agriculture.[122] [123]

Forensics
Further information: DNA profiling Forensic scientists can use DNA in blood, semen, skin, saliva or hair found at a crime scene to identify a matching DNA of an individual, such as a perpetrator. This process is formally termed DNA profiling, but may also be called "genetic fingerprinting". In DNA profiling, the lengths of variable sections of repetitive DNA, such as short tandem repeats and minisatellites, are compared between people. This method is usually an extremely reliable technique for identifying a matching DNA.[124] However, identification can be complicated if the scene is contaminated with DNA from several people.[125] DNA profiling was developed in 1984 by British geneticist Sir Alec Jeffreys,[126] and first used in forensic science to convict Colin Pitchfork in the 1988 Enderby murders case.[127] The development of forensic science,and the ability to now obtain genetic matching on minute samples of blood, skin, saliva or hair has led to a re-examination of a number of cases. Evidence can now be uncovered that was not scientifically possible at the time of the original examination. Combined with the removal of the double jeopardy law, this allows cases to be reopened where previous trials have failed to produce sufficient evidence to convince a jury. People charged with serious crimes may be required to provide a sample of DNA for matching purposes. The most obvious defence to DNA matches obtained forensically is to claim that cross-contamination of evidence has

DNA taken place. This has resulted in meticulous strict handling procedures with new cases of serious crime. DNA profiling is also be used to identify victims of mass casualty incidents.[128] As well as positively identifying bodies or body parts in serious accidents, DNA profiling is being successfully used to identify individual victims in mass war graves - matching to family members.

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Bioinformatics
Further information: Bioinformatics Bioinformatics involves the manipulation, searching, and data mining of biological data, and this includes DNA sequence data. The development of techniques to store and search DNA sequences have led to widely applied advances in computer science, especially string searching algorithms, machine learning and database theory.[129] String searching or matching algorithms, which find an occurrence of a sequence of letters inside a larger sequence of letters, were developed to search for specific sequences of nucleotides.[130] The DNA sequenced may be aligned with other DNA sequences to identify homologous sequences and locate the specific mutations that make them distinct. These techniques, especially multiple sequence alignment, are used in studying phylogenetic relationships and protein function.[131] Data sets representing entire genomes' worth of DNA sequences, such as those produced by the Human Genome Project, are difficult to use without the annotations that identify the locations of genes and regulatory elements on each chromosome. Regions of DNA sequence that have the characteristic patterns associated with protein- or RNA-coding genes can be identified by gene finding algorithms, which allow researchers to predict the presence of particular gene products and their possible functions in an organism even before they have been isolated experimentally.[132] Entire genomes may also be compared which can shed light on the evolutionary history of particular organism and permit the examination of complex evolutionary events.

DNA nanotechnology
Further information: nanotechnology DNA

DNA nanotechnology uses the unique molecular recognition properties of DNA and other nucleic acids to create self-assembling branched DNA complexes with useful properties.[133] DNA is thus used as a structural material rather than as a carrier of biological information. This has led to the creation of two-dimensional The DNA structure at left (schematic shown) will self-assemble into the structure periodic lattices (both tile-based as visualized by atomic force microscopy at right. DNA nanotechnology is the field that well as using the "DNA origami" seeks to design nanoscale structures using the molecular recognition properties of DNA method) as well as three-dimensional molecules. Image from Strong, 2004 (doi:10.1371/journal.pbio.0020073). structures in the shapes of polyhedra.[134] Nanomechanical devices and algorithmic self-assembly have also been demonstrated,[135] and these DNA structures have been used to template the arrangement of other molecules such as gold nanoparticles and streptavidin proteins.[136]

History and anthropology

Further information: Phylogenetics and Genetic genealogy

DNA Because DNA collects mutations over time, which are then inherited, it contains historical information, and, by comparing DNA sequences, geneticists can infer the evolutionary history of organisms, their phylogeny.[137] This field of phylogenetics is a powerful tool in evolutionary biology. If DNA sequences within a species are compared, population geneticists can learn the history of particular populations. This can be used in studies ranging from ecological genetics to anthropology; For example, DNA evidence is being used to try to identify the Ten Lost Tribes of Israel.[138] [139] DNA has also been used to look at modern family relationships, such as establishing family relationships between the descendants of Sally Hemings and Thomas Jefferson. This usage is closely related to the use of DNA in criminal investigations detailed above. Indeed, some criminal investigations have been solved when DNA from crime scenes has matched relatives of the guilty individual.[140]

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History of DNA research

Further information: History of molecular biology DNA was first isolated by the Swiss physician Friedrich Miescher who, in 1869, discovered a microscopic substance in the pus of discarded surgical bandages. As it resided in the nuclei of cells, he called it "nuclein".[141] In 1878, Albrecht Kossel isolated the non-protein component of "nuclein", nucleic acid, and later isolated its five primary nucleobases.[142] In 1919, Phoebus Levene identified the base, sugar and phosphate nucleotide unit.[143] Levene suggested that DNA consisted of a string of nucleotide units linked together through the phosphate groups. However, Levene thought the chain was short and the bases repeated in a fixed order. In 1937 William Astbury produced the first X-ray diffraction patterns that showed that DNA had a regular structure.[144]

James D. Watson and Francis Crick (right), co-originators of the double-helix model, with Maclyn McCarty (left).

In 1927 Nikolai Koltsov proposed that inherited traits would be inherited via a "giant hereditary molecule" which would be made up of "two mirror strands that would replicate in a semi-conservative fashion using each strand as a template".[145] In 1928, Frederick Griffith discovered that traits of the "smooth" form of the Pneumococcus could be transferred to the "rough" form of the same bacteria by mixing killed "smooth" bacteria with the live "rough" form.[146] This system provided the first clear suggestion that DNA carries genetic informationthe AveryMacLeodMcCarty experimentwhen Oswald Avery, along with coworkers Colin MacLeod and Maclyn McCarty, identified DNA as the transforming principle in 1943.[147] DNA's role in heredity was confirmed in 1952, when Alfred Hershey and Martha Chase in the HersheyChase experiment showed that DNA is the genetic material of the T2 phage.[148] In 1953, James D. Watson and Francis Crick suggested what is now accepted as the first correct double-helix model of DNA structure in the journal Nature.[5] Their double-helix, molecular model of DNA was then based on a single X-ray diffraction image (labeled as "Photo 51")[149] taken by Rosalind Franklin and Raymond Gosling in May 1952, as well as the information that the DNA bases are paired also obtained through private communications from Erwin Chargaff in the previous years. Chargaff's rules played a very important role in establishing double-helix configurations for B-DNA as well as A-DNA. Experimental evidence supporting the Watson and Crick model were published in a series of five articles in the same issue of Nature.[150] Of these, Franklin and Gosling's paper was the first publication of their own X-ray diffraction data and original analysis method that partially supported the Watson and Crick model;[31] [151] this issue also contained an article on DNA structure by Maurice Wilkins and two of his colleagues, whose analysis and in vivo B-DNA X-ray patterns also supported the presence in vivo of the double-helical DNA configurations as proposed by

DNA Crick and Watson for their double-helix molecular model of DNA in the previous two pages of Nature.[32] In 1962, after Franklin's death, Watson, Crick, and Wilkins jointly received the Nobel Prize in Physiology or Medicine.[152] However, Nobel rules of the time allowed only living recipients, but a vigorous debate continues on who should receive credit for the discovery.[153] In an influential presentation in 1957, Crick laid out the central dogma of molecular biology, which foretold the relationship between DNA, RNA, and proteins, and articulated the "adaptor hypothesis".[154] Final confirmation of the replication mechanism that was implied by the double-helical structure followed in 1958 through the MeselsonStahl experiment.[155] Further work by Crick and coworkers showed that the genetic code was based on non-overlapping triplets of bases, called codons, allowing Har Gobind Khorana, Robert W. Holley and Marshall Warren Nirenberg to decipher the genetic code.[156] These findings represent the birth of molecular biology.

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References
[1] Russell, Peter (2001). iGenetics. New York: Benjamin Cummings. ISBN0-8053-4553-1. [2] Saenger, Wolfram (1984). Principles of Nucleic Acid Structure. New York: Springer-Verlag. ISBN0-387-90762-9. [3] Alberts, Bruce; Alexander Johnson, Julian Lewis, Martin Raff, Keith Roberts and Peter Walters (2002). Molecular Biology of the Cell; Fourth Edition (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?call=bv. View. . ShowTOC& rid=mboc4. TOC& depth=2). New York and London: Garland Science. ISBN0-8153-3218-1. OCLC145080076 48122761 57023651 69932405. . [4] Butler, John M. (2001). Forensic DNA Typing. Elsevier. ISBN978-0-12-147951-0. OCLC223032110 45406517. pp. 1415. [5] Watson J.D. and Crick F.H.C. (1953). "A Structure for Deoxyribose Nucleic Acid" (http:/ / www. nature. com/ nature/ dna50/ watsoncrick. pdf) (PDF). Nature 171 (4356): 737738. Bibcode1953Natur.171..737W. doi:10.1038/171737a0. PMID13054692. . [6] Mandelkern M, Elias J, Eden D, Crothers D (1981). "The dimensions of DNA in solution". J Mol Biol 152 (1): 15361. doi:10.1016/0022-2836(81)90099-1. PMID7338906. [7] Gregory S; Barlow, KF; McLay, KE; Kaul, R; Swarbreck, D; Dunham, A; Scott, CE; Howe, KL et al. (2006). "The DNA sequence and biological annotation of human chromosome 1". Nature 441 (7091): 31521. Bibcode2006Natur.441..315G. doi:10.1038/nature04727. PMID16710414. [8] Berg J., Tymoczko J. and Stryer L. (2002) Biochemistry. W. H. Freeman and Company ISBN 0-7167-4955-6 [9] Abbreviations and Symbols for Nucleic Acids, Polynucleotides and their Constituents (http:/ / www. chem. qmul. ac. uk/ iupac/ misc/ naabb. html) IUPAC-IUB Commission on Biochemical Nomenclature (CBN). Retrieved 03 January 2006. [10] Ghosh A, Bansal M (2003). "A glossary of DNA structures from A to Z". Acta Crystallogr D 59 (4): 6206. doi:10.1107/S0907444903003251. PMID12657780. [11] Created from PDB 1D65 (http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1D65) [12] Yakovchuk P, Protozanova E, Frank-Kamenetskii MD (2006). "Base-stacking and base-pairing contributions into thermal stability of the DNA double helix". Nucleic Acids Res. 34 (2): 56474. doi:10.1093/nar/gkj454. PMC1360284. PMID16449200. [13] Verma S, Eckstein F (1998). "Modified oligonucleotides: synthesis and strategy for users". Annu. Rev. Biochem. 67: 99134. doi:10.1146/annurev.biochem.67.1.99. PMID9759484. [14] Wing R, Drew H, Takano T, Broka C, Tanaka S, Itakura K, Dickerson R (1980). "Crystal structure analysis of a complete turn of B-DNA". Nature 287 (5784): 7558. Bibcode1980Natur.287..755W. doi:10.1038/287755a0. PMID7432492. [15] Pabo C, Sauer R (1984). "Protein-DNA recognition". Annu Rev Biochem 53: 293321. doi:10.1146/annurev.bi.53.070184.001453. PMID6236744. [16] Clausen-Schaumann H, Rief M, Tolksdorf C, Gaub H (2000). "Mechanical stability of single DNA molecules". Biophys J 78 (4): 19972007. Bibcode2000BpJ....78.1997C. doi:10.1016/S0006-3495(00)76747-6. PMC1300792. PMID10733978. [17] Chalikian T, Vlker J, Plum G, Breslauer K (1999). "A more unified picture for the thermodynamics of nucleic acid duplex melting: A characterization by calorimetric and volumetric techniques". Proc Natl Acad Sci USA 96 (14): 78538. Bibcode1999PNAS...96.7853C. doi:10.1073/pnas.96.14.7853. PMC22151. PMID10393911. [18] deHaseth P, Helmann J (1995). "Open complex formation by Escherichia coli RNA polymerase: the mechanism of polymerase-induced strand separation of double helical DNA". Mol Microbiol 16 (5): 81724. doi:10.1111/j.1365-2958.1995.tb02309.x. PMID7476180. [19] Isaksson J, Acharya S, Barman J, Cheruku P, Chattopadhyaya J (2004). "Single-stranded adenine-rich DNA and RNA retain structural characteristics of their respective double-stranded conformations and show directional differences in stacking pattern". Biochemistry 43 (51): 159966010. doi:10.1021/bi048221v. PMID15609994. [20] Designation of the two strands of DNA (http:/ / www. chem. qmul. ac. uk/ iubmb/ newsletter/ misc/ DNA. html) JCBN/NC-IUB Newsletter 1989, Accessed 07 May 2008 [21] Httenhofer A, Schattner P, Polacek N (2005). "Non-coding RNAs: hope or hype?". Trends Genet 21 (5): 28997. doi:10.1016/j.tig.2005.03.007. PMID15851066. [22] Munroe S (2004). "Diversity of antisense regulation in eukaryotes: multiple mechanisms, emerging patterns". J Cell Biochem 93 (4): 66471. doi:10.1002/jcb.20252. PMID15389973.

DNA
[23] Makalowska I, Lin C, Makalowski W (2005). "Overlapping genes in vertebrate genomes". Comput Biol Chem 29 (1): 112. doi:10.1016/j.compbiolchem.2004.12.006. PMID15680581. [24] Johnson Z, Chisholm S (2004). "Properties of overlapping genes are conserved across microbial genomes". Genome Res 14 (11): 226872. doi:10.1101/gr.2433104. PMC525685. PMID15520290. [25] Lamb R, Horvath C (1991). "Diversity of coding strategies in influenza viruses". Trends Genet 7 (8): 2616. PMID1771674. [26] Benham C, Mielke S (2005). "DNA mechanics". Annu Rev Biomed Eng 7: 2153. doi:10.1146/annurev.bioeng.6.062403.132016. PMID16004565. [27] Champoux J (2001). "DNA topoisomerases: structure, function, and mechanism". Annu Rev Biochem 70: 369413. doi:10.1146/annurev.biochem.70.1.369. PMID11395412. [28] Wang J (2002). "Cellular roles of DNA topoisomerases: a molecular perspective". Nat Rev Mol Cell Biol 3 (6): 43040. doi:10.1038/nrm831. PMID12042765. [29] Basu H, Feuerstein B, Zarling D, Shafer R, Marton L (1988). "Recognition of Z-RNA and Z-DNA determinants by polyamines in solution: experimental and theoretical studies". J Biomol Struct Dyn 6 (2): 299309. PMID2482766. [30] Franklin RE, Gosling RG (6 March 1953). "The Structure of Sodium Thymonucleate Fibres I. The Influence of Water Content" (http:/ / hekto. med. unc. edu:8080/ CARTER/ carter_WWW/ Bioch_134/ PDF_files/ Franklin_Gossling. pdf). Acta Crystallogr 6 (89): 6737. doi:10.1107/S0365110X53001939. . Franklin RE, Gosling RG (1953). "The structure of sodium thymonucleate fibres. II. The cylindrically symmetrical Patterson function". Acta Crystallogr 6 (89): 67885. doi:10.1107/S0365110X53001940. [31] Franklin, Rosalind and Gosling, Raymond (1953). "Molecular Configuration in Sodium Thymonucleate. Franklin R. and Gosling R.G" (http:/ / www. nature. com/ nature/ dna50/ franklingosling. pdf) (PDF). Nature 171 (4356): 7401. Bibcode1953Natur.171..740F. doi:10.1038/171740a0. PMID13054694. . [32] Wilkins M.H.F., A.R. Stokes A.R. & Wilson, H.R. (1953). "Molecular Structure of Deoxypentose Nucleic Acids" (http:/ / www. nature. com/ nature/ dna50/ wilkins. pdf) (PDF). Nature 171 (4356): 738740. Bibcode1953Natur.171..738W. doi:10.1038/171738a0. PMID13054693. . [33] Leslie AG, Arnott S, Chandrasekaran R, Ratliff RL (1980). "Polymorphism of DNA double helices". J. Mol. Biol. 143 (1): 4972. doi:10.1016/0022-2836(80)90124-2. PMID7441761. [34] Baianu, I.C. (1980). "Structural Order and Partial Disorder in Biological systems". Bull. Math. Biol. 42 (4): 137141. http:/ / cogprints. org/ 3822/ [35] Hosemann R., Bagchi R.N., Direct analysis of diffraction by matter, North-Holland Publs., Amsterdam New York, 1962. [36] Baianu, I.C. (1978). "X-ray scattering by partially disordered membrane systems". Acta Crystallogr A 34 (5): 751753. Bibcode1978AcCrA..34..751B. doi:10.1107/S0567739478001540. [37] Wahl M, Sundaralingam M (1997). "Crystal structures of A-DNA duplexes". Biopolymers 44 (1): 4563. doi:10.1002/(SICI)1097-0282(1997)44:1<45::AID-BIP4>3.0.CO;2-#. PMID9097733. [38] Lu XJ, Shakked Z, Olson WK (2000). "A-form conformational motifs in ligand-bound DNA structures". J. Mol. Biol. 300 (4): 81940. doi:10.1006/jmbi.2000.3690. PMID10891271. [39] Rothenburg S, Koch-Nolte F, Haag F (2001). "DNA methylation and Z-DNA formation as mediators of quantitative differences in the expression of alleles". Immunol Rev 184: 28698. doi:10.1034/j.1600-065x.2001.1840125.x. PMID12086319. [40] Oh D, Kim Y, Rich A (2002). "Z-DNA-binding proteins can act as potent effectors of gene expression in vivo". Proc. Natl. Acad. Sci. U.S.A. 99 (26): 1666671. Bibcode2002PNAS...9916666O. doi:10.1073/pnas.262672699. PMC139201. PMID12486233. [41] "Arsenic-loving bacteria may help in hunt for alien life" (http:/ / www. bbc. co. uk/ news/ science-environment-11886943). BBC News. December 2, 2010. . Retrieved 2010-12-02. [42] Bortman, Henry (2010-12-02). "Arsenic-Eating Bacteria Opens New Possibilities for Alien Life" (http:/ / www. space. com/ scienceastronomy/ arsenic-bacteria-alien-life-101202. html). Space.Com web site (http:/ / www. space. com/ ) (Space.com). . Retrieved 2010-12-02. [43] "Arsenic-eating microbe may redefine chemistry of life" (http:/ / www. nature. com/ news/ 2010/ 101202/ full/ news. 2010. 645. html). Nature News. 2 December 2010. . Retrieved 2010-12-02. [44] Greider C, Blackburn E (1985). "Identification of a specific telomere terminal transferase activity in Tetrahymena extracts". Cell 43 (2 Pt 1): 40513. doi:10.1016/0092-8674(85)90170-9. PMID3907856. [45] Nugent C, Lundblad V (1998). "The telomerase reverse transcriptase: components and regulation". Genes Dev 12 (8): 107385. doi:10.1101/gad.12.8.1073. PMID9553037. [46] Wright W, Tesmer V, Huffman K, Levene S, Shay J (1997). "Normal human chromosomes have long G-rich telomeric overhangs at one end". Genes Dev 11 (21): 28019. doi:10.1101/gad.11.21.2801. PMC316649. PMID9353250. [47] Created from NDB UD0017 (http:/ / ndbserver. rutgers. edu/ atlas/ xray/ structures/ U/ ud0017/ ud0017. html) [48] Burge S, Parkinson G, Hazel P, Todd A, Neidle S (2006). "Quadruplex DNA: sequence, topology and structure". Nucleic Acids Res 34 (19): 540215. doi:10.1093/nar/gkl655. PMC1636468. PMID17012276. [49] Parkinson G, Lee M, Neidle S (2002). "Crystal structure of parallel quadruplexes from human telomeric DNA". Nature 417 (6891): 87680. doi:10.1038/nature755. PMID12050675. [50] Griffith J, Comeau L, Rosenfield S, Stansel R, Bianchi A, Moss H, de Lange T (1999). "Mammalian telomeres end in a large duplex loop". Cell 97 (4): 50314. doi:10.1016/S0092-8674(00)80760-6. PMID10338214.

73

DNA
[51] Seeman NC (2005). "DNA enables nanoscale control of the structure of matter". Q. Rev. Biophys. 38 (4): 36371. doi:10.1017/S0033583505004087. PMID16515737. [52] Painter PC, Mosher LE, Rhoads C (1982). "Low-frequency modes in the Raman spectra of proteins". Biopolymers 21 (7): 146972. doi:10.1002/bip.360210715. PMID7115900. [53] Urabe H, Tominaga Y, Kubota K (1983). "Experimental evidence of collective vibrations in DNA double helix (Raman spectroscopy)". Journal of Chemical Physics 78 (10): 59375939. Bibcode1983JChPh..78.5937U. doi:10.1063/1.444600. [54] Chou KC (1984). "Low-frequency vibrations of DNA molecules". Biochem. J. 221 (1): 2731. PMC1143999. PMID6466317. [55] Chou KC, Maggiora GM, Mao B (1989). "Quasi-continuum models of twist-like and accordion-like low-frequency motions in DNA". Biophys. J. 56 (2): 295305. Bibcode1989BpJ....56..295C. doi:10.1016/S0006-3495(89)82676-1. PMC1280479. PMID2775828. [56] Klose R, Bird A (2006). "Genomic DNA methylation: the mark and its mediators". Trends Biochem Sci 31 (2): 8997. doi:10.1016/j.tibs.2005.12.008. PMID16403636. [57] Bird A (2002). "DNA methylation patterns and epigenetic memory". Genes Dev 16 (1): 621. doi:10.1101/gad.947102. PMID11782440. [58] Walsh C, Xu G (2006). "Cytosine methylation and DNA repair". Curr Top Microbiol Immunol. Current Topics in Microbiology and Immunology 301: 283315. doi:10.1007/3-540-31390-7_11. ISBN3-540-29114-8. PMID16570853. [59] Kriaucionis S, Heintz N (2009). "The nuclear DNA base 5-hydroxymethylcytosine is present in Purkinje neurons and the brain". Science 324 (5929): 92930. Bibcode2009Sci...324..929K. doi:10.1126/science.1169786. PMID19372393. [60] Ratel D, Ravanat J, Berger F, Wion D (2006). "N6-methyladenine: the other methylated base of DNA". Bioessays 28 (3): 30915. doi:10.1002/bies.20342. PMC2754416. PMID16479578. [61] Gommers-Ampt J, Van Leeuwen F, de Beer A, Vliegenthart J, Dizdaroglu M, Kowalak J, Crain P, Borst P (1993). "beta-D-glucosyl-hydroxymethyluracil: a novel modified base present in the DNA of the parasitic protozoan T. brucei". Cell 75 (6): 112936. doi:10.1016/0092-8674(93)90322-H. PMID8261512. [62] Created from PDB 1JDG (http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1JDG) [63] Douki T, Reynaud-Angelin A, Cadet J, Sage E (2003). "Bipyrimidine photoproducts rather than oxidative lesions are the main type of DNA damage involved in the genotoxic effect of solar UVA radiation". Biochemistry 42 (30): 92216. doi:10.1021/bi034593c. PMID12885257. [64] Cadet J, Delatour T, Douki T, Gasparutto D, Pouget J, Ravanat J, Sauvaigo S (1999). "Hydroxyl radicals and DNA base damage". Mutat Res 424 (12): 921. doi:10.1016/S0027-5107(99)00004-4. PMID10064846. [65] Beckman KB, Ames BN (1997). "Oxidative decay of DNA". J. Biol. Chem. 272 (32): 196336. doi:10.1074/jbc.272.32.19633. PMID9289489. [66] Valerie K, Povirk L (2003). "Regulation and mechanisms of mammalian double-strand break repair". Oncogene 22 (37): 5792812. doi:10.1038/sj.onc.1206679. PMID12947387. [67] Ferguson L, Denny W (1991). "The genetic toxicology of acridines". Mutat Res 258 (2): 12360. PMID1881402. [68] Stephens T, Bunde C, Fillmore B (2000). "Mechanism of action in thalidomide teratogenesis". Biochem Pharmacol 59 (12): 148999. doi:10.1016/S0006-2952(99)00388-3. PMID10799645. [69] Jeffrey A (1985). "DNA modification by chemical carcinogens". Pharmacol Ther 28 (2): 23772. doi:10.1016/0163-7258(85)90013-0. PMID3936066. [70] Braa M, Cacho M, Gradillas A, de Pascual-Teresa B, Ramos A (2001). "Intercalators as anticancer drugs". Curr Pharm Des 7 (17): 174580. doi:10.2174/1381612013397113. PMID11562309. [71] Venter J; Adams, MD; Myers, EW; Li, PW; Mural, RJ; Sutton, GG; Smith, HO; Yandell, M et al. (2001). "The sequence of the human genome". Science 291 (5507): 130451. Bibcode2001Sci...291.1304V. doi:10.1126/science.1058040. PMID11181995. [72] Thanbichler M, Wang S, Shapiro L (2005). "The bacterial nucleoid: a highly organized and dynamic structure". J Cell Biochem 96 (3): 50621. doi:10.1002/jcb.20519. PMID15988757. [73] Wolfsberg T, McEntyre J, Schuler G (2001). "Guide to the draft human genome". Nature 409 (6822): 8246. doi:10.1038/35057000. PMID11236998. [74] Gregory T (2005). "The C-value enigma in plants and animals: a review of parallels and an appeal for partnership". Ann Bot (Lond) 95 (1): 13346. doi:10.1093/aob/mci009. PMID15596463. [75] The ENCODE Project Consortium (2007). "Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project". Nature 447 (7146): 799816. Bibcode2007Natur.447..799B. doi:10.1038/nature05874. PMC2212820. PMID17571346. [76] Created from PDB 1MSW (http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1MSW) [77] Pidoux A, Allshire R (2005). "The role of heterochromatin in centromere function". Philos Trans R Soc Lond B Biol Sci 360 (1455): 56979. doi:10.1098/rstb.2004.1611. PMC1569473. PMID15905142. [78] Harrison P, Hegyi H, Balasubramanian S, Luscombe N, Bertone P, Echols N, Johnson T, Gerstein M (2002). "Molecular Fossils in the Human Genome: Identification and Analysis of the Pseudogenes in Chromosomes 21 and 22". Genome Res 12 (2): 27280. doi:10.1101/gr.207102. PMC155275. PMID11827946. [79] Harrison P, Gerstein M (2002). "Studying genomes through the aeons: protein families, pseudogenes and proteome evolution". J Mol Biol 318 (5): 115574. doi:10.1016/S0022-2836(02)00109-2. PMID12083509. [80] Alb M (2001). "Replicative DNA polymerases". Genome Biol 2 (1): REVIEWS3002. doi:10.1186/gb-2001-2-1-reviews3002. PMC150442. PMID11178285. [81] Sandman K, Pereira S, Reeve J (1998). "Diversity of prokaryotic chromosomal proteins and the origin of the nucleosome". Cell Mol Life Sci 54 (12): 135064. doi:10.1007/s000180050259. PMID9893710.

74

DNA
[82] Dame RT (2005). "The role of nucleoid-associated proteins in the organization and compaction of bacterial chromatin". Mol. Microbiol. 56 (4): 85870. doi:10.1111/j.1365-2958.2005.04598.x. PMID15853876. [83] Luger K, Mder A, Richmond R, Sargent D, Richmond T (1997). "Crystal structure of the nucleosome core particle at 2.8 A resolution". Nature 389 (6648): 25160. Bibcode1997Natur.389..251L. doi:10.1038/38444. PMID9305837. [84] Jenuwein T, Allis C (2001). "Translating the histone code". Science 293 (5532): 107480. doi:10.1126/science.1063127. PMID11498575. [85] Ito T (2003). "Nucleosome assembly and remodelling". Curr Top Microbiol Immunol. Current Topics in Microbiology and Immunology 274: 122. doi:10.1007/978-3-642-55747-7_1. ISBN978-3-540-44208-0. PMID12596902. [86] Thomas J (2001). "HMG1 and 2: architectural DNA-binding proteins". Biochem Soc Trans 29 (Pt 4): 395401. doi:10.1042/BST0290395. PMID11497996. [87] Grosschedl R, Giese K, Pagel J (1994). "HMG domain proteins: architectural elements in the assembly of nucleoprotein structures". Trends Genet 10 (3): 94100. doi:10.1016/0168-9525(94)90232-1. PMID8178371. [88] Iftode C, Daniely Y, Borowiec J (1999). "Replication protein A (RPA): the eukaryotic SSB". Crit Rev Biochem Mol Biol 34 (3): 14180. doi:10.1080/10409239991209255. PMID10473346. [89] Created from PDB 1LMB (http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1LMB) [90] Myers L, Kornberg R (2000). "Mediator of transcriptional regulation". Annu Rev Biochem 69: 72949. doi:10.1146/annurev.biochem.69.1.729. PMID10966474. [91] Spiegelman B, Heinrich R (2004). "Biological control through regulated transcriptional coactivators". Cell 119 (2): 15767. doi:10.1016/j.cell.2004.09.037. PMID15479634. [92] Li Z, Van Calcar S, Qu C, Cavenee W, Zhang M, Ren B (2003). "A global transcriptional regulatory role for c-Myc in Burkitt's lymphoma cells". Proc Natl Acad Sci USA 100 (14): 81649. Bibcode2003PNAS..100.8164L. doi:10.1073/pnas.1332764100. PMC166200. PMID12808131. [93] Created from PDB 1RVA (http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1RVA) [94] Bickle T, Krger D (1993). "Biology of DNA restriction". Microbiol Rev 57 (2): 43450. PMC372918. PMID8336674. [95] Doherty A, Suh S (2000). "Structural and mechanistic conservation in DNA ligases". Nucleic Acids Res 28 (21): 40518. doi:10.1093/nar/28.21.4051. PMC113121. PMID11058099. [96] Schoeffler A, Berger J (2005). "Recent advances in understanding structure-function relationships in the type II topoisomerase mechanism". Biochem Soc Trans 33 (Pt 6): 146570. doi:10.1042/BST20051465. PMID16246147. [97] Tuteja N, Tuteja R (2004). "Unraveling DNA helicases. Motif, structure, mechanism and function". Eur J Biochem 271 (10): 184963. doi:10.1111/j.1432-1033.2004.04094.x. PMID15128295. [98] Joyce C, Steitz T (1995). "Polymerase structures and function: variations on a theme?". J Bacteriol 177 (22): 63219. PMC177480. PMID7592405. [99] Hubscher U, Maga G, Spadari S (2002). "Eukaryotic DNA polymerases". Annu Rev Biochem 71: 13363. doi:10.1146/annurev.biochem.71.090501.150041. PMID12045093. [100] Johnson A, O'Donnell M (2005). "Cellular DNA replicases: components and dynamics at the replication fork". Annu Rev Biochem 74: 283315. doi:10.1146/annurev.biochem.73.011303.073859. PMID15952889. [101] Tarrago-Litvak L, Androla M, Nevinsky G, Sarih-Cottin L, Litvak S (1 May 1994). "The reverse transcriptase of HIV-1: from enzymology to therapeutic intervention". FASEB J 8 (8): 497503. PMID7514143. [102] Martinez E (2002). "Multi-protein complexes in eukaryotic gene transcription". Plant Mol Biol 50 (6): 92547. doi:10.1023/A:1021258713850. PMID12516863. [103] Created from PDB 1M6G (http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1M6G) [104] Cremer T, Cremer C (2001). "Chromosome territories, nuclear architecture and gene regulation in mammalian cells". Nat Rev Genet 2 (4): 292301. doi:10.1038/35066075. PMID11283701. [105] Pl C, Papp B, Lercher M (2006). "An integrated view of protein evolution". Nat Rev Genet 7 (5): 33748. doi:10.1038/nrg1838. PMID16619049. [106] O'Driscoll M, Jeggo P (2006). "The role of double-strand break repair insights from human genetics". Nat Rev Genet 7 (1): 4554. doi:10.1038/nrg1746. PMID16369571. [107] Visp S, Defais M (1997). "Mammalian Rad51 protein: a RecA homologue with pleiotropic functions". Biochimie 79 (910): 58792. doi:10.1016/S0300-9084(97)82007-X. PMID9466696. [108] Neale MJ, Keeney S (2006). "Clarifying the mechanics of DNA strand exchange in meiotic recombination". Nature 442 (7099): 1538. Bibcode2006Natur.442..153N. doi:10.1038/nature04885. PMID16838012. [109] Dickman M, Ingleston S, Sedelnikova S, Rafferty J, Lloyd R, Grasby J, Hornby D (2002). "The RuvABC resolvasome". Eur J Biochem 269 (22): 5492501. doi:10.1046/j.1432-1033.2002.03250.x. PMID12423347. [110] Orgel L (2004). "Prebiotic chemistry and the origin of the RNA world". Crit Rev Biochem Mol Biol 39 (2): 99123. doi:10.1080/10409230490460765. PMID15217990. [111] Davenport R (2001). "Ribozymes. Making copies in the RNA world". Science 292 (5520): 1278. doi:10.1126/science.292.5520.1278a. PMID11360970. [112] Szathmry E (1992). "What is the optimum size for the genetic alphabet?". Proc Natl Acad Sci USA 89 (7): 26148. Bibcode1992PNAS...89.2614S. doi:10.1073/pnas.89.7.2614. PMC48712. PMID1372984.

75

DNA
[113] Lindahl T (1993). "Instability and decay of the primary structure of DNA". Nature 362 (6422): 70915. Bibcode1993Natur.362..709L. doi:10.1038/362709a0. PMID8469282. [114] Vreeland R, Rosenzweig W, Powers D (2000). "Isolation of a 250 million-year-old halotolerant bacterium from a primary salt crystal". Nature 407 (6806): 897900. doi:10.1038/35038060. PMID11057666. [115] Hebsgaard M, Phillips M, Willerslev E (2005). "Geologically ancient DNA: fact or artefact?". Trends Microbiol 13 (5): 21220. doi:10.1016/j.tim.2005.03.010. PMID15866038. [116] Nickle D, Learn G, Rain M, Mullins J, Mittler J (2002). "Curiously modern DNA for a "250 million-year-old" bacterium". J Mol Evol 54 (1): 1347. doi:10.1007/s00239-001-0025-x. PMID11734907. [117] Callahan, M.P.; Smith, K.E.; Cleaves, H.J.; Ruzica, J.; Stern, J.C.; Glavin, D.P.; House, C.H.; Dworkin, J.P. (11 August 2011). "Carbonaceous meteorites contain a wide range of extraterrestrial nucleobases" (http:/ / www. pnas. org/ content/ early/ 2011/ 08/ 10/ 1106493108). PNAS. doi:10.1073/pnas.1106493108. . Retrieved 2011-08-15. [118] Steigerwald, John (08 August 2011). "NASA Researchers: DNA Building Blocks Can Be Made in Space" (http:/ / www. nasa. gov/ topics/ solarsystem/ features/ dna-meteorites. html). NASA. . Retrieved 2011-08-10. [119] ScienceDaily Staff (09 August 2011). "DNA Building Blocks Can Be Made in Space, NASA Evidence Suggests" (http:/ / www. sciencedaily. com/ releases/ 2011/ 08/ 110808220659. htm). ScienceDaily. . Retrieved 2011-08-09. [120] Goff SP, Berg P (1976). "Construction of hybrid viruses containing SV40 and lambda phage DNA segments and their propagation in cultured monkey cells". Cell 9 (4 PT 2): 695705. doi:10.1016/0092-8674(76)90133-1. PMID189942. [121] Houdebine L (2007). "Transgenic animal models in biomedical research". Methods Mol Biol 360: 163202. doi:10.1385/1-59745-165-7:163. ISBN1-59745-165-7. PMID17172731. [122] Daniell H, Dhingra A (2002). "Multigene engineering: dawn of an exciting new era in biotechnology". Curr Opin Biotechnol 13 (2): 13641. doi:10.1016/S0958-1669(02)00297-5. PMID11950565. [123] Job D (2002). "Plant biotechnology in agriculture". Biochimie 84 (11): 110510. doi:10.1016/S0300-9084(02)00013-5. PMID12595138. [124] Collins A, Morton N (1994). "Likelihood ratios for DNA identification". Proc Natl Acad Sci USA 91 (13): 600711. Bibcode1994PNAS...91.6007C. doi:10.1073/pnas.91.13.6007. PMC44126. PMID8016106. [125] Weir B, Triggs C, Starling L, Stowell L, Walsh K, Buckleton J (1997). "Interpreting DNA mixtures". J Forensic Sci 42 (2): 21322. PMID9068179. [126] Jeffreys A, Wilson V, Thein S (1985). "Individual-specific 'fingerprints' of human DNA". Nature 316 (6023): 769. Bibcode1985Natur.316...76J. doi:10.1038/316076a0. PMID2989708. [127] Colin Pitchfork first murder conviction on DNA evidence also clears the prime suspect (http:/ / web. archive. org/ web/ 20061214004903/ http:/ / www. forensic. gov. uk/ forensic_t/ inside/ news/ list_casefiles. php?case=1) Forensic Science Service Accessed 23 December 2006 [128] "DNA Identification in Mass Fatality Incidents" (http:/ / massfatality. dna. gov/ Introduction/ ). National Institute of Justice. September 2006. . [129] Baldi, Pierre; Brunak, Soren (2001). Bioinformatics: The Machine Learning Approach. MIT Press. ISBN978-0-262-02506-5. OCLC45951728. [130] Gusfield, Dan. Algorithms on Strings, Trees, and Sequences: Computer Science and Computational Biology. Cambridge University Press, 15 January 1997. ISBN 978-0-521-58519-4. [131] Sjlander K (2004). "Phylogenomic inference of protein molecular function: advances and challenges". Bioinformatics 20 (2): 1709. doi:10.1093/bioinformatics/bth021. PMID14734307. [132] Mount DM (2004). Bioinformatics: Sequence and Genome Analysis (2 ed.). Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press. ISBN0-87969-712-1. OCLC55106399. [133] Rothemund PW (2006). "Folding DNA to create nanoscale shapes and patterns". Nature 440 (7082): 297302. Bibcode2006Natur.440..297R. doi:10.1038/nature04586. PMID16541064. [134] Andersen ES, Dong M, Nielsen MM (2009). "Self-assembly of a nanoscale DNA box with a controllable lid". Nature 459 (7243): 736. Bibcode2009Natur.459...73A. doi:10.1038/nature07971. PMID19424153. [135] Ishitsuka Y, Ha T (2009). "DNA nanotechnology: a nanomachine goes live". Nat Nanotechnol 4 (5): 2812. Bibcode2009NatNa...4..281I. doi:10.1038/nnano.2009.101. PMID19421208. [136] Aldaye FA, Palmer AL, Sleiman HF (2008). "Assembling materials with DNA as the guide". Science 321 (5897): 17959. Bibcode2008Sci...321.1795A. doi:10.1126/science.1154533. PMID18818351. [137] Wray G; Martindale, Mark Q. (2002). "Dating branches on the Tree of Life using DNA". Genome Biol 3 (1): REVIEWS0001. doi:10.1046/j.1525-142X.1999.99010.x. PMC150454. PMID11806830. [138] Lost Tribes of Israel, NOVA, PBS airdate: 22 February 2000. Transcript available from PBS.org (http:/ / www. pbs. org/ wgbh/ nova/ transcripts/ 2706israel. html). Retrieved 4 March 2006. [139] Kleiman, Yaakov. "The Cohanim/DNA Connection: The fascinating story of how DNA studies confirm an ancient biblical tradition". (http:/ / www. aish. com/ societywork/ sciencenature/ the_cohanim_-_dna_connection. asp) aish.com (January 13, 2000). Retrieved 4 March 2006. [140] Bhattacharya, Shaoni. "Killer convicted thanks to relative's DNA". (http:/ / www. newscientist. com/ article. ns?id=dn4908) newscientist.com (20 April 2004). Retrieved 22 December 06.

76

DNA
[141] Dahm R (2008). "Discovering DNA: Friedrich Miescher and the early years of nucleic acid research". Hum. Genet. 122 (6): 56581. doi:10.1007/s00439-007-0433-0. PMID17901982. [142] Jones, Mary Ellen (September 1953). "Albrecht Kossel, A Biographical Sketch". Yale Journal of Biology and Medicine (National Center for Biotechnology Information) 26 (1): 8097. PMC2599350. PMID13103145. [143] Levene P, (1 December 1919). "The structure of yeast nucleic acid". J Biol Chem 40 (2): 41524. [144] Astbury W, (1947). "Nucleic acid". Symp. SOC. Exp. Biol. 1 (66). [145] Valery N. Soyfer (2001). "The consequences of political dictatorship for Russian science". Nature Reviews Genetics 2 (9): 723729. doi:10.1038/35088598. PMID11533721. [146] Lorenz MG, Wackernagel W (1994). "Bacterial gene transfer by natural genetic transformation in the environment". Microbiol. Rev. 58 (3): 563602. PMC372978. PMID7968924. [147] Avery O, MacLeod C, McCarty M (1944). "STUDIES ON THE CHEMICAL NATURE OF THE SUBSTANCE INDUCING TRANSFORMATION OF PNEUMOCOCCAL TYPES : INDUCTION OF TRANSFORMATION BY A DESOXYRIBONUCLEIC ACID FRACTION ISOLATED FROM PNEUMOCOCCUS TYPE III". J Exp Med 79 (2): 137158. doi:10.1084/jem.79.2.137. PMC2135445. PMID19871359. [148] Hershey A, Chase M (1952). "INDEPENDENT FUNCTIONS OF VIRAL PROTEIN AND NUCLEIC ACID IN GROWTH OF BACTERIOPHAGE". J Gen Physiol 36 (1): 3956. doi:10.1085/jgp.36.1.39. PMC2147348. PMID12981234. [149] The B-DNA X-ray pattern on the right of this linked image (http:/ / osulibrary. oregonstate. edu/ specialcollections/ coll/ pauling/ dna/ pictures/ sci9. 001. 5. html) was obtained by Rosalind Franklin and Raymond Gosling in May 1952 at high hydration levels of DNA and it has been labeled as "Photo 51" [150] Nature Archives Double Helix of DNA: 50 Years (http:/ / www. nature. com/ nature/ dna50/ archive. html) [151] "Original X-ray diffraction image" (http:/ / osulibrary. oregonstate. edu/ specialcollections/ coll/ pauling/ dna/ pictures/ franklin-typeBphoto. html). Osulibrary.oregonstate.edu. . Retrieved 2011-02-06. [152] The Nobel Prize in Physiology or Medicine 1962 (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1962/ ) Nobelprize .org Accessed 22 December 06 [153] Brenda Maddox (23 January 2003). "The double helix and the 'wronged heroine'" (http:/ / www. biomath. nyu. edu/ index/ course/ hw_articles/ nature4. pdf) (PDF). Nature 421 (6921): 407408. doi:10.1038/nature01399. PMID12540909. . [154] Crick, F.H.C. On degenerate templates and the adaptor hypothesis (PDF). (http:/ / genome. wellcome. ac. uk/ assets/ wtx030893. pdf) genome.wellcome.ac.uk (Lecture, 1955). Retrieved 22 December 2006. [155] Meselson M, Stahl F (1958). "THE REPLICATION OF DNA IN ESCHERICHIA COLI". Proc Natl Acad Sci USA 44 (7): 67182. Bibcode1958PNAS...44..671M. doi:10.1073/pnas.44.7.671. PMC528642. PMID16590258. [156] The Nobel Prize in Physiology or Medicine 1968 (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1968/ ) Nobelprize.org Accessed 22 December 06

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Further reading
Berry, Andrew; Watson, James D. (2003). DNA: the secret of life. New York: Alfred A. Knopf. ISBN0-375-41546-7. Calladine, Chris R.; Drew, Horace R.; Luisi, Ben F. and Travers, Andrew A. (2003). Understanding DNA: the molecule & how it works. Amsterdam: Elsevier Academic Press. ISBN0-12-155089-3. Dennis, Carina; Julie Clayton (2003). 50 years of DNA. Basingstoke: Palgrave Macmillan. ISBN1-4039-1479-6. Judson, Horace F. 1979. The Eighth Day of Creation: Makers of the Revolution in Biology. Touchstone Books, ISBN 0-671-22540-5. 2nd edition: Cold Spring Harbor Laboratory Press, 1996 paperback: ISBN 0-87969-478-5. Olby, Robert C. (1994). The path to the double helix: the discovery of DNA. New York: Dover Publications. ISBN0-486-68117-3., first published in October 1974 by MacMillan, with foreword by Francis Crick;the definitive DNA textbook,revised in 1994 with a 9 page postscript Micklas, David. 2003. DNA Science: A First Course. Cold Spring Harbor Press: ISBN 978-0879696368 Ridley, Matt (2006). Francis Crick: discoverer of the genetic code. Ashland, OH: Eminent Lives, Atlas Books. ISBN0-06-082333-X. Olby, Robert C. (2009). Francis Crick: A Biography. Plainview, N.Y: Cold Spring Harbor Laboratory Press. ISBN0-87969-798-9. Rosenfeld, Israel. 2010. DNA: A Graphic Guide to the Molecule that Shook the World. Columbia University Press: ISBN 978-0231142717 Schultz, Mark and Zander Cannon. 2009. The Stuff of Life: A Graphic Guide to Genetics and DNA. Hill and Wang: ISBN 0809089475

DNA Stent, Gunther Siegmund; Watson, James D. (1980). The double helix: a personal account of the discovery of the structure of DNA. New York: Norton. ISBN0-393-95075-1. Watson, James D. 2004. DNA: The Secret of Life. Random House: ISBN 978-0099451846 Wilkins, Maurice (2003). The third man of the double helix the autobiography of Maurice Wilkins. Cambridge, Eng: University Press. ISBN0-19-860665-6.

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External links
DNA (http://www.dmoz.org/Science/Biology/Biochemistry_and_Molecular_Biology/Biomolecules/ Nucleic_Acids/DNA//) at the Open Directory Project DNA binding site prediction on protein (http://pipe.scs.fsu.edu/displar.html) DNA coiling to form chromosomes (http://biostudio.com/c_ education mac.htm) DNA the Double Helix Game (http://nobelprize.org/educational_games/medicine/dna_double_helix/) From the official Nobel Prize web site DNA under electron microscope (http://www.fidelitysystems.com/Unlinked_DNA.html) Dolan DNA Learning Center (http://www.dnalc.org/) Double Helix: 50 years of DNA (http://www.nature.com/nature/dna50/archive.html), Nature Proteopedia DNA (http://www.proteopedia.org/wiki/index.php/DNA) Double Helix 19532003 (http://www.ncbe.reading.ac.uk/DNA50/) National Centre for Biotechnology Education Francis Crick and James Watson talking on the BBC in 1962, 1972, and 1974 (http://www.bbc.co.uk/bbcfour/ audiointerviews/profilepages/crickwatson1.shtml) Genetic Education Modules for Teachers (http://www.genome.gov/10506718)DNA from the Beginning Study Guide Guide to DNA cloning (http://www.blackwellpublishing.com/trun/artwork/Animations/cloningexp/ cloningexp.html) Olby R (2003). "Quiet debut for the double helix" (http://chem-faculty.ucsd.edu/joseph/CHEM13/DNA1. pdf). Nature 421 (6921): 4025. doi:10.1038/nature01397. PMID12540907. DNA from the Beginning (http://www.dnaftb.org/dnaftb/) Another DNA Learning Center site on DNA, genes, and heredity from Mendel to the human genome project. DNA Lab, demonstrates how to extract DNA from wheat using readily available equipment and supplies. (http:// ca.youtube.com/watch?v=iyb7fwduuGM) PDB Molecule of the Month pdb23_1 (http://www.rcsb.org/pdb/static.do?p=education_discussion/ molecule_of_the_month/pdb23_1.html) Rosalind Franklin's contributions to the study of DNA (http://mason.gmu.edu/~emoody/rfranklin.html) The Register of Francis Crick Personal Papers 1938 2007 (http://orpheus.ucsd.edu/speccoll/testing/html/ mss0660a.html#abstract) at Mandeville Special Collections Library, University of California, San Diego U.S. National DNA Day (http://www.genome.gov/10506367)watch videos and participate in real-time chat with top scientists Clue to chemistry of heredity found (http://www.nytimes.com/packages/pdf/science/dna-article.pdf) The New York Times June 1953. First American newspaper coverage of the discovery of the DNA structure An Introduction to DNA and Chromosomes (http://hopes.stanford.edu/basics/dna/b0.html) from HOPES: Huntington's Disease Outreach Project for Education at Stanford

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Proteins and amino acids

Protein
Proteins (pronounced/protinz/) are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of adjacent amino acid residues. The sequence of amino acids in a protein is defined by the sequence of a gene, which is encoded in the genetic code. In general, the genetic code specifies 20 standard amino acids; however, in certain organisms the genetic code can include selenocysteineand in certain archaeapyrrolysine. Shortly after or even during synthesis, the residues in a protein are often chemically modified by posttranslational modification, which alters the physical and chemical properties, folding, stability, activity, and ultimately, the function of the proteins. Sometimes proteins have non-peptide groups attached, which can be called prosthetic groups or cofactors. Proteins can also work together to achieve a particular function, and they often associate to form stable protein complexes.

One of the most distinguishing features of polypeptides is their ability to fold into a globular state. The extent to which proteins fold into a defined structure varies widely. Some proteins fold into a highly rigid structure with small fluctuations and are therefore considered to be single structure. Other proteins undergo large rearrangements from one conformation to another. This conformational change is often associated with a signaling event. Thus, the structure of a protein serves as a medium through which to regulate either the function of a protein or activity of an enzyme. Not all proteins require a folding process in order to function, as some function in an unfolded state. Like other biological macromolecules such as polysaccharides and nucleic acids, proteins are essential parts of organisms and participate in virtually every process within cells. Many proteins are enzymes that catalyze biochemical reactions and are vital to metabolism. Proteins also have structural or mechanical functions, such as actin and myosin in muscle and the proteins in the cytoskeleton, which form a system of scaffolding that maintains cell shape. Other proteins are important in cell signaling, immune responses, cell adhesion, and the cell cycle. Proteins are also necessary in animals' diets, since animals cannot synthesize all the amino acids they need and must obtain essential amino acids from food. Through the process of digestion, animals break down ingested protein into free amino acids that are then used in metabolism. Proteins were first described by the Dutch chemist Gerardus Johannes Mulder and named by the Swedish chemist Jns Jacob Berzelius in 1838. Early nutritional scientists such as the German Carl von Voit believed that protein was the most important nutrient for maintaining the structure of the body, because it was generally believed that "flesh makes flesh."[1] The central role of proteins as enzymes in living organisms was however not fully appreciated until 1926, when James B. Sumner showed that the enzyme urease was in fact a protein.[2] The first protein to be sequenced was insulin, by Frederick Sanger, who won the Nobel Prize for this achievement in 1958. The first protein

A representation of the 3D structure of the protein myoglobin showing colored alpha helices. This protein was the first to have its structure solved by X-ray crystallography. Towards the right-center among the coils, a prosthetic group called a heme group is shown colored largely in green.

Protein structures to be solved were hemoglobin and myoglobin, by Max Perutz and Sir John Cowdery Kendrew, respectively, in 1958.[3] [4] The three-dimensional structures of both proteins were first determined by X-ray diffraction analysis; Perutz and Kendrew shared the 1962 Nobel Prize in Chemistry for these discoveries. Proteins may be purified from other cellular components using a variety of techniques such as ultracentrifugation, precipitation, electrophoresis, and chromatography; the advent of genetic engineering has made possible a number of methods to facilitate purification. Methods commonly used to study protein structure and function include immunohistochemistry, site-directed mutagenesis, nuclear magnetic resonance and mass spectrometry. Distributed computing is a relatively new tool researchers are using to examine the infamously complex interactions that govern protein folding; the statistical analysis techniques employed to calculate a protein's probable tertiary structure from its amino acid sequence (primary structure) are well-suited for the distributed computing environment, which has made this otherwise prohibitively expensive and time consuming problem significantly more manageable.

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Biochemistry
Most proteins consist of linear polymers built from series of up to 20 different L--amino acids. All proteinogenic amino acids possess common structural features, including an -carbon to which an amino group, a carboxyl group, and a variable side chain are bonded. Only proline differs from this basic structure as it contains an unusual ring to the N-end amine group, which forces the CONH amide moiety into a fixed conformation.[5] The side chains of the standard amino acids, detailed in the list of standard amino acids, have a great variety of chemical structures and properties; it is the combined effect of all of the amino acid side chains in a protein that ultimately determines its Chemical structure of the peptide bond (bottom) and the three-dimensional structure and its chemical reactivity.[6] The three-dimensional structure of a peptide bond between an alanine and an adjacent amino acid (top/inset) amino acids in a polypeptide chain are linked by peptide bonds. Once linked in the protein chain, an individual amino acid is called a residue, and the linked series of carbon, nitrogen, and oxygen atoms are known as the main chain or protein backbone.[7] The peptide bond has two resonance forms that contribute some double-bond character and inhibit rotation around its axis, so that the alpha carbons are roughly coplanar. The other two dihedral angles in the Resonance structures of the peptide bond that links individual amino peptide bond determine the local shape assumed by the acids to form a protein polymer protein backbone.[8] The end of the protein with a free carboxyl group is known as the C-terminus or carboxy terminus, whereas the end with a free amino group is known as the N-terminus or amino terminus. The words protein, polypeptide, and peptide are a little ambiguous and can overlap in meaning. Protein is generally used to refer to the complete biological molecule in a stable conformation, whereas peptide is generally reserved for a short amino acid oligomers often lacking a stable three-dimensional structure. However, the boundary between the two is not well defined and usually lies near 2030 residues.[9] Polypeptide can refer to any single linear chain of amino acids, usually regardless of length, but often implies an absence of a defined conformation.

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Synthesis
Proteins are assembled from amino acids using information encoded in genes. Each protein has its own unique amino acid sequence that is specified by the nucleotide sequence of the gene encoding this protein. The genetic code is a set of three-nucleotide sets called codons and each three-nucleotide combination designates an amino acid, for example AUG (adenine-uracil-guanine) is the code for methionine. Because DNA contains four nucleotides, the total number of possible codons is 64; hence, there is some redundancy in the genetic code, with some amino acids specified by more than one codon.[10] Genes A ribosome produces a protein using mRNA as template. encoded in DNA are first transcribed into pre-messenger RNA (mRNA) by proteins such as RNA polymerase. Most organisms then process the pre-mRNA (also known as a primary transcript) using various forms of Post-transcriptional modification to form the mature mRNA, which is then used as a template for protein synthesis by the ribosome. In prokaryotes the mRNA may either be used as soon as it is produced, or be bound by a ribosome after having moved away from the nucleoid. In contrast, eukaryotes make mRNA in the cell nucleus The DNA sequence of a gene encodes the amino acid sequence of a protein. and then translocate it across the nuclear membrane into the cytoplasm, where protein synthesis then takes place. The rate of protein synthesis is higher in prokaryotes than eukaryotes and can reach up to 20 amino acids per second.[11] The process of synthesizing a protein from an mRNA template is known as translation. The mRNA is loaded onto the ribosome and is read three nucleotides at a time by matching each codon to its base pairing anticodon located on a transfer RNA molecule, which carries the amino acid corresponding to the codon it recognizes. The enzyme aminoacyl tRNA synthetase "charges" the tRNA molecules with the correct amino acids. The growing polypeptide is often termed the nascent chain. Proteins are always biosynthesized from N-terminus to C-terminus.[10] The size of a synthesized protein can be measured by the number of amino acids it contains and by its total molecular mass, which is normally reported in units of daltons (synonymous with atomic mass units), or the derivative unit kilodalton (kDa). Yeast proteins are on average 466 amino acids long and 53 kDa in mass.[9] The largest known proteins are the titins, a component of the muscle sarcomere, with a molecular mass of almost 3,000 kDa and a total length of almost 27,000 amino acids.[12]

Chemical synthesis
Short proteins can also be synthesized chemically by a family of methods known as peptide synthesis, which rely on organic synthesis techniques such as chemical ligation to produce peptides in high yield.[13] Chemical synthesis allows for the introduction of non-natural amino acids into polypeptide chains, such as attachment of fluorescent probes to amino acid side chains.[14] These methods are useful in laboratory biochemistry and cell biology, though generally not for commercial applications. Chemical synthesis is inefficient for polypeptides longer than about 300 amino acids, and the synthesized proteins may not readily assume their native tertiary structure. Most chemical synthesis methods proceed from C-terminus to N-terminus, opposite the biological reaction.[15]

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Structure
Further information: Protein structure prediction Mostproteins fold into unique 3-dimensional structures. The shape into which a protein naturally folds is known as its native conformation.[16] Although many proteins can fold unassisted, simply through the chemical properties of their amino acids, others require the aid of molecular chaperones to fold into their native states.[17] Biochemists often refer to four distinct aspects of a protein's structure:[18] Primary structure: the amino acid sequence. Secondary structure: regularly repeating local structures stabilized by hydrogen bonds. The most common examples are the alpha helix, beta sheet and turns. Because secondary structures are local, many regions of different secondary structure can be present in the same protein molecule.

The crystal structure of the chaperonin. Chaperonins assist protein folding.

Tertiary structure: the overall shape of a single protein molecule; the spatial relationship of the secondary structures to one another. Tertiary structure is generally stabilized by nonlocal interactions, most commonly the formation of a hydrophobic core, but also through salt bridges, hydrogen bonds, disulfide bonds, and even posttranslational modifications. The term "tertiary structure" is often used as synonymous with the term fold. The tertiary structure is what controls the basic function of the protein. Quaternary structure: the structure formed by several protein molecules (polypeptide chains), usually called protein subunits in this context, which function as a single protein complex.

Three possible representations of the three-dimensional structure of the protein triose phosphate isomerase. Left: all-atom representation colored by atom type. Middle: Simplified representation illustrating the backbone conformation, colored by secondary structure. Right: Solvent-accessible surface representation colored by residue type (acidic residues red, basic residues blue, polar residues green, nonpolar residues white)

Proteins are not entirely rigid molecules. In addition to these levels of structure, proteins may shift between several related structures while they perform their functions. In the context of these functional rearrangements, these tertiary or quaternary structures are usually referred to as "conformations", and transitions between them are called conformational changes. Such changes are often induced by the binding of a substrate molecule to an enzyme's active site, or the physical region of the protein that participates in chemical catalysis. In solution proteins also undergo variation in structure through thermal vibration and the collision with other molecules.[19]

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Proteins can be informally divided into three main classes, which correlate with typical tertiary structures: globular proteins, fibrous proteins, and membrane proteins. Almost all globular proteins are soluble and many are enzymes. Fibrous proteins are often Molecular surface of several proteins showing their comparative sizes. From left to structural, such as collagen, the major right are: immunoglobulin G (IgG, an antibody), hemoglobin, insulin (a hormone), component of connective tissue, or keratin, adenylate kinase (an enzyme), and glutamine synthetase (an enzyme). the protein component of hair and nails. Membrane proteins often serve as receptors or provide channels for polar or charged molecules to pass through the cell membrane.[20] A special case of intramolecular hydrogen bonds within proteins, poorly shielded from water attack and hence promoting their own dehydration, are called dehydrons.[21]

Structure determination
Discovering the tertiary structure of a protein, or the quaternary structure of its complexes, can provide important clues about how the protein performs its function. Common experimental methods of structure determination include X-ray crystallography and NMR spectroscopy, both of which can produce information at atomic resolution. However, NMR experiments are able to provide information from which a subset of distances between pairs of atoms can be estimated, and the final possible conformations for a protein are determined by solving a distance geometry problem. Dual polarisation interferometry is a quantitative analytical method for measuring the overall protein conformation and conformational changes due to interactions or other stimulus. Circular dichroism is another laboratory technique for determining internal beta sheet/ helical composition of proteins. Cryoelectron microscopy is used to produce lower-resolution structural information about very large protein complexes, including assembled viruses;[22] a variant known as electron crystallography can also produce high-resolution information in some cases, especially for two-dimensional crystals of membrane proteins.[23] Solved structures are usually deposited in the Protein Data Bank (PDB), a freely available resource from which structural data about thousands of proteins can be obtained in the form of Cartesian coordinates for each atom in the protein.[24] Many more gene sequences are known than protein structures. Further, the set of solved structures is biased toward proteins that can be easily subjected to the conditions required in X-ray crystallography, one of the major structure determination methods. In particular, globular proteins are comparatively easy to crystallize in preparation for X-ray crystallography. Membrane proteins, by contrast, are difficult to crystallize and are underrepresented in the PDB.[25] Structural genomics initiatives have attempted to remedy these deficiencies by systematically solving representative structures of major fold classes. Protein structure prediction methods attempt to provide a means of generating a plausible structure for proteins whose structures have not been experimentally determined.

Cellular functions
Proteins are the chief actors within the cell, said to be carrying out the duties specified by the information encoded in genes.[9] With the exception of certain types of RNA, most other biological molecules are relatively inert elements upon which proteins act. Proteins make up half the dry weight of an Escherichia coli cell, whereas other macromolecules such as DNA and RNA make up only 3% and 20%, respectively.[26] The set of proteins expressed in a particular cell or cell type is known as its proteome.

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The chief characteristic of proteins that also allows their diverse set of functions is their ability to bind other molecules specifically and tightly. The region of the protein responsible for binding another molecule is known as the binding site and is often a depression or "pocket" on the molecular surface. This binding ability is mediated by the tertiary structure of the protein, which defines the binding site pocket, and by the chemical properties of the surrounding amino acids' side chains. Protein binding can be extraordinarily tight and specific; for example, the ribonuclease inhibitor protein binds to human The enzyme hexokinase is shown as a angiogenin with a sub-femtomolar dissociation constant (<1015 M) conventional ball-and-stick molecular model. To scale in the top right-hand corner are two of its but does not bind at all to its amphibian homolog onconase (>1 M). substrates, ATP and glucose. Extremely minor chemical changes such as the addition of a single methyl group to a binding partner can sometimes suffice to nearly eliminate binding; for example, the aminoacyl tRNA synthetase specific to the amino acid valine discriminates against the very similar side chain of the amino acid isoleucine.[27] Proteins can bind to other proteins as well as to small-molecule substrates. When proteins bind specifically to other copies of the same molecule, they can oligomerize to form fibrils; this process occurs often in structural proteins that consist of globular monomers that self-associate to form rigid fibers. Proteinprotein interactions also regulate enzymatic activity, control progression through the cell cycle, and allow the assembly of large protein complexes that carry out many closely related reactions with a common biological function. Proteins can also bind to, or even be integrated into, cell membranes. The ability of binding partners to induce conformational changes in proteins allows the construction of enormously complex signaling networks.[28] Importantly, as interactions between proteins are reversible, and depend heavily on the availability of different groups of partner proteins to form aggregates that are capable to carry out discrete sets of function, study of the interactions between specific proteins is a key to understand important aspects of cellular function, and ultimately the properties that distinguish particular cell types.[29] [30]

Enzymes
The best-known role of proteins in the cell is as enzymes, which catalyze chemical reactions. Enzymes are usually highly specific and accelerate only one or a few chemical reactions. Enzymes carry out most of the reactions involved in metabolism, as well as manipulating DNA in processes such as DNA replication, DNA repair, and transcription. Some enzymes act on other proteins to add or remove chemical groups in a process known as posttranslational modification. About 4,000 reactions are known to be catalyzed by enzymes.[31] The rate acceleration conferred by enzymatic catalysis is often enormousas much as 1017-fold increase in rate over the uncatalyzed reaction in the case of orotate decarboxylase (78 million years without the enzyme, 18 milliseconds with the enzyme).[32] The molecules bound and acted upon by enzymes are called substrates. Although enzymes can consist of hundreds of amino acids, it is usually only a small fraction of the residues that come in contact with the substrate, and an even smaller fractionthree to four residues on averagethat are directly involved in catalysis.[33] The region of the enzyme that binds the substrate and contains the catalytic residues is known as the active site.

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Cell signaling and ligand binding

Many proteins are involved in the process of cell signaling and signal transduction. Some proteins, such as insulin, are extracellular proteins that transmit a signal from the cell in which they were synthesized to other cells in distant tissues. Others are membrane proteins that act as receptors whose main function is to bind a signaling molecule and induce a biochemical response in the cell. Many receptors have a binding site exposed on the cell surface and an effector domain within the cell, which may have enzymatic activity or may undergo a conformational change detected by other proteins within the cell.[34] Antibodies are protein components of an adaptive immune system whose main function is to bind antigens, or foreign substances in the body, and target them for destruction. Antibodies can be secreted into the extracellular environment or anchored in the membranes of specialized B cells known as plasma cells. Whereas enzymes are limited in their binding affinity for their substrates by the necessity of conducting their reaction, antibodies have no such constraints. An antibody's binding affinity to its target is extraordinarily high.[35]

Many ligand transport proteins bind particular small biomolecules and transport them to other locations in the body of a multicellular organism. These proteins must have a high binding affinity when their ligand is present in high concentrations, but must also release the ligand when it is present at low concentrations in the target tissues. The canonical example of a ligand-binding protein is haemoglobin, which transports oxygen from the lungs to other organs and tissues in all vertebrates and has close homologs in every biological kingdom.[36] Lectins are sugar-binding proteins which are highly specific for their sugar moieties. Lectins typically play a role in biological recognition phenomena involving cells and proteins.[37] Receptors and hormones are highly specific binding proteins. Transmembrane proteins can also serve as ligand transport proteins that alter the permeability of the cell membrane to small molecules and ions. The membrane alone has a hydrophobic core through which polar or charged molecules cannot diffuse. Membrane proteins contain internal channels that allow such molecules to enter and exit the cell. Many ion channel proteins are specialized to select for only a particular ion; for example, potassium and sodium channels often discriminate for only one of the two ions.[38]

Ribbon diagram of a mouse antibody against cholera that binds a carbohydrate antigen

Structural proteins
Structural proteins confer stiffness and rigidity to otherwise-fluid biological components. Most structural proteins are fibrous proteins; for example, actin and tubulin are globular and soluble as monomers, but polymerize to form long, stiff fibers that make up the cytoskeleton, which allows the cell to maintain its shape and size. Collagen and elastin are critical components of connective tissue such as cartilage, and keratin is found in hard or filamentous structures such as hair, nails, feathers, hooves, and some animal shells.[39] Other proteins that serve structural functions are motor proteins such as myosin, kinesin, and dynein, which are capable of generating mechanical forces. These proteins are crucial for cellular motility of single celled organisms and the sperm of many multicellular organisms which reproduce sexually. They also generate the forces exerted by contracting muscles.[40]

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Methods of study
As some of the most commonly studied biological molecules, the activities and structures of proteins are examined both in vitro and in vivo. In vitro studies of purified proteins in controlled environments are useful for learning how a protein carries out its function: for example, enzyme kinetics studies explore the chemical mechanism of an enzyme's catalytic activity and its relative affinity for various possible substrate molecules. By contrast, in vivo experiments on proteins' activities within cells or even within whole organisms can provide complementary information about where a protein functions and how it is regulated.

Protein purification
In order to perform in vitro analysis, a protein must be purified away from other cellular components. This process usually begins with cell lysis, in which a cell's membrane is disrupted and its internal contents released into a solution known as a crude lysate. The resulting mixture can be purified using ultracentrifugation, which fractionates the various cellular components into fractions containing soluble proteins; membrane lipids and proteins; cellular organelles, and nucleic acids. Precipitation by a method known as salting out can concentrate the proteins from this lysate. Various types of chromatography are then used to isolate the protein or proteins of interest based on properties such as molecular weight, net charge and binding affinity.[41] The level of purification can be monitored using various types of gel electrophoresis if the desired protein's molecular weight and isoelectric point are known, by spectroscopy if the protein has distinguishable spectroscopic features, or by enzyme assays if the protein has enzymatic activity. Additionally, proteins can be isolated according their charge using electrofocusing.[42] For natural proteins, a series of purification steps may be necessary to obtain protein sufficiently pure for laboratory applications. To simplify this process, genetic engineering is often used to add chemical features to proteins that make them easier to purify without affecting their structure or activity. Here, a "tag" consisting of a specific amino acid sequence, often a series of histidine residues (a "His-tag"), is attached to one terminus of the protein. As a result, when the lysate is passed over a chromatography column containing nickel, the histidine residues ligate the nickel and attach to the column while the untagged components of the lysate pass unimpeded. A number of different tags have been developed to help researchers purify specific proteins from complex mixtures.[43]

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Cellular localization
The study of proteins in vivo is often concerned with the synthesis and localization of the protein within the cell. Although many intracellular proteins are synthesized in the cytoplasm and membrane-bound or secreted proteins in the endoplasmic reticulum, the specifics of how proteins are targeted to specific organelles or cellular structures is often unclear. A useful technique for assessing cellular localization uses genetic engineering to express in a cell a fusion protein or chimera consisting of the natural protein of interest linked to a "reporter" such as green fluorescent protein (GFP).[44] The fused protein's position within the cell can be cleanly and efficiently visualized using microscopy,[45] as shown in the figure opposite. Other methods for elucidating the cellular location of proteins requires the use of known compartmental markers for regions such as the ER, the Golgi, lysosomes/vacuoles, mitochondria, chloroplasts, plasma membrane, etc. With the use of fluorescently tagged versions Proteins in different cellular compartments and structures tagged with green of these markers or of antibodies to known fluorescent protein (here, white) markers, it becomes much simpler to identify the localization of a protein of interest. For example, indirect immunofluorescence will allow for fluorescence colocalization and demonstration of location. Fluorescent dyes are used to label cellular compartments for a similar purpose.[46] Other possibilities exist, as well. For example, immunohistochemistry usually utilizes an antibody to one or more proteins of interest that are conjugated to enzymes yielding either luminescent or chromogenic signals that can be compared between samples, allowing for localization information. Another applicable technique is cofractionation in sucrose (or other material) gradients using isopycnic centrifugation.[47] While this technique does not prove colocalization of a compartment of known density and the protein of interest, it does increase the likelihood, and is more amenable to large-scale studies. Finally, the gold-standard method of cellular localization is immunoelectron microscopy. This technique also uses an antibody to the protein of interest, along with classical electron microscopy techniques. The sample is prepared for normal electron microscopic examination, and then treated with an antibody to the protein of interest that is conjugated to an extremely electro-dense material, usually gold. This allows for the localization of both ultrastructural details as well as the protein of interest.[48] Through another genetic engineering application known as site-directed mutagenesis, researchers can alter the protein sequence and hence its structure, cellular localization, and susceptibility to regulation. This technique even allows the incorporation of unnatural amino acids into proteins, using modified tRNAs,[49] and may allow the rational design of new proteins with novel properties.[50]

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Proteomics and bioinformatics

The total complement of proteins present at a time in a cell or cell type is known as its proteome, and the study of such large-scale data sets defines the field of proteomics, named by analogy to the related field of genomics. Key experimental techniques in proteomics include 2D electrophoresis,[51] which allows the separation of a large number of proteins, mass spectrometry,[52] which allows rapid high-throughput identification of proteins and sequencing of peptides (most often after in-gel digestion), protein microarrays,[53] which allow the detection of the relative levels of a large number of proteins present in a cell, and two-hybrid screening, which allows the systematic exploration of proteinprotein interactions.[54] The total complement of biologically possible such interactions is known as the interactome.[55] A systematic attempt to determine the structures of proteins representing every possible fold is known as structural genomics.[56] The large amount of genomic and proteomic data available for a variety of organisms, including the human genome, allows researchers to efficiently identify homologous proteins in distantly related organisms by sequence alignment. Sequence profiling tools can perform more specific sequence manipulations such as restriction enzyme maps, open reading frame analyses for nucleotide sequences, and secondary structure prediction. From this data phylogenetic trees can be constructed and evolutionary hypotheses developed using special software like ClustalW regarding the ancestry of modern organisms and the genes they express. The field of bioinformatics seeks to assemble, annotate, and analyze genomic and proteomic data, applying computational techniques to biological problems such as gene finding and cladistics.

Structure prediction and simulation

Complementary to the field of structural genomics, protein structure prediction seeks to develop efficient ways to provide plausible models for proteins whose structures have not yet been determined experimentally.[57] The most successful type of structure prediction, known as homology modeling, relies on the existence of a "template" structure with sequence similarity to the protein being modeled; structural genomics' goal is to provide sufficient representation in solved structures to model most of those that remain.[58] Although producing accurate models remains a challenge when only distantly related template structures are available, it has been suggested that sequence alignment is the bottleneck in this process, as quite accurate models can be produced if a "perfect" sequence alignment is known.[59] Many structure prediction methods have served to inform the emerging field of protein engineering, in which novel protein folds have already been designed.[60] A more complex computational problem is the prediction of intermolecular interactions, such as in molecular docking and proteinprotein interaction prediction.[61] The processes of protein folding and binding can be simulated using such technique as molecular mechanics, in particular, molecular dynamics and Monte Carlo, which increasingly take advantage of parallel and distributed computing (Folding@Home project;[62] molecular modeling on GPU). The folding of small alpha-helical protein domains such as the villin headpiece[63] and the HIV accessory protein[64] have been successfully simulated in silico, and hybrid methods that combine standard molecular dynamics with quantum mechanics calculations have allowed exploration of the electronic states of rhodopsins.[65]

Nutrition
Further information: Protein (nutrient) Most microorganisms and plants can biosynthesize all 20 standard amino acids, while animals (including humans) must obtain some of the amino acids from the diet.[26] The amino acids that an organism cannot synthesize on its own are referred to as essential amino acids. Key enzymes that synthesize certain amino acids are not present in animals such as aspartokinase, which catalyzes the first step in the synthesis of lysine, methionine, and threonine from aspartate. If amino acids are present in the environment, microorganisms can conserve energy by taking up the amino acids from their surroundings and downregulating their biosynthetic pathways.

Protein In animals, amino acids are obtained through the consumption of foods containing protein. Ingested proteins are then broken down into amino acids through digestion, which typically involves denaturation of the protein through exposure to acid and hydrolysis by enzymes called proteases. Some ingested amino acids are used for protein biosynthesis, while others are converted to glucose through gluconeogenesis, or fed into the citric acid cycle. This use of protein as a fuel is particularly important under starvation conditions as it allows the body's own proteins to be used to support life, particularly those found in muscle.[66] Amino acids are also an important dietary source of nitrogen.

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History and etymology

Further information: History of molecular biology Proteins were recognized as a distinct class of biological molecules in the eighteenth century by Antoine Fourcroy and others, distinguished by the molecules' ability to coagulate or flocculate under treatments with heat or acid. Noted examples at the time included albumin from egg whites, blood serum albumin, fibrin, and wheat gluten. Dutch chemist Gerardus Johannes Mulder carried out elemental analysis of common proteins and found that nearly all proteins had the same empirical formula, C400H620N100O120P1S1.[67] He came to the erroneous conclusion that they might be composed of a single type of (very large) molecule. The term "protein" to describe these molecules was proposed in 1838 by Mulder's associate Jns Jacob Berzelius; protein is derived from the Greek word (proteios), meaning "primary",[68] "in the lead", or "standing in front".[69] Mulder went on to identify the products of protein degradation such as the amino acid leucine for which he found a (nearly correct) molecular weight of 131 Da.[67] The difficulty in purifying proteins in large quantities made them very difficult for early protein biochemists to study. Hence, early studies focused on proteins that could be purified in large quantities, e.g., those of blood, egg white, various toxins, and digestive/metabolic enzymes obtained from slaughterhouses. In the 1950s, the Armour Hot Dog Co. purified 1kg of pure bovine pancreatic ribonuclease A and made it freely available to scientists; this gesture helped ribonuclease A become a major target for biochemical study for the following decades.[67] Linus Pauling is credited with the successful prediction of regular protein secondary structures based on hydrogen bonding, an idea first put forth by William Astbury in 1933.[70] Later work by Walter Kauzmann on denaturation,[71] [72] based partly on previous studies by Kaj Linderstrm-Lang,[73] contributed an understanding of protein folding and structure mediated by hydrophobic interactions. In 1949 Fred Sanger correctly determined the amino acid sequence of insulin, thus conclusively demonstrating that proteins consisted of linear polymers of amino acids rather than branched chains, colloids, or cyclols.[74] The first atomic-resolution structures of proteins were solved by X-ray crystallography in the 1960s and by NMR in the 1980s. As of 2009, the Protein Data Bank has over 55,000 atomic-resolution structures of proteins.[75] In more recent times, cryo-electron microscopy of large macromolecular assemblies[76] and computational protein structure prediction of small protein domains[77] are two methods approaching atomic resolution.

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Footnotes
[1] Bischoff TLW, Voit, C (1860) (in German). Die Gesetze der Ernaehrung des Pflanzenfressers durch neue Untersuchungen festgestellt. Leipzig, Heidelberg. [2] Sumner JB (1926). "The isolation and crystallization of the enzyme urease. Preliminary paper" (http:/ / www. jbc. org/ content/ 69/ 2/ 435. full. pdf+ html) (PDF). Journal of Biological Chemistry 69 (2): 43541. . [3] Muirhead H, Perutz M (1963). "Structure of hemoglobin. A three-dimensional fourier synthesis of reduced human hemoglobin at 5.5 resolution". Nature 199 (4894): 63338. doi:10.1038/199633a0. PMID14074546. [4] Kendrew J, Bodo G, Dintzis H, Parrish R, Wyckoff H, Phillips D (1958). "A three-dimensional model of the myoglobin molecule obtained by X-ray analysis". Nature 181 (4610): 66266. Bibcode1958Natur.181..662K. doi:10.1038/181662a0. PMID13517261. [5] Nelson DL, Cox MM (2005). Lehninger's Principles of Biochemistry (4th ed.). New York, New York: W. H. Freeman and Company. [6] Gutteridge A, Thornton JM (2005). "Understanding nature's catalytic toolkit". Trends in Biochemical Sciences 30 (11): 62229. doi:10.1016/j.tibs.2005.09.006. PMID16214343. [7] Murray et al., p. 19. [8] Murray et al., p. 31. [9] Lodish H, Berk A, Matsudaira P, Kaiser CA, Krieger M, Scott MP, Zipurksy SL, Darnell J (2004). Molecular Cell Biology (5th ed.). New York, New York: WH Freeman and Company. [10] van Holde and Mathews, pp. 100242. [11] Dobson CM (2000). "The nature and significance of protein folding". In Pain RH (ed.). Mechanisms of Protein Folding. Oxford, Oxfordshire: Oxford University Press. pp.128. ISBN0-19-963789-X. [12] Fulton A, Isaacs W (1991). "Titin, a huge, elastic sarcomeric protein with a probable role in morphogenesis". Bioessays 13 (4): 15761. doi:10.1002/bies.950130403. PMID1859393. [13] Bruckdorfer T, Marder O, Albericio F (2004). "From production of peptides in milligram amounts for research to multi-tons quantities for drugs of the future". Current Pharmaceutical Biotechnology 5 (1): 2943. doi:10.2174/1389201043489620. PMID14965208. [14] Schwarzer D, Cole P (2005). "Protein semisynthesis and expressed protein ligation: chasing a protein's tail". Current Opinions in Chemical Biology 9 (6): 56169. doi:10.1016/j.cbpa.2005.09.018. PMID16226484. [15] Kent SB (2009). "Total chemical synthesis of proteins". Chemical Society Reviews 38 (2): 33851. doi:10.1039/b700141j. PMID19169452. [16] Murray et al., p. 36. [17] Murray et al., p. 37. [18] Murray et al., pp. 3034. [19] van Holde and Mathews, pp. 36875. [20] van Holde and Mathews, pp. 16585. [21] Fernndez A, Scott R (2003). "Dehydron: a structurally encoded signal for protein interaction" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0006-3495(03)74619-0). Biophysical Journal 85 (3): 191428. Bibcode2003BpJ....85.1914F. doi:10.1016/S0006-3495(03)74619-0. PMC1303363. PMID12944304. . [22] Branden and Tooze, pp. 34041. [23] Gonen T, Cheng Y, Sliz P, Hiroaki Y, Fujiyoshi Y, Harrison SC, Walz T (2005). "Lipid-protein interactions in double-layered two-dimensional AQP0 crystals". Nature 438 (7068): 63338. doi:10.1038/nature04321. PMC1350984. PMID16319884. [24] Standley DM, Kinjo AR, Kinoshita K, Nakamura H (2008). "Protein structure databases with new web services for structural biology and biomedical research" (http:/ / bib. oxfordjournals. org/ cgi/ pmidlookup?view=long& pmid=18430752). Briefings in Bioinformatics 9 (4): 27685. doi:10.1093/bib/bbn015. PMID18430752. . [25] Walian P, Cross TA, Jap BK (2004). "Structural genomics of membrane proteins" (http:/ / genomebiology. com/ 2004/ 5/ 4/ 215). Genome Biology 5 (4): 215. doi:10.1186/gb-2004-5-4-215. PMC395774. PMID15059248. . [26] Voet D, Voet JG. (2004). Biochemistry Vol 1 3rd ed. Wiley: Hoboken, NJ. [27] Sankaranarayanan R, Moras D (2001). "The fidelity of the translation of the genetic code". Acta Biochimica Polonica 48 (2): 32335. PMID11732604. [28] van Holde and Mathews, pp. 83049. [29] Copland JA, Sheffield-Moore M, Koldzic-Zivanovic N, Gentry S, Lamprou G, Tzortzatou-Stathopoulou F, Zoumpourlis V, Urban RJ, Vlahopoulos SA (2009). "Sex steroid receptors in skeletal differentiation and epithelial neoplasia: is tissue-specific intervention possible?". BioEssays: news and reviews in molecular, cellular and developmental biology 31 (6): 62941. doi:10.1002/bies.200800138. PMID19382224. [30] Samarin S, Nusrat A (2009). "Regulation of epithelial apical junctional complex by Rho family GTPases". Frontiers in bioscience: a journal and virtual library 14: 112942. PMID19273120. [31] Bairoch A (2000). "The ENZYME database in 2000" (http:/ / www. expasy. org/ NAR/ enz00. pdf). Nucleic Acids Research 28 (1): 304305. doi:10.1093/nar/28.1.304. PMC102465. PMID10592255. . [32] Radzicka A, Wolfenden R (1995). "A proficient enzyme". Science 6 (267): 9093. doi:10.1126/science.7809611. PMID7809611. [33] EBI External Services (2010-01-20). "The Catalytic Site Atlas at The European Bioinformatics Institute" (http:/ / www. ebi. ac. uk/ thornton-srv/ databases/ CSA/ ). Ebi.ac.uk. . Retrieved 2011-01-16. [34] Branden and Tooze, pp. 25181.

Protein
[35] van Holde and Mathews, pp. 24750. [36] van Holde and Mathews, pp. 22029. [37] Rdiger H, Siebert HC, Sols D, Jimnez-Barbero J, Romero A, von der Lieth CW, Diaz-Mario T, Gabius HJ (2000). "Medicinal chemistry based on the sugar code: fundamentals of lectinology and experimental strategies with lectins as targets". Current Medicinal Chemistry 7 (4): 389416. PMID10702616. [38] Branden and Tooze, pp. 23234. [39] van Holde and Mathews, pp. 17881. [40] van Holde and Mathews, pp. 25864; 272. [41] Murray et al., pp. 2124. [42] Hey J, Posch A, Cohen A, Liu N, Harbers A (2008). "Fractionation of complex protein mixtures by liquid-phase isoelectric focusing". Methods in Molecular Biology 424: 22539. doi:10.1007/978-1-60327-064-9_19. PMID18369866. [43] Terpe K (2003). "Overview of tag protein fusions: from molecular and biochemical fundamentals to commercial systems". Applied Microbiology and Biotechnology 60 (5): 52333. doi:10.1007/s00253-002-1158-6. PMID12536251. [44] Stepanenko OV, Verkhusha VV, Kuznetsova IM, Uversky VN, Turoverov KK (2008). "Fluorescent proteins as biomarkers and biosensors: throwing color lights on molecular and cellular processes". Current Protein & Peptide Science 9 (4): 33869. doi:10.2174/138920308785132668. PMC2904242. PMID18691124. [45] Yuste R (2005). "Fluorescence microscopy today". Nature Methods 2 (12): 902904. doi:10.1038/nmeth1205-902. PMID16299474. [46] Margolin W (2000). "Green fluorescent protein as a reporter for macromolecular localization in bacterial cells" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S1046-2023(99)90906-4). Methods (San Diego, Calif.) 20 (1): 6272. doi:10.1006/meth.1999.0906. PMID10610805. . [47] Walker JH, Wilson K (2000). Principles and Techniques of Practical Biochemistry. Cambridge, UK: Cambridge University Press. pp.28789. ISBN0-521-65873-X. [48] Mayhew TM, Lucocq JM (2008). "Developments in cell biology for quantitative immunoelectron microscopy based on thin sections: a review". Histochemistry and Cell Biology 130 (2): 299313. doi:10.1007/s00418-008-0451-6. PMC2491712. PMID18553098. [49] Hohsaka T, Sisido M (2002). "Incorporation of non-natural amino acids into proteins". Current Opinion in Chemical Biology 6 (6): 80915. doi:10.1016/S1367-5931(02)00376-9. PMID12470735. [50] Cedrone F, Mnez A, Qumneur E (2000). "Tailoring new enzyme functions by rational redesign". Current Opinion in Structural Biology 10 (4): 40510. doi:10.1016/S0959-440X(00)00106-8. PMID10981626. [51] Grg A, Weiss W, Dunn MJ (2004). "Current two-dimensional electrophoresis technology for proteomics". Proteomics 4 (12): 366585. doi:10.1002/pmic.200401031. PMID15543535. [52] Conrotto P, Souchelnytskyi S (2008). "Proteomic approaches in biological and medical sciences: principles and applications" (http:/ / www. exp-oncology. com. ua/ en/ archives/ 36/ 699. html). Experimental Oncology 30 (3): 17180. PMID18806738. . [53] Joos T, Bachmann J (2009). "Protein microarrays: potentials and limitations" (http:/ / www. bioscience. org/ 2009/ v14/ af/ 3534/ fulltext. htm). Frontiers in Bioscience 14 (14): 437685. doi:10.2741/3534. PMID19273356. . [54] Koegl M, Uetz P (2007). "Improving yeast two-hybrid screening systems" (http:/ / bfgp. oxfordjournals. org/ cgi/ pmidlookup?view=long& pmid=18218650). Briefings in Functional Genomics & Proteomics 6 (4): 30212. doi:10.1093/bfgp/elm035. PMID18218650. . [55] Plewczyski D, Ginalski K (2009). "The interactome: predicting the proteinprotein interactions in cells". Cellular & Molecular Biology Letters 14 (1): 122. doi:10.2478/s11658-008-0024-7. PMID18839074. [56] Zhang C, Kim SH (2003). "Overview of structural genomics: from structure to function". Current Opinion in Chemical Biology 7 (1): 2832. doi:10.1016/S1367-5931(02)00015-7. PMID12547423. [57] Zhang Y (2008). "Progress and challenges in protein structure prediction". Current Opinions in Structural Biology 18 (3): 34248. doi:10.1016/j.sbi.2008.02.004. PMC2680823. PMID18436442. [58] Xiang Z (2006). "Advances in homology protein structure modeling". Current Protein and Peptide Science 7 (3): 21727. doi:10.2174/138920306777452312. PMC1839925. PMID16787261. [59] Zhang Y, Skolnick J (2005). "The protein structure prediction problem could be solved using the current PDB library" (http:/ / www. pnas. org/ cgi/ pmidlookup?view=long& pmid=15653774). Proceedings of the National Academy of Sciences U.S.A. 102 (4): 102934. doi:10.1073/pnas.0407152101. PMC545829. PMID15653774. . [60] Kuhlman B, Dantas G, Ireton GC, Varani G, Stoddard BL, Baker D (2003). "Design of a novel globular protein fold with atomic-level accuracy" (http:/ / www. sciencemag. org/ cgi/ pmidlookup?view=long& pmid=14631033). Science 302 (5649): 136468. Bibcode2003Sci...302.1364K. doi:10.1126/science.1089427. PMID14631033. . [61] Ritchie DW (2008). "Recent progress and future directions in proteinprotein docking". Current Protein and Peptide Science 9 (1): 115. doi:10.2174/138920308783565741. PMID18336319. [62] Scheraga HA, Khalili M, Liwo A (2007). "Protein-folding dynamics: overview of molecular simulation techniques". Annual Review of Physical Chemistry 58: 5783. doi:10.1146/annurev.physchem.58.032806.104614. PMID17034338. [63] Zagrovic B, Snow CD, Shirts MR, Pande VS (2002). "Simulation of folding of a small alpha-helical protein in atomistic detail using worldwide-distributed computing". Journal of Molecular Biology 323 (5): 92737. doi:10.1016/S0022-2836(02)00997-X. PMID12417204. [64] Herges T, Wenzel W (2005). "In silico folding of a three helix protein and characterization of its free-energy landscape in an all-atom force field". Physical Review Letters 94 (1): 018101. Bibcode2005PhRvL..94a8101H. doi:10.1103/PhysRevLett.94.018101. PMID15698135. [65] Hoffmann M, Wanko M, Strodel P, Knig PH, Frauenheim T, Schulten K, Thiel W, Tajkhorshid E, Elstner M (2006). "Color tuning in rhodopsins: the mechanism for the spectral shift between bacteriorhodopsin and sensory rhodopsin II". Journal of the American Chemical

91

Protein
Society 128 (33): 1080818. doi:10.1021/ja062082i. PMID16910676. [66] Brosnan J (June 2003). "Interorgan amino acid transport and its regulation" (http:/ / jn. nutrition. org/ cgi/ content/ full/ 133/ 6/ 2068S). Journal of Nutrition 133 (6 Suppl 1): 2068S72S. PMID12771367. . [67] Perrett D (2007). "From 'protein' to the beginnings of clinical proteomics". Proteomics Clinical Applications 1 (8): 72038. doi:10.1002/prca.200700525. PMID21136729. [68] New Oxford Dictionary of English [69] Reynolds JA, Tanford C (2003). Nature's Robots: A History of Proteins (Oxford Paperbacks). New York, New York: Oxford University Press. p.15. ISBN0-19-860694-X. [70] Pauling L, Corey RB, Branson HR (1951). "The structure of proteins: two hydrogen-bonded helical configurations of the polypeptide chain" (http:/ / www. pnas. org/ site/ misc/ Protein8. pdf) (PDF). Proceedings of the National Academy of Sciences U.S.A. 37 (5): 23540. Bibcode1951PNAS...37..235P. doi:10.1073/pnas.37.5.235. PMC1063348. PMID14834145. . [71] Kauzmann W (1956). "Structural factors in protein denaturation". Journal of Cellular Physiology. Supplement 47 (Suppl 1): 11331. doi:10.1002/jcp.1030470410. PMID13332017. [72] Kauzmann W (1959). "Some factors in the interpretation of protein denaturation". Advances in Protein Chemistry 14: 163. doi:10.1016/S0065-3233(08)60608-7. PMID14404936. [73] Kalman SM, Linderstrom-Lang K, Ottesen M, Richards FM (1955). "Degradation of ribonuclease by subtilisin". Biochimica et Biophysica Acta 16 (2): 29799. doi:10.1016/0006-3002(55)90224-9. PMID14363272. [74] Sanger F (1949). "The terminal peptides of insulin". Biochemical Journal 45 (5): 56374. PMC1275055. PMID15396627. [75] "RCSB Protein Data Bank" (http:/ / www. rcsb. org/ pdb/ home/ home. do). . Retrieved 2009-04-14. [76] Zhou ZH (2008). "Towards atomic resolution structural determination by single-particle cryo-electron microscopy". Current Opinion in Structural Biology 18 (2): 21828. doi:10.1016/j.sbi.2008.03.004. PMC2714865. PMID18403197. [77] Keskin O, Tuncbag N, Gursoy A (2008). "Characterization and prediction of protein interfaces to infer protein-protein interaction networks". Current Pharmaceutical Biotechnology 9 (2): 6776. doi:10.2174/138920108783955191. PMID18393863.

92

References
Branden C, Tooze J (1999). Introduction to Protein Structure. New York: Garland Pub. ISBN0-8153-2305-0. Murray RF, Harper HW, Granner DK, Mayes PA, Rodwell VW (2006). Harper's Illustrated Biochemistry. New York: Lange Medical Books/McGraw-Hill. ISBN0-07-146197-3. Van Holde KE, Mathews CK (1996). Biochemistry. Menlo Park, California: Benjamin/Cummings Pub. Co., Inc. ISBN0-8053-3931-0.

External links
Databases and projects
Comparative Toxicogenomics Database (http://ctd.mdibl.org/) curates proteinchemical interactions, as well as gene/proteindisease relationships and chemical-disease relationships. Bioinformatic Harvester (http://harvester.fzk.de) A Meta search engine (29 databases) for gene and protein information. The Protein Databank (http://www.rcsb.org) (see also PDB Molecule of the Month (http://www.rcsb.org/ pdb/static.do?p=education_discussion/molecule_of_the_month/index.html), presenting short accounts on selected proteins from the PDB) Proteopedia Life in 3D (http://www.proteopedia.org): rotatable, zoomable 3D model with wiki annotations for every known protein molecular structure. UniProt the Universal Protein Resource (http://www.expasy.uniprot.org) neXtProt Exploring the universe of human proteins (http://www.nextprot.org): human-centric protein knowledge resource The Protein Naming Utility (http://www.jcvi.org/pn-utility) Human Protein Atlas (http://www.proteinatlas.org) NCBI Entrez Protein database (http://www.ncbi.nlm.nih.gov/sites/entrez?db=protein) NCBI Protein Structure database (http://www.ncbi.nlm.nih.gov/sites/entrez?db=structure) Human Protein Reference Database (http://www.hprd.org/)

Protein Human Proteinpedia (http://www.humanproteinpedia.org/) Folding@Home (Stanford University) (http://folding.stanford.edu/)

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Tutorials and educational websites

"An Introduction to Proteins" (http://hopes.stanford.edu/basics/proteins/p0.html) from HOPES (Huntington's Disease Outreach Project for Education at Stanford) Proteins: Biogenesis to Degradation The Virtual Library of Biochemistry and Cell Biology (http://www. biochemweb.org/proteins.shtml)

Amino acid
Amino acids (pronounced/mino ..., mano ..., mno .../) are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen. They are particularly important in biochemistry, where the term usually refers to alpha-amino acids. An alpha-amino acid has the generic formula H2NCHRCOOH, where R is an organic substituent;[1] the amino group is attached to the carbon atom immediately adjacent to the carboxylate group The generic structure of an alpha amino acid in its (the carbon). Other types of amino acid exist when the amino unionized form group is attached to a different carbon atom; for example, in gamma-amino acids (such as gamma-amino-butyric acid) the carbon atom to which the amino group attaches is separated from the carboxylate group by two other carbon atoms. The various alpha-amino acids differ in which side-chain (R-group) is attached to their alpha carbon, and can vary in size from just one hydrogen atom in glycine to a large heterocyclic group in tryptophan. Amino acids are critical to life, and have many functions in metabolism. One particularly important function is to serve as the building blocks of proteins, which are linear chains of amino acids. Amino acids can be linked together in varying sequences to form a vast variety of proteins.[2] Twenty-two amino acids

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are naturally incorporated into polypeptides and are called proteinogenic or standard amino acids. Of these, 20 are encoded by the universal genetic code. Nine standard amino acids are called "essential" for humans because they cannot be created from other compounds by the human body, and so must be taken in as food. Due to their central role in biochemistry, amino acids are important in nutrition and are commonly used in nutrition supplements, fertilizers, food technology and industry. In industry, applications include the production of biodegradable plastics, drugs, and chiral catalysts.

The 21 amino acids found in eukaryotes, grouped according to their side-chains' pKas and charge at physiological pH 7.4

History
The first few amino acids were discovered in the early 19th century. In 1806, the French chemists Louis-Nicolas Vauquelin and Pierre Jean Robiquet isolated a compound in asparagus that proved to be asparagine, the first amino acid to be discovered.[3] [4] Another amino acid that was discovered in the early 19th century was cystine, in 1810,[5] although its monomer, cysteine, was discovered much later, in 1884.[4] [6] Glycine and leucine were also discovered around this time, in 1820.[7] Usage of the term amino acid in the English language is from 1898.[8]

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General structure
Further information: Proteinogenic amino acid In the structure shown at the top of the page, R represents a side-chain specific to each amino acid. The carbon atom next to the carboxyl group is called the carbon and amino acids with a side-chain bonded to this carbon are referred to as alpha amino acids. These are the most common form found in nature. In the alpha amino acids, the carbon is a chiral carbon atom, with the exception of glycine.[9] In amino acids that have a carbon chain attached to the carbon (such as lysine, shown to the right) the carbons are labeled in order as , , , , and so on.[10] In some amino acids, the amine group is attached to the or -carbon, and these are therefore referred to as beta or gamma amino acids. Amino acids are usually classified by the properties of their side-chain into four groups. The side-chain can make an amino acid a weak acid or a weak base, and a hydrophile if the side-chain is polar or a hydrophobe if it is nonpolar.[9] The chemical structures of the 22 standard amino acids, along with their chemical properties, are described more fully in the article on these proteinogenic amino acids.

Lysine with the carbon atoms in the side-chain labeled

The phrase "branched-chain amino acids" or BCAA refers to the amino acids having aliphatic side-chains that are non-linear; these are leucine, isoleucine, and valine. Proline is the only proteinogenic amino acid whose side-group links to the -amino group and, thus, is also the only proteinogenic amino acid containing a secondary amine at this position.[9] In chemical terms, proline is, therefore, an imino acid, since it lacks a primary amino group,[11] although it is still classed as an amino acid in the current biochemical nomenclature,[12] and may also be called an "N-alkylated alpha-amino acid".[13]

Isomerism
Of the standard -amino acids, all but glycine can exist in either of two optical isomers, called L or D amino acids, which are mirror images of each other (see also Chirality). While L-amino acids represent all of the amino acids found in proteins during translation in the ribosome, D-amino acids are found in some proteins produced by enzyme posttranslational modifications after translation and translocation to the endoplasmic reticulum, as in exotic sea-dwelling organisms such as The two optical isomers of alanine, D-Alanine cone snails.[14] They are also abundant components of the and L-Alanine peptidoglycan cell walls of bacteria,[15] and D-serine may act as a neurotransmitter in the brain.[16] The L and D convention for amino acid configuration refers not to the optical activity of the amino acid itself, but rather to the optical activity of the isomer of glyceraldehyde from which that amino acid can, in theory, be synthesized (D-glyceraldehyde is dextrorotary; L-glyceraldehyde is levorotary). In alternative fashion, the (S) and (R) designators are used to indicate the absolute stereochemistry. Almost all of the amino acids in proteins are (S) at the carbon, with cysteine being (R) and glycine non-chiral.[17] Cysteine is unusual since it has a sulfur atom at the second position in its side-chain, which has a larger atomic mass than the groups attached to the first carbon, which is attached to the -carbon in the other standard amino acids, thus the (R) instead of (S).

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Zwitterions
The amine and carboxylic acid functional groups found in amino acids allow them to have amphiprotic properties.[9] Carboxylic acid groups (-CO2H) can be deprotonated to become negative carboxylates (-CO2- ), and -amino groups (NH2-) can be protonated to become positive + -ammonium groups ( NH3-). At pH An amino acid in its (1) unionized and (2) zwitterionic forms values greater than the pKa of the carboxylic acid group (mean for the 20 common amino acids is about 2.2, see the table of amino acid structures above), the negative carboxylate ion predominates. At pH values lower than the pKa of the -ammonium group (mean for the 20 common -amino acids is about 9.4), the nitrogen is predominantly protonated as a positively charged -ammonium group. Thus, at pH between 2.2 and 9.4, the predominant form adopted by -amino acids contains a negative carboxylate and a positive -ammonium group, as shown in structure (2) on the right, so has net zero charge. This molecular state is known as a zwitterion, from the German Zwitter meaning hermaphrodite or hybrid.[18] Below pH 2.2, the predominant form will have a neutral carboxylic acid group and a positive -ammonium ion (net charge +1), and above pH 9.4, a negative carboxylate and neutral -amino group (net charge -1). The fully neutral form (structure (1) on the right) is a very minor species in aqueous solution throughout the pH range (less than 1 part in 107). Amino acids also exist as zwitterions in the solid phase, and crystallize with salt-like properties unlike typical organic acids or amines.

Isoelectric point
At pH values between the two pKa values, the zwitterion predominates, but coexists in dynamic equilibrium with small amounts of net negative and net positive ions. At the exact midpoint between the two pKa values, the trace amount of net negative and trace of net positive ions exactly balance, so that average net charge of all forms present is zero.[19] This pH is known as the isoelectric point pI, so pI = (pKa1 + pKa2). The individual amino acids all have slightly different pKa values, so have different isoelectric points. For amino acids with charged side-chains, the pKa of the side-chain is involved. Thus for Asp, Glu with negative side-chains, pI = (pKa1 + pKaR), where pKaR is the side-chain pKa. Cysteine also has potentially negative side-chain with pKaR = 8.14, so pI should be calculated as for Asp and Glu, even though the side-chain is not significantly charged at neutral pH. For His, Lys, and Arg with positive side-chains, pI = (pKaR + pKa2). Amino acids have zero mobility in electrophoresis at their isoelectric point, although this behaviour is more usually exploited for peptides and proteins than single amino acids. Zwitterions have minimum solubility at their isolectric point and some amino acids (in particular, with non-polar side-chains) can be isolated by precipitation from water by adjusting the pH to the required isoelectric point.

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Occurrence and functions in biochemistry

Standard amino acids
Amino acids are the structural units that make up proteins. They join together to form short polymer chains called peptides or longer chains called either polypeptides or proteins. These polymers are linear and unbranched, with each amino acid within the chain attached to two neighboring amino acids. The process of making proteins is called translation and involves the step-by-step addition of amino acids to a A polypeptide is an unbranched chain of amino acids. growing protein chain by a ribozyme that is [20] called a ribosome. The order in which the amino acids are added is read through the genetic code from an mRNA template, which is a RNA copy of one of the organism's genes. Twenty-two amino acids are naturally incorporated into polypeptides and are called proteinogenic or natural amino acids.[9] Of these, 20 are encoded by the universal genetic code. The remaining 2, selenocysteine and pyrrolysine, are incorporated into proteins by unique synthetic mechanisms. Selenocysteine is incorporated when the mRNA being translated includes a SECIS element, which causes the UGA codon to encode selenocysteine instead of a stop codon.[21] Pyrrolysine is used by some methanogenic archaea in enzymes that they use to produce methane. It is coded for with the codon UAG, which is normally a stop codon in other organisms.[22] This UAG codon is followed by a PYLIS downstream sequence.[23]

Non-standard amino acids

Aside from the 22 standard amino acids, there are many other amino acids that are called non-proteinogenic or non-standard. Those either are not found in proteins (for example carnitine, GABA), or are not produced directly and in isolation by standard cellular machinery (for example, hydroxyproline and selenomethionine). Non-standard amino acids that are found in proteins are formed by post-translational modification, which is modification after translation during protein synthesis. These modifications are often essential for the The amino acid selenocysteine function or regulation of a protein; for example, the carboxylation of glutamate allows for better binding of calcium cations,[24] and the hydroxylation of proline is critical for maintaining connective tissues.[25] Another example is the formation of hypusine in the translation initiation factor EIF5A, through modification of a lysine residue.[26] Such modifications can also determine the localization of the protein, e.g., the addition of long hydrophobic groups can cause a protein to bind to a phospholipid membrane.[27]

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Some nonstandard amino acids are not found in proteins. Examples include lanthionine, 2-aminoisobutyric acid, dehydroalanine, and the neurotransmitter gamma-aminobutyric acid. Nonstandard amino acids often occur as intermediates in the metabolic pathways for standard amino acids for example, ornithine and citrulline occur in the urea cycle, part of amino acid catabolism (see below).[28] A -alanine and its -alanine isomer rare exception to the dominance of -amino acids in biology is the -amino acid beta alanine (3-aminopropanoic acid), which is used in plants and microorganisms in the synthesis of pantothenic acid (vitamin B5), a component of coenzyme A.[29]

In human nutrition
Further information: Protein in nutrition and Amino acid synthesis When taken up into the human body from the diet, the 22 standard amino acids either are used to synthesize proteins and other biomolecules or are oxidized to urea and carbon dioxide as a source of energy.[30] The oxidation pathway starts with the removal of the amino group by a transaminase, the amino group is then fed into the urea cycle. The other product of transamidation is a keto acid that enters the citric acid cycle.[31] Glucogenic amino acids can also be converted into glucose, through gluconeogenesis.[32] Pyrrolysine trait is restricted to several microbes, and only one organism has both Pyl and Sec. Of the 22 standard amino acids, 9 are called essential amino acids because the human body cannot synthesize them from other compounds at the level needed for normal growth, so they must be obtained from food.[33] In addition, cysteine, taurine, tyrosine, histidine, and arginine are semiessential amino-acids in children, because the metabolic pathways that synthesize these amino acids are not fully developed.[34] [35] The amounts required also depend on the age and health of the individual, so it is hard to make general statements about the dietary requirement for some amino acids.
Essential Histidine Isoleucine Leucine Lysine Methionine Nonessential Alanine Arginine* Asparagine Aspartic acid Cysteine*

Phenylalanine Glutamic acid Threonine Tryptophan Valine Glutamine* Glycine Ornithine* Proline* Selenocysteine* Serine* Taurine* Tyrosine*

(*) Essential only in certain cases.[36] [37]

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Non-protein functions
Further information: Amino acid neurotransmitter In humans, non-protein amino acids also have important roles as metabolic intermediates, such as in the biosynthesis of the neurotransmitter gamma-aminobutyric acid. Many amino acids are used to synthesize other molecules, for example: Tryptophan is a precursor of the neurotransmitter serotonin.[38] Tyrosine is a precursor of the neurotransmitter dopamine. Glycine is a precursor of porphyrins such as heme.[39] Arginine is a precursor of nitric oxide.[40] Ornithine and S-adenosylmethionine are precursors of polyamines.[41] Aspartate, glycine, and glutamine are precursors of nucleotides.[42] Phenylalanine is a precursor of various phenylpropanoids, which are important in plant metabolism.

However, not all of the functions of other abundant non-standard amino acids are known. For example, taurine is a major amino acid in muscle and brain tissues, but, although many functions have been proposed, its precise role in the body has not been determined.[43] Some non-standard amino acids are used as defenses against herbivores in plants.[44] For example canavanine is an analogue of arginine that is found in many legumes,[45] and in particularly large amounts in Canavalia gladiata (sword bean).[46] This amino acid protects the plants from predators such as insects and can cause illness in people if some types of legumes are eaten without processing.[47] The non-protein amino acid mimosine is found in other species of legume, particularly Leucaena leucocephala.[48] This compound is an analogue of tyrosine and can poison animals that graze on these plants.

Uses in technology
Amino acids are used for a variety of applications in industry, but their main use is as additives to animal feed. This is necessary, since many of the bulk components of these feeds, such as soybeans, either have low levels or lack some of the essential amino acids: Lysine, methionine, threonine, and tryptophan are most important in the production of these feeds.[49] In this industry, amino acids are also used to chelate metal cations in order to improve the absorption of minerals from supplements, which may be required to improve the health or production of these animals.[50] The food industry is also a major consumer of amino acids, in particular, glutamic acid, which is used as a flavor enhancer,[51] and Aspartame (aspartyl-phenylalanine-1-methyl ester) as a low-calorie artificial sweetener.[52] Similar technology to that used for animal nutrition is employed in the human nutrition industry to alleviate symptoms of mineral deficiencies, such as anemia, by improving mineral absorption and reducing negative side effects from inorganic mineral supplementation. [53] The chelating ability of amino acids has been used in fertilizers for agriculture to facilitate the delivery of minerals to plants in order to correct mineral deficiencies, such as iron chlorosis. These fertilizers are also used to prevent deficiencies from occurring and improving the overall health of the plants.[54] The remaining production of amino acids is used in the synthesis of drugs and cosmetics.[49]

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Amino acid derivative 5-HTP (5-hydroxytryptophan) L-DOPA (L-dihydroxyphenylalanine) Eflornithine

Pharmaceutical application Experimental treatment for depression. Treatment for Parkinsonism. [56] [57] [55]

Drug that inhibits ornithine decarboxylase and is used in the treatment of sleeping sickness.

Expanded genetic code

Since 2001, 40 non-natural amino acids have been added into protein by creating a unique codon (recoding) and a corresponding transfer-RNA:aminoacyl tRNA-synthetase pair to encode it with diverse physicochemical and biological properties in order to be used as a tool to exploring protein structure and function or to create novel or enhanced proteins.[58] [59]

Chemical building blocks

Further information: Asymmetric synthesis Amino acids are important as low-cost feedstocks. These compounds are used in chiral pool synthesis as enantiomerically-pure building blocks.[60] Amino acids have been investigated as precursors chiral catalysts, e.g., for asymmetric hydrogenation reactions, although no commercial applications exist.[61]

Biodegradable plastics
Further information: Biodegradable plastics and Biopolymers Amino acids are under development as components of a range of biodegradable polymers. These materials have applications as environmentally friendly packaging and in medicine in drug delivery and the construction of prosthetic implants. These polymers include polypeptides, polyamides, polyesters, polysulfides, and polyurethanes with amino acids either forming part of their main chains or bonded as side-chains. These modifications alter the physical properties and reactivities of the polymers.[62] An interesting example of such materials is polyaspartate, a water-soluble biodegradable polymer that may have applications in disposable diapers and agriculture.[63] Due to its solubility and ability to chelate metal ions, polyaspartate is also being used as a biodegradeable anti-scaling agent and a corrosion inhibitor.[64] [65] In addition, the aromatic amino acid tyrosine is being developed as a possible replacement for toxic phenols such as bisphenol A in the manufacture of polycarbonates.[66]

Reactions
As amino acids have both a primary amine group and a primary carboxyl group, these chemicals can undergo most of the reactions associated with these functional groups. These include nucleophilic addition, amide bond formation and imine formation for the amine group and esterification, amide bond formation and decarboxylation for the carboxylic acid group.[67] The combination of these functional groups allow amino acids to be effective polydentate ligands for metal-amino acid chelates.[68] The multiple side-chains of amino acids can also undergo chemical reactions.[69] The types of these reactions are determined by the groups on these side-chains and are, therefore, different between the various types of amino acid.

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Chemical synthesis
Several methods exist to synthesize amino acids. One of the oldest methods begins with the bromination at the -carbon of a carboxylic acid. The Strecker amino acid synthesis Nucleophilic substitution with ammonia then converts the alkyl bromide to the amino acid.[70] In alternative fashion, the Strecker amino acid synthesis involves the treatment of an aldehyde with potassium cyanide and ammonia, this produces an -amino nitrile as an intermediate. Hydrolysis of the nitrile in acid then yields a -amino acid.[71] Using ammonia or ammonium salts in this reaction gives unsubstituted amino acids, while substituting primary and secondary amines will yield substituted amino acids.[72] Likewise, using ketones, instead of aldehydes, gives ,-disubstituted amino [73] acids. The classical synthesis gives racemic mixtures of -amino acids as products, but several alternative procedures using asymmetric auxiliaries [74] or asymmetric catalysts [75] [76] have been developed.[77] At the current time, the most-adopted method is an automated synthesis on a solid support (e.g., polystyrene beads), using protecting groups (e.g., Fmoc and t-Boc) and activating groups (e.g., DCC and DIC).

Peptide bond formation

As both the amine and carboxylic acid groups of amino acids can react to form amide bonds, one amino acid molecule can react with another and become joined through an amide linkage. This polymerization of amino acids is what creates proteins. This condensation reaction yields the newly formed peptide bond and a molecule of water. In cells, this reaction does not occur directly; instead the amino acid is first activated by attachment to a transfer RNA molecule through an ester bond. This aminoacyl-tRNA is produced in an ATP-dependent reaction carried out by an aminoacyl tRNA synthetase.[78] This The condensation of two amino acids to form a peptide bond aminoacyl-tRNA is then a substrate for the ribosome, which catalyzes the attack of the amino group of the elongating protein chain on the ester bond.[79] As a result of this mechanism, all proteins made by ribosomes are synthesized starting at their N-terminus and moving towards their C-terminus. However, not all peptide bonds are formed in this way. In a few cases, peptides are synthesized by specific enzymes. For example, the tripeptide glutathione is an essential part of the defenses of cells against oxidative stress. This peptide is synthesized in two steps from free amino acids.[80] In the first step gamma-glutamylcysteine synthetase condenses cysteine and glutamic acid through a peptide bond formed between the side-chain carboxyl of the glutamate (the gamma carbon of this side-chain) and the amino group of the cysteine. This dipeptide is then condensed with glycine by glutathione synthetase to form glutathione.[81]

Amino acid In chemistry, peptides are synthesized by a variety of reactions. One of the most-used in solid-phase peptide synthesis uses the aromatic oxime derivatives of amino acids as activated units. These are added in sequence onto the growing peptide chain, which is attached to a solid resin support.[82] The ability to easily synthesize vast numbers of different peptides by varying the types and order of amino acids (using combinatorial chemistry) has made peptide synthesis particularly important in creating libraries of peptides for use in drug discovery through high-throughput screening.[83]

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Biosynthesis
In plants, nitrogen is first assimilated into organic compounds in the form of glutamate, formed from alpha-ketoglutarate and ammonia in the mitochondrion. In order to form other amino acids, the plant uses transaminases to move the amino group to another alpha-keto carboxylic acid. For example, aspartate aminotransferase converts glutamate and oxaloacetate to alpha-ketoglutarate and aspartate.[84] Other organisms use transaminases for amino acid synthesis, too. Nonstandard amino acids are usually formed through modifications to standard amino acids. For example, homocysteine is formed through the transsulfuration pathway or by the demethylation of methionine via the intermediate metabolite S-adenosyl methionine,[43] while hydroxyproline is made by a posttranslational modification of proline.[85] Microorganisms and plants can synthesize many uncommon amino acids. For example, some microbes make 2-aminoisobutyric acid and lanthionine, which is a sulfide-bridged derivative of alanine. Both of these amino acids are found in peptidic lantibiotics such as alamethicin.[86] While in plants, 1-aminocyclopropane-1-carboxylic acid is a small disubstituted cyclic amino acid that is a key intermediate in the production of the plant hormone ethylene.[87]

Catabolism
Degradation of an amino acid often involves deamination by moving its amino group to alpha-ketoglutarate, forming glutamate. This process involves transaminases, often the same as those used in amination during synthesis. In many vertebrates, the amino group is then removed through the urea cycle and is excreted in the form of urea. However, amino acid degradation can produce uric acid or ammonia instead. For example, serine dehydratase converts serine to pyruvate and ammonia.[89]
Catabolism of proteinogenic amino acids. Amino acids can be classified according [88] to the properties of their main products as either of the following: Glucogenic, with the products having the ability to form glucose by gluconeogenesisKetogenic, with the products not having the ability to form glucose. These products may still be used for ketogenesis or lipid synthesis.Amino acids catabolized into both glucogenic and ketogenic products.

Physicochemical properties of amino acids

The 20 amino acids encoded directly by the genetic code can be divided into several groups based on their properties. Important factors are charge, hydrophilicity or hydrophobicity, size, and functional groups.[9] These properties are important for protein structure and proteinprotein interactions. The water-soluble proteins tend to have their hydrophobic residues (Leu, Ile, Val, Phe, and Trp) buried in the middle of the protein, whereas hydrophilic side-chains are exposed to the aqueous solvent. The integral membrane proteins tend to have outer rings of exposed hydrophobic amino acids that anchor them into the lipid bilayer. In the case part-way between

Amino acid these two extremes, some peripheral membrane proteins have a patch of hydrophobic amino acids on their surface that locks onto the membrane. In similar fashion, proteins that have to bind to positively-charged molecules have surfaces rich with negatively charged amino acids like glutamate and aspartate, while proteins binding to negatively-charged molecules have surfaces rich with positively charged chains like lysine and arginine. There are different hydrophobicity scales of amino acid residues.[90] Some amino acids have special properties such as cysteine, that can form covalent disulfide bonds to other cysteine residues, proline that forms a cycle to the polypeptide backbone, and glycine that is more flexible than other amino acids. Many proteins undergo a range of posttranslational modifications, when additional chemical groups are attached to the amino acids in proteins. Some modifications can produce hydrophobic lipoproteins,[91] or hydrophilic glycoproteins.[92] These type of modification allow the reversible targeting of a protein to a membrane. For example, the addition and removal of the fatty acid palmitic acid to cysteine residues in some signaling proteins causes the proteins to attach and then detach from cell membranes.[93]

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Table of standard amino acid abbreviations and properties

Amino Acid 3-Letter [94] 1-Letter [94] Side-chain [94] polarity nonpolar polar polar polar polar polar polar nonpolar polar Side-chain charge [94] (pH 7.4) neutral positive neutral negative neutral negative neutral neutral positive(10%) neutral(90%) neutral neutral positive neutral neutral neutral neutral neutral neutral neutral neutral Hydropathy [95] index 1.8 4.5 3.5 3.5 2.5 3.5 3.5 0.4 3.2 211 5.9 250 0.3 Absorbance [96] max(nm) at max (x103 [96] M1 cm1)

Alanine Arginine Asparagine Aspartic acid Cysteine Glutamic acid Glutamine Glycine Histidine

Ala Arg Asn Asp Cys Glu Gln Gly His

A R N D C E Q G H

Isoleucine Leucine Lysine Methionine Phenylalanine Proline Serine Threonine Tryptophan Tyrosine Valine

Ile Leu Lys Met Phe Pro Ser Thr Trp Tyr Val

I L K M F P S T W Y V

nonpolar nonpolar polar nonpolar nonpolar nonpolar polar polar nonpolar polar nonpolar

4.5 3.8 3.9 1.9 2.8 1.6 0.8 0.7 0.9 1.3 4.2 280, 219 274, 222, 193 5.6, 47.0 1.4, 8.0, 48.0 257, 206, 188 0.2, 9.3, 60.0

In addition, there are two additional amino acids that are incorporated by overriding stop codons:

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21st and 22nd amino acids Selenocysteine Pyrrolysine

3-Letter Sec Pyl

1-Letter U O

In addition to the specific amino acid codes, placeholders are used in cases where chemical or crystallographic analysis of a peptide or protein cannot conclusively determine the identity of a residue.
Ambiguous Amino Acids Asparagine or aspartic acid Glutamine or glutamic acid Leucine or Isoleucine Unspecified or unknown amino acid 3-Letter Asx Glx Xle Xaa 1-Letter B Z J X

Unk is sometimes used instead of Xaa, but is less standard. In addition, many non-standard amino acids have a specific code. For example, several peptide drugs, such as Bortezomib and MG132, are artificially synthesized and retain their protecting groups, which have specific codes. Bortezomib is Pyz-Phe-boroLeu, and MG132 is Z-Leu-Leu-Leu-al. To aid in the analysis of protein structure, photocrosslinking amino acid analogues are available. These include photoleucine (pLeu) and photomethionine (pMet).[97]

References and notes

[1] Proline is an exception to this general formula. It lacks the NH2 group because of the cyclization of the side-chain and is known as an imino acid; it falls under the category of special structured amino acids. [2] "The Structures of Life" (http:/ / publications. nigms. nih. gov/ structlife/ chapter1. html). National Institute of General Medical Sciences. . Retrieved 2008-05-20. [3] Vauquelin LN, Robiquet PJ (1806). "The discovery of a new plant principle in Asparagus sativus". Annales de Chimie 57: 8893. [4] Anfinsen CB, Edsall JT, Richards FM (1972). Advances in Protein Chemistry. New York: Academic Press. pp.99, 103. ISBN978-0-12-034226-6. [5] Wollaston WH (1810). "On cystic oxide, a new species of urinary calculus". Philosophical Transactions of the Royal Society of London 100 (0): 22330. doi:10.1098/rstl.1810.0015. [6] Baumann E (1884). "ber cystin und cystein" (http:/ / vlp. mpiwg-berlin. mpg. de/ library/ data/ lit16533). Z Physiol Chemie 8 (4): 299305. . Retrieved 28 March 2011. [7] Braconnot HM (1820). "Sur la conversion des matires animales en nouvelles substances par le moyen de l'acide sulfurique". Ann Chim Phys Ser 2 13: 11325. [8] "etymonline.com entry for amino" (http:/ / www. etymonline. com/ index. php?term=amino). www.etymonline.com. . Retrieved 2010-07-19. [9] Creighton, Thomas H. (1993). "Chapter 1". Proteins: structures and molecular properties. San Francisco: W. H. Freeman. ISBN978-0-7167-7030-5. [10] "Nomenclature and Symbolism for Amino Acids and Peptides" (http:/ / www. chem. qmul. ac. uk/ iupac/ AminoAcid/ AA1n2. html). IUPAC-IUB Joint Commission on Biochemical Nomenclature. 1983. . Retrieved 2008-11-17. [11] Jodidi, S. L. (1926-03-01). "The Formol Titration of Certain Amino Acids". Journal of the American Chemical Society 48 (3): 751753. doi:10.1021/ja01414a033. [12] Liebecq, Claude, ed (1992). Biochemical Nomenclature and Related Documents (2nd ed.). Portland Press. pp.3969. ISBN978-1-85578-005-7. [13] Smith, Anthony D. (1997). Oxford dictionary of biochemistry and molecular biology. Oxford: Oxford University Press. pp.535. ISBN978-0-19-854768-6. OCLC37616711. [14] Pisarewicz K, Mora D, Pflueger FC, Fields GB, Mar F (May 2005). "Polypeptide chains containing D-gamma-hydroxyvaline". Journal of the American Chemical Society 127 (17): 620715. doi:10.1021/ja050088m. PMID15853325. [15] van Heijenoort J (March 2001). "Formation of the glycan chains in the synthesis of bacterial peptidoglycan". Glycobiology 11 (3): 25R36R. doi:10.1093/glycob/11.3.25R. PMID11320055. [16] Wolosker H, Dumin E, Balan L, Foltyn VN (July 2008). "D-amino acids in the brain: D-serine in neurotransmission and neurodegeneration". The FEBS Journal 275 (14): 351426. doi:10.1111/j.1742-4658.2008.06515.x. PMID18564180.

Amino acid
[17] Hatem, Salama Mohamed Ali (2006). "Gas chromatographic determination of Amino Acid Enantiomers in tobacco and bottled wines" (http:/ / geb. uni-giessen. de/ geb/ volltexte/ 2006/ 3038/ index. html). University of Giessen. . Retrieved 2008-11-17. [18] Simmons, William J.; Gerhard Meisenberg (2006). Principles of medical biochemistry. Mosby Elsevier. p.19. ISBN0-323-02942-6. [19] Fennema OR. Food Chemistry 3rd Ed. CRC Press. pp.3278. ISBN0-8247-9691-8. [20] Rodnina MV, Beringer M, Wintermeyer W (January 2007). "How ribosomes make peptide bonds". Trends in Biochemical Sciences 32 (1): 206. doi:10.1016/j.tibs.2006.11.007. PMID17157507. [21] Driscoll DM, Copeland PR (2003). "Mechanism and regulation of selenoprotein synthesis". Annual Review of Nutrition 23 (1): 1740. doi:10.1146/annurev.nutr.23.011702.073318. PMID12524431. [22] Krzycki JA (December 2005). "The direct genetic encoding of pyrrolysine". Current Opinion in Microbiology 8 (6): 70612. doi:10.1016/j.mib.2005.10.009. PMID16256420. [23] Thobald-Dietrich A, Gieg R, Rudinger-Thirion J (2005). "Evidence for the existence in mRNAs of a hairpin element responsible for ribosome dependent pyrrolysine insertion into proteins". Biochimie 87 (910): 8137. doi:10.1016/j.biochi.2005.03.006. PMID16164991. [24] Vermeer C (March 1990). "Gamma-carboxyglutamate-containing proteins and the vitamin K-dependent carboxylase". The Biochemical Journal 266 (3): 62536. PMC1131186. PMID2183788. [25] Bhattacharjee A, Bansal M (March 2005). "Collagen structure: the Madras triple helix and the current scenario". IUBMB Life 57 (3): 16172. doi:10.1080/15216540500090710. PMID16036578. [26] Park MH (February 2006). "The post-translational synthesis of a polyamine-derived amino acid, hypusine, in the eukaryotic translation initiation factor 5A (eIF5A)". Journal of Biochemistry 139 (2): 1619. doi:10.1093/jb/mvj034. PMC2494880. PMID16452303. [27] Blenis J, Resh MD (December 1993). "Subcellular localization specified by protein acylation and phosphorylation". Current Opinion in Cell Biology 5 (6): 9849. doi:10.1016/0955-0674(93)90081-Z. PMID8129952. [28] Curis E, Nicolis I, Moinard C, et al. (November 2005). "Almost all about citrulline in mammals". Amino Acids 29 (3): 177205. doi:10.1007/s00726-005-0235-4. PMID16082501. [29] Coxon KM, Chakauya E, Ottenhof HH, et al. (August 2005). "Pantothenate biosynthesis in higher plants". Biochemical Society Transactions 33 (Pt 4): 7436. doi:10.1042/BST0330743. PMID16042590. [30] Sakami W, Harrington H (1963). "Amino acid metabolism". Annual Review of Biochemistry 32 (1): 35598. doi:10.1146/annurev.bi.32.070163.002035. PMID14144484. [31] Brosnan JT (April 2000). "Glutamate, at the interface between amino acid and carbohydrate metabolism" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=10736367). The Journal of Nutrition 130 (4S Suppl): 988S90S. PMID10736367. . [32] Young VR, Ajami AM (September 2001). "Glutamine: the emperor or his clothes?" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=11533293). The Journal of Nutrition 131 (9 Suppl): 2449S59S; discussion 2486S7S. PMID11533293. . [33] Young VR (August 1994). "Adult amino acid requirements: the case for a major revision in current recommendations" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=8064412). The Journal of Nutrition 124 (8 Suppl): 1517S1523S. PMID8064412. . [34] Imura K, Okada A (January 1998). "Amino acid metabolism in pediatric patients". Nutrition 14 (1): 1438. doi:10.1016/S0899-9007(97)00230-X. PMID9437700. [35] Loureno R, Camilo ME (2002). "Taurine: a conditionally essential amino acid in humans? An overview in health and disease". Nutricin Hospitalaria 17 (6): 26270. PMID12514918. [36] Frst P, Stehle P (June 2004). "What are the essential elements needed for the determination of amino acid requirements in humans?" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=15173430). The Journal of Nutrition 134 (6 Suppl): 1558S1565S. PMID15173430. . [37] Reeds PJ (July 2000). "Dispensable and indispensable amino acids for humans" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=10867060). The Journal of Nutrition 130 (7): 1835S40S. PMID10867060. . [38] Savelieva KV, Zhao S, Pogorelov VM, et al. (2008). Bartolomucci, Alessandro. ed. "Genetic disruption of both tryptophan hydroxylase genes dramatically reduces serotonin and affects behavior in models sensitive to antidepressants". PloS ONE 3 (10): e3301. Bibcode2008PLoSO...3.3301S. doi:10.1371/journal.pone.0003301. PMC2565062. PMID18923670. [39] Shemin D, Rittenberg D (1 December 1946). "The biological utilization of glycine for the synthesis of the protoporphyrin of hemoglobin" (http:/ / www. jbc. org/ cgi/ reprint/ 166/ 2/ 621). Journal of Biological Chemistry 166 (2): 6215. PMID20276176. . [40] Tejero J, Biswas A, Wang ZQ, et al. (November 2008). "Stabilization and characterization of a heme-oxy reaction intermediate in inducible nitric-oxide synthase". The Journal of Biological Chemistry 283 (48): 33498507. doi:10.1074/jbc.M806122200. PMC2586280. PMID18815130. [41] Rodrguez-Caso C, Montaez R, Cascante M, Snchez-Jimnez F, Medina MA (August 2006). "Mathematical modeling of polyamine metabolism in mammals". The Journal of Biological Chemistry 281 (31): 21799812. doi:10.1074/jbc.M602756200. PMID16709566. [42] Stryer, Lubert; Berg, Jeremy Mark; Tymoczko, John L. (2002). Biochemistry. San Francisco: W.H. Freeman. pp.6938. ISBN0-7167-4684-0. [43] Brosnan JT, Brosnan ME (June 2006). "The sulfur-containing amino acids: an overview" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=16702333). The Journal of Nutrition 136 (6 Suppl): 1636S1640S. PMID16702333. . [44] Hylin, John W. (1969). "Toxic peptides and amino acids in foods and feeds". Journal of Agricultural and Food Chemistry 17 (3): 4926. doi:10.1021/jf60163a003. [45] Turner, B. L.; Harborne, J. B. (1967). "Distribution of canavanine in the plant kingdom". Phytochemistry 6 (6): 86366. doi:10.1016/S0031-9422(00)86033-1.

105

Amino acid
[46] Ekanayake S, Skog K, Asp NG (May 2007). "Canavanine content in sword beans (Canavalia gladiata): analysis and effect of processing". Food and Chemical Toxicology 45 (5): 797803. doi:10.1016/j.fct.2006.10.030. PMID17187914. [47] Rosenthal GA (2001). "L-Canavanine: a higher plant insecticidal allelochemical". Amino Acids 21 (3): 31930. doi:10.1007/s007260170017. PMID11764412. [48] Hammond AC (May 1995). "Leucaena toxicosis and its control in ruminants" (http:/ / jas. fass. org/ cgi/ pmidlookup?view=long& pmid=7665380). Journal of Animal Science 73 (5): 148792. PMID7665380. . [49] Leuchtenberger W, Huthmacher K, Drauz K (November 2005). "Biotechnological production of amino acids and derivatives: current status and prospects". Applied Microbiology and Biotechnology 69 (1): 18. doi:10.1007/s00253-005-0155-y. PMID16195792. [50] Ashmead, H. DeWayne (1993). The Role of Amino Acid Chelates in Animal Nutrition. Westwood: Noyes Publications. [51] Garattini S (April 2000). "Glutamic acid, twenty years later" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=10736350). The Journal of Nutrition 130 (4S Suppl): 901S9S. PMID10736350. . [52] Stegink LD (July 1987). "The aspartame story: a model for the clinical testing of a food additive" (http:/ / www. ajcn. org/ cgi/ pmidlookup?view=long& pmid=3300262). The American Journal of Clinical Nutrition 46 (1 Suppl): 20415. PMID3300262. . [53] Albion Laboratories, Inc.. "Albion Ferrochel Website" (http:/ / www. albionferrochel. com). . Retrieved Cited: July 12, 2011. [54] Ashmead, H. DeWayne (1986). Foliar Feeding of Plants with Amino Acid Chelates. Park Ridge: Noyes Publications. [55] Turner EH, Loftis JM, Blackwell AD (March 2006). "Serotonin a la carte: supplementation with the serotonin precursor 5-hydroxytryptophan". Pharmacology & Therapeutics 109 (3): 32538. doi:10.1016/j.pharmthera.2005.06.004. PMID16023217. [56] Kostrzewa RM, Nowak P, Kostrzewa JP, Kostrzewa RA, Brus R (March 2005). "Peculiarities of L: -DOPA treatment of Parkinson's disease". Amino Acids 28 (2): 15764. doi:10.1007/s00726-005-0162-4. PMID15750845. [57] Heby O, Persson L, Rentala M (August 2007). "Targeting the polyamine biosynthetic enzymes: a promising approach to therapy of African sleeping sickness, Chagas' disease, and leishmaniasis". Amino Acids 33 (2): 35966. doi:10.1007/s00726-007-0537-9. PMID17610127. [58] Xie J, Schultz PG (December 2005). "Adding amino acids to the genetic repertoire". Curr Opin Chem Biol 9 (6): 54854. doi:10.1016/j.cbpa.2005.10.011. PMID16260173. [59] Wang Q, Parrish AR, Wang L (March 2009). "Expanding the genetic code for biological studies". Chem. Biol. 16 (3): 32336. doi:10.1016/j.chembiol.2009.03.001. PMC2696486. PMID19318213. [60] Hanessian, S. (1993). "Reflections on the total synthesis of natural products: Art, craft, logic, and the chiron approach". Pure and Applied Chemistry 65 (6): 1189204. doi:10.1351/pac199365061189. [61] Blaser, Hans Ulrich (1992). "The chiral pool as a source of enantioselective catalysts and auxiliaries". Chemical Reviews 92 (5): 93552. doi:10.1021/cr00013a009. [62] Sanda, Fumio; Endo, Takeshi (1999). "Feature Article Syntheses and functions of polymers based on amino acids". Macromolecular Chemistry and Physics 200 (12): 265161. doi:10.1002/(SICI)1521-3935(19991201)200:12<2651::AID-MACP2651>3.0.CO;2-P. [63] Gross, R. A.; Kalra, B. (2002). "Biodegradable Polymers for the Environment" (http:/ / www. sciencemag. org/ cgi/ content/ abstract/ 297/ 5582/ 803). Science 297 (5582): 803807. Bibcode2002Sci...297..803G. doi:10.1126/science.297.5582.803. PMID12161646. . [64] Low, K. C.; Wheeler, A. P.; Koskan, L. P. (1996). Commercial poly(aspartic acid) and Its Uses. Advances in Chemistry Series. 248. Washington, D.C.: American Chemical Society. [65] Thombre, S.M.; Sarwade, B.D. (2005). "Synthesis and Biodegradability of Polyaspartic Acid: A Critical Review" (http:/ / www. informaworld. com/ index/ 718581646. pdf). Journal of Macromolecular Science, Part A 42 (9): 12991315. doi:10.1080/10601320500189604. . [66] Bourke, S. L.; Kohn, J. (2003). "Polymers derived from the amino acid l-tyrosine: polycarbonates, polyarylates and copolymers with poly(ethylene glycol)" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0169409X03000383). Advanced Drug Delivery Reviews 55 (4): 447466. doi:10.1016/S0169-409X(03)00038-3. PMID12706045. . [67] Elmore, Donald Trevor; Barrett, G. C. (1998). Amino acids and peptides. Cambridge, UK: Cambridge University Press. pp.4860. ISBN0-521-46827-2. [68] Konar, Sanjit; et al. (2010). "Structural determination and characterization of copper and zinc bis-glycinates with X-ray crystallography and mass spectrometry". Journal of Coordination Chemistry 63 (19). [69] Gutteridge A, Thornton JM (November 2005). "Understanding nature's catalytic toolkit". Trends in Biochemical Sciences 30 (11): 6229. doi:10.1016/j.tibs.2005.09.006. PMID16214343. [70] McMurry, John (1996). Organic chemistry. Pacific Grove, CA, USA: Brooks/Cole. p.1064. ISBN0-534-23832-7. [71] Strecker, Adolph (1850). "Ueber die knstliche Bildung der Milchsure und einen neuen, dem Glycocoll homologen Krper". Justus Liebigs Annalen der Chemie 75 (1): 2745. doi:10.1002/jlac.18500750103. [72] Strecker, Adolph (1854). "Ueber einen neuen aus Aldehyd - Ammoniak und Blausure entstehenden Krper". Justus Liebigs Annalen der Chemie 91 (3): 34951. doi:10.1002/jlac.18540910309. [73] Masumoto S, Usuda H, Suzuki M, Kanai M, Shibasaki M (May 2003). "Catalytic enantioselective Strecker reaction of ketoimines". Journal of the American Chemical Society 125 (19): 56345. doi:10.1021/ja034980. PMID12733893. [74] Davis, F. A.; Reddy, Rajarathnam E.; Portonovo, Padma S. (1994). "Asymmetric strecker synthesis using enantiopure sulfinimines: A convenient synthesis of -amino acids". Tetrahedron Letters 35 (50): 9351. doi:10.1016/S0040-4039(00)78540-6. [75] Ishitani, Haruro; Komiyama, Susumu; Hasegawa, Yoshiki; Kobayashi, Sh (2000). "Catalytic Asymmetric Strecker Synthesis. Preparation of Enantiomerically Pure -Amino Acid Derivatives from Aldimines and Tributyltin Cyanide or Achiral Aldehydes, Amines, and Hydrogen Cyanide Using a Chiral Zirconium Catalyst". Journal of the American Chemical Society 122 (5): 7626. doi:10.1021/ja9935207.

106

Amino acid
[76] Huang, Jinkun; Corey, E. J. (2004). "A New Chiral Catalyst for the Enantioselective Strecker Synthesis of -Amino Acids". Orgic Letters 62 (6): 50279. doi:10.1021/ol047698w. PMID15606127. [77] Duthaler, Rudolf O. (1994). "Recent developments in the stereoselective synthesis of -aminoacids". Tetrahedron 50 (6): 15391650. doi:10.1016/S0040-4020(01)80840-1. [78] Ibba M, Sll D (May 2001). "The renaissance of aminoacyl-tRNA synthesis" (http:/ / www. nature. com/ embor/ journal/ v2/ n5/ full/ embor420. html). EMBO Reports 2 (5): 3827. doi:10.1093/embo-reports/kve095 (inactive 2010-02-18). PMC1083889. PMID11375928. . [79] Lengyel P, Sll D (June 1969). "Mechanism of protein biosynthesis" (http:/ / mmbr. asm. org/ cgi/ pmidlookup?view=long& pmid=4896351). Bacteriological Reviews 33 (2): 264301. PMC378322. PMID4896351. . [80] Wu G, Fang YZ, Yang S, Lupton JR, Turner ND (March 2004). "Glutathione metabolism and its implications for health" (http:/ / jn. nutrition. org/ cgi/ pmidlookup?view=long& pmid=14988435). The Journal of Nutrition 134 (3): 48992. PMID14988435. . [81] Meister A (November 1988). "Glutathione metabolism and its selective modification" (http:/ / www. jbc. org/ cgi/ pmidlookup?view=long& pmid=3053703). The Journal of Biological Chemistry 263 (33): 172058. PMID3053703. . [82] Carpino, Louis A. (1992). "1-Hydroxy-7-azabenzotriazole. An efficient peptide coupling additive". Journal of the American Chemical Society 115 (10): 43978. doi:10.1021/ja00063a082. [83] Marasco D, Perretta G, Sabatella M, Ruvo M (October 2008). "Past and future perspectives of synthetic peptide libraries". Current Protein & Peptide Science 9 (5): 44767. doi:10.2174/138920308785915209. PMID18855697. [84] Jones, Russell Celyn; Buchanan, Bob B.; Gruissem, Wilhelm (2000). Biochemistry & molecular biology of plants. Rockville, Md: American Society of Plant Physiologists. pp.3712. ISBN0-943088-39-9. [85] Kivirikko KI, Pihlajaniemi T (1998). "Collagen hydroxylases and the protein disulfide isomerase subunit of prolyl 4-hydroxylases". Advances in Enzymology and Related Areas of Molecular Biology 72: 32598. PMID9559057. [86] Whitmore L, Wallace BA (May 2004). "Analysis of peptaibol sequence composition: implications for in vivo synthesis and channel formation". European Biophysics Journal 33 (3): 2337. doi:10.1007/s00249-003-0348-1. PMID14534753. [87] Alexander L, Grierson D (October 2002). "Ethylene biosynthesis and action in tomato: a model for climacteric fruit ripening". Journal of Experimental Botany 53 (377): 203955. doi:10.1093/jxb/erf072. PMID12324528. [88] Chapter 20 (Amino Acid Degradation and Synthesis) in: Denise R., PhD. Ferrier. Lippincott's Illustrated Reviews: Biochemistry (Lippincott's Illustrated Reviews). Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-2265-9. [89] Stryer, Lubert; Berg, Jeremy Mark; Tymoczko, John L. (2002). Biochemistry. San Francisco: W.H. Freeman. pp.63949. ISBN0-7167-4684-0. [90] Urry, Dan W. (2004). "The change in Gibbs free energy for hydrophobic association: Derivation and evaluation by means of inverse temperature transitions". Chemical Physics Letters 399 (13): 17783. doi:10.1016/S0009-2614(04)01565-9. [91] Magee T, Seabra MC (April 2005). "Fatty acylation and prenylation of proteins: what's hot in fat". Current Opinion in Cell Biology 17 (2): 1906. doi:10.1016/j.ceb.2005.02.003. PMID15780596. [92] Pilobello KT, Mahal LK (June 2007). "Deciphering the glycocode: the complexity and analytical challenge of glycomics". Current Opinion in Chemical Biology 11 (3): 3005. doi:10.1016/j.cbpa.2007.05.002. PMID17500024. [93] Smotrys JE, Linder ME (2004). "Palmitoylation of intracellular signaling proteins: regulation and function". Annual Review of Biochemistry 73 (1): 55987. doi:10.1146/annurev.biochem.73.011303.073954. PMID15189153. [94] Hausman, Robert E.; Cooper, Geoffrey M. (2004). The cell: a molecular approach. Washington, D.C: ASM Press. p.51. ISBN0-87893-214-3. [95] Kyte J, Doolittle RF (May 1982). "A simple method for displaying the hydropathic character of a protein". Journal of Molecular Biology 157 (1): 10532. doi:10.1016/0022-2836(82)90515-0. PMID7108955. [96] Freifelder, D. (1983). Physical Biochemistry (2nd ed.). W. H. Freeman and Company. ISBN0-7167-1315-2. [97] Suchanek M, Radzikowska A, Thiele C (April 2005). "Photo-leucine and photo-methionine allow identification of protein-protein interactions in living cells". Nature Methods 2 (4): 2617. doi:10.1038/nmeth752. PMID15782218.

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Further reading
Doolittle, R.F. (1989) Redundancies in protein sequences. In Predictions of Protein Structure and the Principles of Protein Conformation (Fasman, G.D. ed) Plenum Press, New York, pp.599623 David L. Nelson and Michael M. Cox, Lehninger Principles of Biochemistry, 3rd edition, 2000, Worth Publishers, ISBN 1-57259-153-6 Meierhenrich, U.J.: Amino acids and the asymmetry of life, Springer-Verlag, Berlin, New York, 2008. ISBN 978-3-540-76885-2 Morelli, Robert J. "Studies of amino acid absorption from the small intestine." San Francisco: Morelli, 1952.

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External links
Amino acids overview (http://www.peptideguide.com/amino-acids/index.html) physical-chemistry properties, 3D structures, etc. List of Standard Amino Acids (http://www.unc.edu/~bzafer/aminoacids/) The Detailed PDF List of Standard Amino Acids (including 3D depictions) Nomenclature and Symbolism for Amino Acids and Peptides (http://www.chem.qmul.ac.uk/iupac/ AminoAcid/) IUPAC-IUB Joint Commission on Biochemical Nomenclature (JCBN) Molecular Expressions: The Amino Acid Collection (http://micro.magnet.fsu.edu/aminoacids/index.html) Has detailed information and microscopy photographs of each amino acid. Amino acid properties (http://www.russell.embl.de/aas/) Properties of the amino acids (a tool aimed mostly at molecular geneticists trying to understand the meaning of mutations) Synthesis of Amino Acids and Derivatives (http://www.organic-chemistry.org/synthesis/C1C/nitrogen/ alpha-amino-acids2.shtm) Learn the 20 proteinogenic amino acids online (http://www.mathiasbader.de/studium/biology/index. php?lng=en) The origin of the single-letter code for the amino acids (http://www.biology.arizona.edu/biochemistry/ problem_sets/aa/Dayhoff.html) Amino acid solutions pH, titration and isoelectric point calculation free spreadsheet (http://www2.iq.usp.br/ docente/gutz/Curtipot_.html) Interactive Amino Map Web Application at DNA.UTAH.EDU (http://www.dna.utah.edu/utensils/amino. php)

Properties of the twenty amino acids

Proteinogenic amino acids are those amino acids that can be found in proteins and require cellular machinery coded for in the genetic code [1] of any organism for their isolated production. There are 22 standard amino acids, but only 21 are found in eukaryotes. Of the 22, 20 are directly encoded by the universal genetic code. Humans can synthesize 11 of these 20 from each other or from other molecules of intermediary metabolism. The other 9 must be consumed in the diet, and so are called essential amino acids; those are histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine. The remaining two, selenocysteine and pyrrolysine, are incorporated into proteins by unique synthetic mechanisms. The word proteinogenic means "protein building". Proteinogenic amino acids can be assembled into a polypeptide (the subunit of a protein) through a process called translation (the second stage of protein biosynthesis, part of the overall process of gene expression). In contrast, non-proteinogenic amino acids are either not found in proteins (like carnitine, GABA, or L-DOPA), or are not produced directly and in isolation by standard cellular machinery (like hydroxyproline and selenomethionine). The latter often results from posttranslational modification of proteins. There are clear reasons why organisms have not evolved to incorporate certain non-proteinogenic amino acids into proteins: for example, ornithine and homoserine cyclize against the peptide backbone and fragment the protein with relatively short half-lives, while others are toxic because they can be mistakenly incorporated into proteins, such as the arginine analog canavanine. Non-proteinogenic amino acids are found in nonribosomal peptides, which are not produced by the ribosome during translation.

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Structures
The following illustrates the structures and abbreviations of the 21 amino acids that are directly encoded for protein synthesis by the genetic code of eukaryotes. The structures given below are standard chemical structures, not the typical zwitterion forms that exist in aqueous solutions.

Grouped table of 21 amino acids' structures, nomenclature, and their side groups' pKa's.

L-Alanine (Ala/A)

L-Arginine (Arg/R)

L-Asparagine (Asn/N)

L-Aspartic acid (Asp/D)

L-Cysteine (Cys/C)

L-Glutamic acid (Glu/E)

L-Glutamine (Gln/Q)

Glycine (Gly/G)

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L-Histidine (His/H)

L-Isoleucine (Ile/I)

L-Leucine (Leu/L)

L-Lysine (Lys/K)

L-Methionine (Met/M)

L-Phenylalanine (Phe/F)

L-Proline (Pro/P)

L-Serine (Ser/S)

L-Threonine (Thr/T)

L-Tryptophan (Trp/W)

L-Tyrosine (Tyr/Y)

L-Valine (Val/V)

IUPAC/IUBMB now also recommends standard abbreviations for the following two amino acids:

L-Selenocysteine (Sec/U)

L-Pyrrolysine (Pyl/O)

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Non-specific abbreviations
Sometimes the specific identity of an amino acid cannot be determined unambiguously. Certain protein sequencing techniques do not distinguish among certain pairs. Thus, the following codes are used: Asx (B) is "asparagine or aspartic acid" Glx (Z) is "glutamic acid or glutamine" Xle (J) is "leucine or isoleucine" In addition, the symbol X is used to indicate an amino acid that is completely unidentified.

Chemical properties
Following is a table listing the one-letter symbols, the three-letter symbols, and the chemical properties of the side-chains of the standard amino acids. The masses listed are based on weighted averages of the elemental isotopes at their natural abundances. Note that forming a peptide bond results in elimination of a molecule of water, so the mass of an amino acid unit within a protein chain is reduced by 18.01524 Da. General chemical properties
Amino Acid Short Abbrev. Avg. Mass (Da) pI pK1 pK2 (-COOH) (-+NH ) 3 2.35 1.92 1.99 2.10 2.20 2.35 1.80 2.32 2.16 2.33 2.13 2.14 9.87 10.70 9.90 9.47 9.31 9.78 9.33 9.76 9.06 9.74 9.28 8.72

Alanine Cysteine Aspartic acid Glutamic acid Phenylalanine Glycine Histidine Isoleucine Lysine Leucine Methionine Asparagine Pyrrolysine Proline Glutamine Arginine Serine Threonine Selenocysteine Valine Tryptophan Tyrosine

A C D E F G H I K L M N O P Q R S T U V W Y

Ala Cys Asp Glu Phe Gly His Ile Lys Leu Met Asn Pyl Pro Gln Arg Ser Thr Sec Val Trp Tyr

89.09404 121.15404 133.10384 147.13074 165.19184 75.06714 155.15634 131.17464 146.18934 131.17464 149.20784 132.11904

6.01 5.05 2.85 3.15 5.49 6.06 7.60 6.05 9.60 6.01 5.74 5.41

115.13194 146.14594 174.20274 105.09344 119.12034 168.053 117.14784 204.22844 181.19124

6.30 5.65 10.76 5.68 5.60 5.47 6.00 5.89 5.64

1.95 2.17 1.82 2.19 2.09

10.64 9.13 8.99 9.21 9.10

2.39 2.46 2.20

9.74 9.41 9.21

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Side chain properties

Amino Acid Short Abbrev. Side chain Hydro- pKa Polar phobic pH Small Tiny Aromatic or Aliphatic Aromatic Aromatic Aliphatic Aliphatic 67 86 91 109 135 48 118 124 135 124 124 96 van der Waals volume

Alanine Cysteine

A C

Ala Cys Asp Glu Phe Gly His Ile Lys Leu Met Asn Pyl Pro Gln Arg

-CH3 -CH2SH -CH2COOH -CH2CH2COOH -CH2C6H5 -H -CH2-C3H3N2 -CH(CH3)CH2CH3 -(CH2)4NH2 -CH2CH(CH3)2 -CH2CH2SCH3 -CH2CONH2

X X X X X X X -

8.18 3.90 4.07 6.04 -

X X X -

acidic acidic acidic weak basic basic -

X X X X X

X X -

Aspartic acid D Glutamic acid E Phenylalanine F Glycine Histidine Isoleucine Lysine Leucine Methionine Asparagine Pyrrolysine Proline Glutamine Arginine G H I K L M N O P Q R

10.54 X X

-CH2CH2CH2-CH2CH2CONH2

X -

strongly basic weak acidic -

X -

90 114 148

-(CH2)3NH-C(NH)NH2 -CH2OH -CH(OH)CH3 -CH2SeH -CH(CH3)2 -CH2C8H6N -CH2-C6H4OH X X X -

12.48 X

Serine Threonine

S T

Ser Thr Sec Val Trp Tyr

5.73 -

X X -

X X X X -

X -

Aliphatic Aromatic Aromatic

73 93

Selenocysteine U Valine Tryptophan Tyrosine V W Y

105 163 141

10.46 X

Note: The pKa values of amino acids are typically slightly different when the amino acid is inside a protein. Protein pKa calculations are sometimes used to calculate the change in the pKa value of an amino acid in this situation.

Gene expression and biochemistry

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Amino Acid

Short

Abbrev.

Codon(s)

Occurrence in human proteins (%) 7.8 1.9 5.3 6.3 3.9 7.2 2.3 5.3 5.9

Essential in humans

Alanine Cysteine

A C

Ala Cys Asp Glu Phe Gly His Ile Lys Leu Met Asn Pyl Pro Gln Arg Ser Thr Sec Val Trp Tyr Term

GCU, GCC, GCA, GCG UGU, UGC GAU, GAC GAA, GAG UUU, UUC GGU, GGC, GGA, GGG CAU, CAC AUU, AUC, AUA AAA, AAG

Conditionally Conditionally Yes Conditionally Yes Yes Yes Yes Yes -

Aspartic acid D Glutamic acid E Phenylalanine F Glycine Histidine Isoleucine Lysine Leucine Methionine Asparagine Pyrrolysine Proline Glutamine Arginine Serine Threonine G H I K L M N O P Q R S T

UUA, UUG, CUU, CUC, CUA, CUG 9.1 AUG AAU, AAC UAG* CCU, CCC, CCA, CCG CAA, CAG 5.2 4.2 2.3 4.3

Conditionally Yes -

CGU, CGC, CGA, CGG, AGA, AGG 5.1 UCU, UCC, UCA, UCG, AGU, AGC 6.8 ACU, ACC, ACA, ACG UGA** GUU, GUC, GUA, GUG UGG UAU, UAC UAA, UAG, UGA 6.6 1.4 3.2 5.9

Selenocysteine U Valine Tryptophan Tyrosine Stop codon V W Y -

Yes Yes Conditionally -

* UAG is normally the amber stop codon, but encodes pyrrolysine if a PYLIS element is present. ** UGA is normally the opal (or umber) stop codon, but encodes selenocysteine if a SECIS element is present. The stop codon is not an amino acid, but is included for completeness. An essential amino acid cannot be synthesized in humans and must, therefore, be supplied in the diet. Conditionally essential amino acids are not normally required in the diet, but must be supplied exogenously to specific populations that do not synthesize it in adequate amounts.

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Mass spectrometry
In mass spectrometry of peptides and proteins, it is useful to know the masses of the residues. The mass of the peptide or protein is the sum of the residue masses plus the mass of water.[2]
Amino Acid Alanine Cysteine Aspartic acid Glutamic acid Phenylalanine Glycine Histidine Isoleucine Lysine Leucine Methionine Asparagine Pyrrolysine Proline Glutamine Arginine Serine Threonine Selenocysteine Valine Tryptophan Tyrosine Short A C D E F G H I K L M N O P Q R S T U V W Y Abbrev. Ala Cys Asp Glu Phe Gly His Ile Lys Leu Met Asn Pyl Pro Gln Arg Ser Thr Sec Val Trp Tyr Formula C3H5NO C3H5NOS C4H5NO3 C5H7NO3 C9H9NO C2H3NO C6H7N3O C6H11NO C6H12N2O C6H11NO C5H9NOS C4H6N2O2 C12H21N3O3 C5H7NO C5H8N2O2 C6H12N4O C3H5NO2 C4H7NO2 C3H5NOSe C5H9NO C11H10N2O C9H9NO2 Mon. Mass (Da) Avg. Mass (Da) 71.03711 103.00919 115.02694 129.04259 147.06841 57.02146 137.05891 113.08406 128.09496 113.08406 131.04049 114.04293 255.15829 97.05276 128.05858 156.10111 87.03203 101.04768 150.95364 99.06841 186.07931 163.06333 71.0788 103.1388 115.0886 129.1155 147.1766 57.0519 137.1411 113.1594 128.1741 113.1594 131.1986 114.1039 255.3172 97.1167 128.1307 156.1875 87.0782 101.1051 150.0388 99.1326 186.2132 163.1760

Monoisotopic mass

Stoichiometry and metabolic cost in cell

Following table lists the abundance of amino acids in E.coli cell and the metabolic cost (ATP) for synthesis the amino acids. Negative numbers indicate the metabolic processes are energy favorable and do not cost net ATP of the cell.[3] Note that the abundance of amino acids include amino acids in free-form and in polymerization form (proteins).

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Amino acid

Abundance (# of molecules (108) per E. coli cell) 2.9 0.52 1.4 1.5 1.1 3.5 0.54 1.7 2.0 2.6 0.88 1.4 1.3 1.5 1.7 1.2 1.5 0.33 0.79 2.4

ATP cost in synthesis under aerobic condition -1 11 0 -7 -6 -2 1 7 5 -9 21 3 -2 -6 5 -2 6 -7 -8 -2

ATP cost in synthesis under anaerobic condition 1 15 2 -1 2 2 7 11 9 1 23 5 4 0 13 2 8 7 2 2

Alanine Cysteine Aspartic acid Glutamic acid Phenylalanine Glycine Histidine Isoleucine Lysine Leucine Methionine Asparagine Proline Glutamine Arginine Serine Threonine Tryptophan Tyrosine Valine

Remarks
Amino Acid Abbrev. Remarks

Alanine

Ala Very abundant, very versatile. More stiff than glycine, but small enough to pose only small steric limits for the protein conformation. It behaves fairly neutrally, and can be located in both hydrophilic regions on the protein outside and the hydrophobic areas inside. Asx A placeholder when either amino acid may occupy a position.

Asparagine or aspartic acid Cysteine

Cys The sulfur atom bonds readily to heavy metal ions. Under oxidizing conditions, two cysteines can join together in a disulfide bond to form the amino acid cystine. When cystines are part of a protein, insulin for example, the tertiary structure is stabilized, which makes the protein more resistant to denaturation; therefore, disulfide bonds are common in proteins that have to function in harsh environments including digestive enzymes (e.g., pepsin and chymotrypsin) and structural proteins (e.g., keratin). Disulfides are also found in peptides too small to hold a stable shape on their own (eg. insulin). Asp Behaves similarly to glutamic acid. Carries a hydrophilic acidic group with strong negative charge. Usually is located on the outer surface of the protein, making it water-soluble. Binds to positively-charged molecules and ions, often used in enzymes to fix the metal ion. When located inside of the protein, aspartate and glutamate are usually paired with arginine and lysine.

Aspartic acid

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Glutamic acid Phenylalanine

E F

Glu Behaves similar to aspartic acid. Has longer, slightly more flexible side chain. Phe Essential for humans. Phenylalanine, tyrosine, and tryptophan contain large rigid aromatic group on the side-chain. These are the biggest amino acids. Like isoleucine, leucine and valine, these are hydrophobic and tend to orient towards the interior of the folded protein molecule. Phenylalanine can be converted into Tyrosine. Gly Because of the two hydrogen atoms at the carbon, glycine is not optically active. It is the smallest amino acid, rotates easily, adds flexibility to the protein chain. It is able to fit into the tightest spaces, e.g., the triple helix of collagen. As too much flexibility is usually not desired, as a structural component it is less common than alanine. His In even slightly acidic conditions protonation of the nitrogen occurs, changing the properties of histidine and the polypeptide as a whole. It is used by many proteins as a regulatory mechanism, changing the conformation and behavior of the polypeptide in acidic regions such as the late endosome or lysosome, enforcing conformation change in enzymes. However only a few histidines are needed for this, so it is comparatively scarce. Ile Essential for humans. Isoleucine, leucine and valine have large aliphatic hydrophobic side chains. Their molecules are rigid, and their mutual hydrophobic interactions are important for the correct folding of proteins, as these chains tend to be located inside of the protein molecule.

Glycine

Histidine

Isoleucine

Leucine or isoleucine Lysine

Xle A placeholder when either amino acid may occupy a position

Lys Essential for humans. Behaves similarly to arginine. Contains a long flexible side-chain with a positively-charged end. The flexibility of the chain makes lysine and arginine suitable for binding to molecules with many negative charges on their surfaces. E.g., DNA-binding proteins have their active regions rich with arginine and lysine. The strong charge makes these two amino acids prone to be located on the outer hydrophilic surfaces of the proteins; when they are found inside, they are usually paired with a corresponding negatively-charged amino acid, e.g., aspartate or glutamate. Leu Essential for humans. Behaves similar to isoleucine and valine. See isoleucine. Met Essential for humans. Always the first amino acid to be incorporated into a protein; sometimes removed after translation. Like cysteine, contains sulfur, but with a methyl group instead of hydrogen. This methyl group can be activated, and is used in many reactions where a new carbon atom is being added to another molecule. Asn Similar to aspartic acid. Asn contains an amide group where Asp has a carboxyl. Pyl Similar to lysine, with a pyrroline ring attached.

Leucine Methionine

L M

Asparagine Pyrrolysine Proline

N O P

Pro Contains an unusual ring to the N-end amine group, which forces the CO-NH amide sequence into a fixed conformation. Can disrupt protein folding structures like helix or sheet, forcing the desired kink in the protein chain. Common in collagen, where it often undergoes a posttranslational modification to hydroxyproline. Gln Similar to glutamic acid. Gln contains an amide group where Glu has a carboxyl. Used in proteins and as a storage for ammonia. The most abundant Amino Acid in the body. Arg Functionally similar to lysine. Ser Serine and threonine have a short group ended with a hydroxyl group. Its hydrogen is easy to remove, so serine and threonine often act as hydrogen donors in enzymes. Both are very hydrophilic, therefore the outer regions of soluble proteins tend to be rich with them.

Glutamine

Arginine Serine

R S

Threonine Selenocysteine Valine Tryptophan

T U V W

Thr Essential for humans. Behaves similarly to serine. Sec Selenated form of cysteine, which replaces sulfur. Val Essential for humans. Behaves similarly to isoleucine and leucine. See isoleucine. Trp Essential for humans. Behaves similarly to phenylalanine and tyrosine (see phenylalanine). Precursor of serotonin. Naturally fluorescent. Xaa Placeholder when the amino acid is unknown or unimportant. Tyr Behaves similarly to phenylalanine (precursor to Tyrosine) and tryptophan (see phenylalanine). Precursor of melanin, epinephrine, and thyroid hormones. Naturally fluorescent, although fluorescence is usually quenched by energy transfer to tryptophans. Glx A placeholder when either amino acid may occupy a position.

Unknown Tyrosine

X Y

Glutamic acid or glutamine

Properties of the twenty amino acids

117

Catabolism

[4] Amino acids can be classified according to the properties of their main products as either of the following: Glucogenic, with the products having the ability to form glucose by gluconeogenesisKetogenic, with the products not having the ability to form glucose. These products may still be used for ketogenesis or lipid synthesis.Amino acids catabolized into both glucogenic and ketogenic products.

References
[1] Ambrogelly A, Palioura S, Sll D (Jan 2007). "Natural expansion of the genetic code" (http:/ / www. nature. com/ nchembio/ journal/ v3/ n1/ abs/ nchembio847. html). Nat Chem Biol 3 (1): 2935. doi:10.1038/nchembio847. PMID17173027. . [2] "The amino acid masses" (http:/ / education. expasy. org/ student_projects/ isotopident/ htdocs/ aa-list. html). ExPASy. . Retrieved 2009-01-06. [3] Physical Biology of the Cell (Garland Science) p. 178 [4] Chapter 20 (Amino Acid Degradation and Synthesis) in: Denise R., PhD. Ferrier. Lippincott's Illustrated Reviews: Biochemistry (Lippincott's Illustrated Reviews). Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-2265-9.

Nelson, David L.; Cox, Michael M. (2000). Lehninger Principles of Biochemistry (3rd ed.). Worth Publishers. ISBN1-57259-153-6. Kyte, J.; Doolittle, R. F. (1982). "A simple method for displaying the hydropathic character of a protein". J. Mol. Biol. 157 (1): 105132. doi:10.1016/0022-2836(82)90515-0. PMID7108955. Meierhenrich, Uwe J. (2008). Amino acids and the asymmetry of life (1st ed.). Springer. ISBN978-3-540-76885-2.

Myoglobin

118

Myoglobin
Myoglobin

Model of helical domains in myoglobin. Available structures

[1]

PDB 1m6c [2], 1m6m [3], 1mdn [4], 1mnh [5], 1mni [6], 1mnj [7], 1mnk [8], 1mno [9], 1mwc [10], 1mwd [11], 1myg [12], 1myh [13] [14] [15] [16] [17] [18] [19] , 1myi , 1myj , 1pmb , 1yca , 1ycb , 2mm1 Identifiers Symbols External IDs MB
[20]

; MGC13548; PVALB
[21]

OMIM: 160000 Gene Ontology

MGI: 96922

[22]

HomoloGene: 3916

[23]

GeneCards: MB Gene

[24]

Molecular function oxygen transporter activity [25] [26] iron ion binding [27] oxygen binding [28] heme binding [29] metal ion binding Biological process response to hypoxia [31] transport [32] oxygen transport [33] enucleate erythrocyte differentiation
[34] [30]

Sources: Amigo

/ QuickGO

[35]

RNA expression pattern

More reference expression data Orthologs Species Human Mouse

[36]

Myoglobin
[37] [39] [38] [40]

119

Entrez Ensembl UniProt RefSeq (mRNA) RefSeq (protein) Location (UCSC)

4151

17189

ENSG00000198125 P02144
[41] [43]

ENSMUSG00000018893 Q3UVB1
[42] [44]

NM_005368 NP_005359

NM_013593 NP_038621

[45]

[46]

Chr 22: [47] 34.33 34.35 Mb

[49]

Chr 15: [48] 76.84 76.88 Mb

[50]

PubMed search

Myoglobin is an iron- and oxygen-binding protein found in the muscle tissue of vertebrates in general and in almost all mammals. It is related to hemoglobin, which is the iron- and oxygen-binding protein in blood, specifically in the red blood cells. The only time myoglobin is found in the bloodstream is when it is released following muscle injury. It is an abnormal finding, and can be diagnostically relevant when found in blood. [51] Myoglobin (abbreviated Mb) is a single-chain globular protein of 153[52] or 154[53] amino acids, containing a heme (iron-containing porphyrin) prosthetic group in the center around which the remaining apoprotein folds. It has eight alpha helices and a hydrophobic core. It has a molecular weight of 17,699 daltons (with heme), and is the primary oxygen-carrying pigment of muscle tissues.[53] Unlike the blood-borne hemoglobin, to which it is structurally related,[54] this protein does not exhibit cooperative binding of oxygen, since positive cooperativity is a property of multimeric/oligomeric proteins only. Instead, the binding of oxygen by myoglobin is unaffected by the oxygen pressure in the surrounding tissue. Myoglobin is often cited as having an "instant binding tenacity" to oxygen given its hyperbolic oxygen dissociation curve. High concentrations of myoglobin in muscle cells allow organisms to hold their breaths longer. Diving mammals such as whales and seals have muscles with particularly high myoglobin abundance.[51] Myoglobin was the first protein to have its three-dimensional structure revealed.[55] In 1958, John Kendrew and associates successfully determined the structure of myoglobin by high-resolution X-ray crystallography.[56] For this discovery, John Kendrew shared the 1962 Nobel Prize in chemistry with Max Perutz.[57] Despite being one of the most studied proteins in biology, its true physiological function is not yet conclusively established: mice genetically engineered to lack myoglobin are viable, but showed a 30% reduction in cardiac systolic output. They adapted to this deficiency through hypoxic genetic mechanisms and increased vasodilation.[58] In humans myoglobin is encoded by the MB gene.[59]

Myoglobin

120

Meat color
Myoglobin forms pigments responsible for making meat red. The color that meat takes is partly determined by the oxidation states of the iron atom in myoglobin and the oxygen species attached to it. When meat is in its raw state, the iron atom is in the +2 oxidation state, and is bound to a dioxygen molecule (O2). Meat cooked well done is brown because the iron atom is now in the +3 oxidation state, having lost an electron, and is now coordinated by a water molecule. Under some conditions, meat can also remain pink all through cooking, despite being heated to high temperatures. If meat has been exposed to nitrites, it will remain pink because the iron atom is bound to NO, nitric oxide (true of, e.g., corned beef or cured hams). Grilled meats can also take on a pink An X-ray diffraction image for the protein myoglobin "smoke ring" that comes from the iron binding to a molecule of carbon monoxide to give metmyoglobin.[60] Raw meat packed in a carbon monoxide atmosphere also shows this same pink "smoke ring" due to the same coordination chemistry. Notably, the surface of this raw meat also displays the pink color, which is usually associated in consumers' minds with fresh meat. This artificially-induced pink color can persist in the meat for a very long time, reportedly up to one year.[61] Hormel and Cargill are both reported to use this meat-packing process, and meat treated this way has been in the consumer market since 2003.[62] Myoglobin is found in Type I muscle, Type II A and Type II B, but most texts consider myoglobin not to be found in smooth muscle.

Role in disease
Myoglobin is released from damaged muscle tissue (rhabdomyolysis), which has very high concentrations of myoglobin. The released myoglobin is filtered by the kidneys but is toxic to the renal tubular epithelium and so may cause acute renal failure.[63] It is not the myoglobin itself that is toxic (it is a protoxin) but the ferrihemate portion that is dissociated from myoglobin in acidic environments (e.g., acidic urine, lysozymes). Myoglobin is a sensitive marker for muscle injury, making it a potential marker for heart attack in patients with chest pain.[64] However, elevated myoglobin has low specificity for acute myocardial infarction (AMI) and thus CK-MB, cTnT, ECG, and clinical signs should be taken into account to make the diagnosis.

Structure and bonding

Myoglobin contains a porphyrin ring with an iron center. There is a proximal histidine group attached directly to the iron center, and a distal histidine group on the opposite face, not bonded to the iron. Many functional models of myoglobin have been studied. One of the most important is that of picket fence porphyrin by James Collman. This model was used to show the importance of the distal prosthetic group. It serves three functions: 1. To form hydrogen bonds with the dioxygen moiety, increasing the O2 binding constant 2. To prevent the binding of carbon monoxide, whether from within or without the body. Carbon monoxide binds to iron in an end-on fashion, and is hindered by the presence of the distal histidine, which forces it into a bent conformation. CO binds to heme 23,000 times better than O2, but only 200 times better in hemoglobin and myoglobin. Oxygen binds in a bent fashion, which can fit with the distal histidine.[65] 3. To prevent irreversible dimerization of the oxymyoglobin with another deoxymyoglobin species

Myoglobin

121

References
[1] PDB 1MBO (http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1MBO); Takano T (March 1977). "Structure of myoglobin refined at 2.0 resolution. II. Structure of deoxymyoglobin from sperm whale". J. Mol. Biol. 110 (3): 56984. doi:10.1016/S0022-2836(77)80112-5. PMID845960. [2] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1m6c [3] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1m6m [4] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mdn [5] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mnh [6] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mni [7] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mnj [8] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mnk [9] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mno [10] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mwc [11] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1mwd [12] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1myg [13] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1myh [14] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1myi [15] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1myj [16] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1pmb [17] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1yca [18] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=1ycb [19] [20] [21] [22] [23] [24] [25] [26] [27] [28] [29] [30] [31] [32] [33] [34] [35] [36] [37] [38] [39] [40] [41] [42] [43] [44] [45] [46] [47] [48] [49] [50] [51] http:/ / www. rcsb. org/ pdb/ cgi/ explore. cgi?pdbId=2mm1 http:/ / www. genenames. org/ data/ hgnc_data. php?hgnc_id=6915 http:/ / omim. org/ entry/ 160000 http:/ / www. informatics. jax. org/ searches/ accession_report. cgi?id=MGI:96922 http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Retrieve& db=homologene& dopt=HomoloGene& list_uids=3916 http:/ / www. genecards. org/ cgi-bin/ carddisp. pl?id_type=entrezgene& id=4151 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0005344 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0005506 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0019825 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0020037 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0046872 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0001666 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0006810 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0015671 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?view=details& search_constraint=terms& depth=0& query=GO:0043353 http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ gp-assoc. cgi?gp=UniProtKB:P02144 http:/ / www. ebi. ac. uk/ QuickGO/ GProtein?ac=P02144 http:/ / biogps. org/ gene/ 4151/ http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=gene& cmd=retrieve& dopt=default& list_uids=4151& rn=1 http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=gene& cmd=retrieve& dopt=default& list_uids=17189& rn=1 http:/ / www. ensembl. org/ Homo_sapiens/ geneview?gene=ENSG00000198125;db=core http:/ / www. ensembl. org/ Mus_musculus/ geneview?gene=ENSMUSG00000018893;db=core http:/ / www. uniprot. org/ uniprot/ P02144 http:/ / www. uniprot. org/ uniprot/ Q3UVB1 http:/ / www. ncbi. nlm. nih. gov/ entrez/ viewer. fcgi?val=NM_005368 http:/ / www. ncbi. nlm. nih. gov/ entrez/ viewer. fcgi?val=NM_013593 http:/ / www. ncbi. nlm. nih. gov/ entrez/ viewer. fcgi?val=NP_005359 http:/ / www. ncbi. nlm. nih. gov/ entrez/ viewer. fcgi?val=NP_038621 http:/ / genome. ucsc. edu/ cgi-bin/ hgTracks?org=Human& db=hg18& position=chr22:34332757-34349347 http:/ / genome. ucsc. edu/ cgi-bin/ hgTracks?org=Mouse& db=mm8& position=chr15:76842742-76877925 http:/ / www. ncbi. nlm. nih. gov/ sites/ entrez?db=gene& cmd=Link& LinkName=gene_pubmed& from_uid=4151 http:/ / www. ncbi. nlm. nih. gov/ sites/ entrez?db=gene& cmd=Link& LinkName=gene_pubmed& from_uid=17189 Nelson, D. L.; Cox, M. M. (2000). Lehninger Principles of Biochemistry (3rd ed.). New York: Worth Publishers. p.206. ISBN071676203X.

[52] Hendgen-Cotta, U.; Kelm, M.; Rassaf, T. (2009). "A highlight of myoglobin diversity: the nitrite reductase activity during myocardial ischemia-reperfusion". Nitric oxide : biology and chemistry / official journal of the Nitric Oxide Society 22 (2): 7582. doi:10.1016/j.niox.2009.10.003. PMID19836457.

Myoglobin
[53] George A. Ordway and Daniel J. Garry (2004). "Myoglobin: an essential hemoprotein in striated muscle". Journal of Experimental Biology 207 (Pt 20): 34416. doi:10.1242/jeb.01172. PMID15339940. [54] Harvey Lodish, Arnold Berk, Lawrence S. Zipursky, Paul Matsudaira, David Baltimore and James Darnell (2000). "Evolutionary tree showing the globin protein family members myoglobin and hemoglobin" (http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Search& db=books& doptcmdl=GenBookHL& term=myoglobin+ AND+ mcb[book]+ AND+ 105134[uid]& rid=mcb. figgrp. 540). Molecular Cell Biology (4th ed.). W. H. Freeman. ISBN0-7167-3136-3. . [55] (U.S.) National Science Foundation: Protein Data Bank Chronology (Jan. 21, 2004) (http:/ / www. nsf. gov/ news/ news_summ. jsp?cntn_id=100689). Retrieved 3.17.2010 [56] JC Kendrew, G Bodo, HM Dintzis, RG Parrish, H Wyckoff, and DC Phillips (1958). "A Three-Dimensional Model of the Myoglobin Molecule Obtained by X-Ray Analysis". Nature 181 (4610): 6626. Bibcode1958Natur.181..662K. doi:10.1038/181662a0. PMID13517261. [57] The Nobel Prize in Chemistry 1962 (http:/ / nobelprize. org/ chemistry/ laureates/ 1962/ index. html) [58] Mammen PP, Kanatous SB, Yuhanna IS, Shaul PW, Garry MG, Balaban RS, Garry DJ (2003). "Hypoxia-induced left ventricular dysfunction in myoglobin-deficient mice". American Journal of Physiology. Heart and Circulatory Physiology 285 (5): H213241. doi:10.1152/ajpheart.00147.2003. PMID12881221. [59] Akaboshi E (1985). "Cloning of the human myoglobin gene". Gene 33 (3): 2419. doi:10.1016/0378-1119(85)90231-8. PMID2989088. [60] McGee, H (2004). On Food and Cooking: The Science and Lore of the Kitchen. New York: Scribner. p.148. ISBN0-684-80001-2. [61] Minneapolis Star Tribune, Nov. 14, 2007 http:/ / www. startribune. com/ 10223/ story/ 1548852. html [62] Minneapolis Star Tribune, October 31, 2007 http:/ / www. startribune. com/ 535/ story/ 1518775. html [63] Toshio Naka, Daryl Jones, Ian Baldwin, Nigel Fealy, Samantha Bates, Hermann Goehl, Stanislao Morgera, Hans H. Neumayer and Rinaldo Bellomo (2005). "Myoglobin clearance by super high-flux hemofiltration in a case of severe rhabdomyolysis: a case report". Critical Care 9 (2): R905. doi:10.1186/cc3034. PMC1175920. PMID15774055. [64] M. Weber, M. Rau, K. Madlener, A. Elsaesser, D. Bankovic, V. Mitrovic and C. Hamm (2005). "Diagnostic utility of new immunoassays for the cardiac markers cTnI, myoglobin and CK-MB mass". Clinical Biochemistry 38 (11): 102730. doi:10.1016/j.clinbiochem.2005.07.011. PMID16125162. [65] J. P. Collman, J. I. Brauman, T. R. Halbert, and K. S. Suslick (1976). "Nature of Oxygen and Carbon Monoxide Binding to Metalloporphyrins and Heme Proteins" (http:/ / www. pnas. org/ cgi/ content/ abstract/ 73/ 10/ 3333). Proceedings of the National Academy of Sciences of the United States of America 73 (10): 33337. doi:10.1073/pnas.73.10.3333. PMC431107. PMID1068445. .

122

Further reading
J. P. Collman, R. Boulatov, C. J. Sunderland and L. Fu (2004). "Functional Analogues of Cytochrome c Oxidase, Myoglobin, and Hemoglobin". Chem. Rev. 104 (2): 561588. doi:10.1021/cr0206059. PMID14871135. Reeder, BJ; Svistunenko DA, Cooper CE, Wilson MT (December 2004). "The radical and redox chemistry of myoglobin and hemoglobin: from in vitro studies to human pathology". Antioxid Redox Signal 6 (6): 95466. doi:10.1089/ars.2004.6.954. PMID15548893. Schlieper, G; Kim JH, Molojavyi A, Jacoby C, Laussmann T, Flogel U, Godecke A, Schrader J (April 2004). "Adaptation of the myoglobin knockout mouse to hypoxic stress". Am J Physiol Regul Integr Comp Physiol 286 (4): R78692. doi:10.1152/ajpregu.00043.2003. PMID14656764. Takano, T (1977). "Structure of myoglobin refined at 2-0 A resolution. II. Structure of deoxymyoglobin from sperm whale". J. Mol. Biol. 110 (3): 569584. doi:10.1016/S0022-2836(77)80112-5. PMID845960. Roy, A; Sen S, Chakraborti AS (February 2004). "In vitro nonenzymatic glycation enhances the role of myoglobin as a source of oxidative stress". Free Radic Res. 38 (2): 13946. doi:10.1080/10715160310001638038. PMID15104207. Stewart, JM; Blakely JA, Karpowicz PA, Kalanxhi E, Thatcher BJ, Martin BM (March 2004). "Unusually weak oxygen binding, physical properties, partial sequence, autoxidation rate and a potential phosphorylation site of beluga whale (Delphinapterus leucas) myoglobin". Comp Biochem Physiol B Biochem Mol Biol 137 (3): 40112. doi:10.1016/j.cbpc.2004.01.007. PMID15050527. Wu, G; Wainwright LM, Poole RK (2003). "Microbial globins". Adv Microb Physiol 47: 255310. doi:10.1016/S0065-2911(03)47005-7. PMID14560666.

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123

External links
The Myoglobin Protein (http://macromoleculeinsights.com/myoglobin.php) Protein Database featured molecule (http://pdbdev.sdsc.edu:48346/pdb/molecules/mb1.html) Online 'Mendelian Inheritance in Man' (OMIM) 160000 (http://omim.org/entry/160000) human genetics Which Cut Is Older? (It's a Trick Question) (http://www.nytimes.com/2006/02/21/national/21meat.html) New York Times, February 21, 2006 article regarding meat industry use of carbon monoxide to keep meat looking red. Stores React to Meat Reports (http://www.nytimes.com/2006/03/01/dining/01meat.html) New York Times, March 1, 2006 article on the use of carbon monoxide to make meat appear fresh.

Hemoglobin

124

Hemoglobin
Hemoglobin, human, adult
(heterotetramer, ()2)

Structure of human hemoglobin. The protein's and subunits are in red and blue, and the iron-containing heme groups in green. From PDB 1GZX

[1]Proteopedia Hemoglobin [2]

Protein type Function Cofactor(s)

-

metalloprotein, globulin oxygen-transport heme (4)

Subunit Name Hb-1 Hb-2 Hb-

Gene

Chromosomal Locus Chr. 16 p13.3 Chr. 16 p13.3 Chr. 11 p15.5 [3] [3] [4]

HBA1 HBA2 HBB

Hemoglobin (English pronunciation: /himlobn/; also rendered as haemoglobin and abbreviated Hb or Hgb) is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates,[5] with the exception of the fish family Channichthyidae,[6] as well as the tissues of some invertebrates. Hemoglobin in the blood carries oxygen from the respiratory organs (lungs or gills) to the rest of the body (i.e.,the tissues) where it releases the oxygen to burn nutrients to provide energy to power the functions of the organism, and collects the resultant carbon dioxide to bring it back to the respiratory organs to be dispensed from the organism. In mammals, the protein makes up about 97% of the red blood cells' dry content, and around 35% of the total content (including water). Hemoglobin has an oxygen binding capacity of 1.34 ml O2 per gram of hemoglobin,[7] which increases the total blood oxygen capacity seventy-fold compared to dissolved oxygen in blood. The mammalian hemoglobin molecule can bind (carry) up to four oxygen molecules.[8] Hemoglobin is involved in the transport of other gases: it carries some of the body's respiratory carbon dioxide (about 10% of the total) as carbaminohemoglobin, in which CO2 is bound to the globin protein. The molecule also carries the important regulatory molecule nitric oxide bound to a globin protein thiol group, releasing it at the same time as oxygen.[9] Hemoglobin is also found outside red blood cells and their progenitor lines. Other cells that contain hemoglobin include the A9 dopaminergic neurons in the substantia nigra, macrophages, alveolar cells, and mesangial cells in the kidney. In these tissues, hemoglobin has a non-oxygen-carrying function as an antioxidant and a regulator of iron

Hemoglobin metabolism.[10] Hemoglobin and hemoglobin-like molecules are also found in many invertebrates, fungi, and plants. In these organisms, hemoglobins may carry oxygen, or they may act to transport and regulate other things such as carbon dioxide, nitric oxide, hydrogen sulfide and sulfide. A variant of the molecule, called leghemoglobin, is used to scavenge oxygen, to keep it from poisoning anaerobic systems, such as nitrogen-fixing nodules of leguminous plants.

125

Research history
The oxygen-carrying protein hemoglobin was discovered by Hnefeld in 1840.[11] In 1851,[12] Otto Funke published a series of articles in which he described growing hemoglobin crystals by successively diluting red blood cells with a solvent such as pure water, alcohol or ether, followed by slow evaporation of the solvent from the resulting protein solution.[13] Hemoglobin's reversible oxygenation was described a few years later by Felix Hoppe-Seyler.[14] In 1959 Max Perutz determined the molecular structure of hemoglobin by X-ray crystallography.[15] resulted in his sharing with John Kendrew the 1962 Nobel Prize in Chemistry.
[16]

This work

The role of hemoglobin in the blood was elucidated by physiologist Claude Bernard. The name hemoglobin is derived from the words heme and globin, reflecting the fact that each subunit of hemoglobin is a globular protein with an embedded heme group. Each heme group contains one iron atom, that can bind one oxygen molecule through ion-induced dipole forces. The most common type of hemoglobin in mammals contains four such subunits.

Genetics
Hemoglobin consists mostly of protein (the "globin" chains) subunits, and these proteins, in turn, are folded chains of a large number of different amino acids called polypeptides. The amino acid sequence of any polypeptide created by a cell, is in turn determined by the stretches of DNA called genes. In all proteins, it is the amino acid sequence, which determines the protein's chemical properties and function. There is more than one hemoglobin gene. The amino acid sequences of the globin proteins in hemoglobins usually differ between species. These differences grow with evolutionary distance between species. For example, the most common hemoglobin sequences in humans and chimpanzees are nearly identical, differing by only one amino acid in both the alpha and the beta globin protein chains. These differences grow larger between less closely related species. Even within a species, different variants of hemoglobin always exist, although one sequence is usually a "most common" one in each species. Mutations in the genes for the hemoglobin protein in a species result in hemoglobin variants.[17] [18] Many of these mutant forms of hemoglobin cause no disease. Some of these mutant forms of hemoglobin, however, cause a group of hereditary diseases termed the hemoglobinopathies. The best known hemoglobinopathy is sickle-cell disease, which was the first human disease whose mechanism was understood at the molecular level. A (mostly) separate set of diseases called thalassemias involves underproduction of normal and sometimes abnormal hemoglobins, through problems and mutations in globin gene regulation. All these diseases produce anemia.[19] Variations in hemoglobin amino acid sequences, as with other proteins, may be adaptive. For example, recent studies have suggested genetic variants in deer mice that help explain how deer mice that live in the mountains are able to survive in the thin air that accompanies high altitudes. A researcher from the University of Nebraska-Lincoln found mutations in four different genes that can account for differences between deer mice that live in lowland prairies versus the mountains. After examining wild mice captured from both highlands and lowlands, it was found that: the genes of the two breeds are virtually identicalexcept for those that govern the oxygen-carrying capacity of their hemoglobin. The genetic difference enables highland mice to make more efficient use of their oxygen, since less is available at higher altitudes, such as those in the mountains.[20] Mammoth hemoglobin featured mutations that allowed for oxygen delivery at lower temperatures, thus enabling mammoths to migrate to higher latitudes during the

Hemoglobin Pleistocene.[21]

126

Synthesis
Hemoglobin (Hb) is synthesized in a complex series of steps. The heme part is synthesized in a series of steps in the mitochondria and the cytosol of immature red blood cells, while the globin protein parts are synthesized by ribosomes in the cytosol.[22] Production of Hb continues in the cell throughout its early development from the proerythroblast to the reticulocyte in the bone marrow. At this point, the nucleus is lost in mammalian red blood cells, but not in birds and many other species. Even after the loss of the nucleus in mammals, residual ribosomal RNA allows further synthesis of Hb until the reticulocyte loses its RNA soon after entering the vasculature (this hemoglobin-synthetic RNA in fact gives the reticulocyte its reticulated appearance and name).

Structure
Hemoglobin has a quaternary structure characteristic of many multi-subunit globular proteins.[23] Most of the amino acids in hemoglobin form alpha helices, connected by short non-helical segments. Hydrogen bonds stabilize the helical sections inside this protein, causing attractions within the molecule, folding each polypeptide chain into a specific shape.[24] Hemoglobin's quaternary structure comes from its four subunits in roughly a tetrahedral arrangement.[23] In most vertebrates, the hemoglobin molecule is an assembly of four globular protein subunits. Each subunit is composed of a protein chain tightly associated with a non-protein heme group. Each protein chain arranges into a set of alpha-helix structural segments connected together in a globin fold arrangement, so called because this Heme b group arrangement is the same folding motif used in other heme/globin proteins such as myoglobin.[25] [26] This folding pattern contains a pocket that strongly binds the heme group. A heme group consists of an iron (Fe) ion (charged atom) held in a heterocyclic ring, known as a porphyrin. This porphyrin ring consists of four pyrrole molecules cyclically linked together (by methene bridges) with the iron ion bound in the center.[27] The iron ion, which is the site of oxygen binding, coordinates with the four nitrogens in the center of the ring, which all lie in one plane. The iron is bound strongly (covalently) to the globular protein via the imidazole ring of the F8 histidine residue (also known as the proximal histidine) below the porphyrin ring. A sixth position can reversibly bind oxygen by a coordinate covalent bond,[28] completing the octahedral group of six ligands. Oxygen binds in an "end-on bent" geometry where one oxygen atom binds Fe and the other protrudes at an angle. When oxygen is not bound, a very weakly bonded water molecule fills the site, forming a distorted octahedron. Even though carbon dioxide is carried by hemoglobin, it does not compete with oxygen for the iron-binding positions, but is actually bound to the protein chains of the structure. The iron ion may be either in the Fe2+ or in the Fe3+ state, but ferrihemoglobin (methemoglobin) (Fe3+) cannot bind oxygen.[29] In binding, oxygen temporarily and reversibly oxidizes (Fe2+) to (Fe3+) while oxygen temporally turns into superoxide, thus iron must exist in the +2 oxidation state to bind oxygen. If superoxide ion associated to Fe3+ is protonated the hemoglobin iron will remain oxidized and incapable to bind oxygen. In such cases, the enzyme methemoglobin reductase will be able to eventually reactivate methemoglobin by reducing the iron center. In adult humans, the most common hemoglobin type is a tetramer (which contains 4 subunit proteins) called hemoglobin A, consisting of two and two subunits non-covalently bound, each made of 141 and 146 amino acid

Hemoglobin residues, respectively. This is denoted as 22. The subunits are structurally similar and about the same size. Each subunit has a molecular weight of about 17,000daltons, for a total molecular weight of the tetramer of about 64,000daltons (64,458 g/mol).[30] Thus, 1 g/dL = 0.1551mmol/L. Hemoglobin A is the most intensively studied of the hemoglobin molecules. In human infants, the hemoglobin molecule is made up of 2 chains and 2 gamma chains. The gamma chains are gradually replaced by chains as the infant grows.[31] The four polypeptide chains are bound to each other by salt bridges, hydrogen bonds, and the hydrophobic effect.

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Oxygen saturation
In general, hemoglobin can be saturated with oxygen molecules (oxyhemoglobin), or desaturated with oxygen molecules (deoxyhemoglobin).[32] Oxyhemoglobin Oxyhemoglobin is formed during physiological respiration when oxygen binds to the heme component of the protein hemoglobin in red blood cells. This process occurs in the pulmonary capillaries adjacent to the alveoli of the lungs. The oxygen then travels through the blood stream to be dropped off at cells where it is utilized in glycolysis and in the production of ATP by the process of oxidative phosphorylation. It does not, however, help to counteract a decrease in blood pH. Ventilation, or breathing, may reverse this condition by removal of carbon dioxide, thus causing a shift up in pH.[33] Hemoglobin exists in two forms, a taut form (T) and a relaxed form (R). Various factors such as low pH, high CO2 and high 2,3 BPG at the level of the tissues favor the taut form, which has low oxygen affinity and releases oxygen in the tissues. Conversely, a high pH, low CO2, or low 2,3 BPG favors the relaxed form which can better bind oxygen. Deoxygenated hemoglobin Deoxygenated hemoglobin is the form of hemoglobin without the bound oxygen. The absorption spectra of oxyhemoglobin and deoxyhemoglobin differ. The oxyhemoglobin has significantly lower absorption of the 660nm wavelength than deoxyhemoglobin, while at 940nm its absorption is slightly higher. This difference is used for measurement of the amount of oxygen in patient's blood by an instrument called pulse oximeter. This difference also accounts for the presentation of cyanosis, the blue to purplish color that tissues develop during hypoxia.

Iron's oxidation state in oxyhemoglobin

Assigning oxygenated hemoglobin's oxidation state is difficult because oxyhemoglobin (Hb-O2), by experimental measurement, is diamagnetic (no net unpaired electrons), yet the low-energy electron configurations in both oxygen and iron are paramagnetic (suggesting at least one unpaired electron in the complex). The lowest-energy form of oxygen, and the lowest energy forms of the relevant oxidation states of iron, are these: Triplet oxygen, the lowest energy molecular oxygen species, has two unpaired electrons in antibonding * molecular orbitals. Iron(II) tends to exist in a high-spin configuration where unpaired electrons exist in Eg antibonding orbitals. Iron(III) has an odd number of electrons, and thus must have one or more unpaired electrons, in any energy state. All of these structures are paramagnetic (have unpaired electrons), not diamagnetic. Thus, a non-intuitive (e.g., a higher-energy for at least one species) distribution of electrons in the combination of iron and oxygen must exist, in order to explain the observed diamagnetism and no unpaired electrons. The three logical possibilities to produce diamagnetic (no net spin) Hb-O2 are:

Hemoglobin 1. Low-spin Fe2+ binds to singlet oxygen. Both low-spin iron and singlet oxygen are diamagnetic. However, the singlet form of oxygen is the higher-energy form of the molecule. 2. Low-spin Fe3+ binds to .O2- (the superoxide ion) and the two unpaired electrons couple antiferromagnetically, giving diamagnetic properties. 3. Low-spin Fe4+ binds to peroxide, O22-. Both are diamagnetic. Direct experimental data: X-ray photoelectron spectroscopy suggests iron has an oxidation state of approximately 3.2 infrared stretching frequencies of the O-O bond suggests a bond length fitting with superoxide (a bond order of about 1.6, with superoxide being 1.5). X-ray Absorption Near Edge Structures at the iron K-edge. The energy shift of 5 eV between Deoxyhemoglobin and Oxyhemoglobin, as for all the Methemoglobin species, strongly suggests an actual local charge closer to Fe3+ than Fe2+.[34] [35] [36] Thus, the nearest formal oxidation state of iron in Hb-O2 is the +3 state, with oxygen in the -1 state (as superoxide .O2-). The diamagnetism in this configuration arises from the single unpaired electron on superoxide aligning antiferromagnetically from the single unpaired electron on iron, to give no net spin to the entire configuration, in accordance with diamagnetic oxyhemoglobin from experiment.[37] [38] The second choice of the three logical possibilities above for diamagnetic oxyhemoglobin being found correct by experiment, is not surprising: singlet oxygen (possibility #1) and large separations of charge (possibility #3) are both unfavorably high-energy states. Iron's shift to a higher oxidation state in Hb-O2 decreases the atom's size, and allows it into the plane of the porphyrin ring, pulling on the coordinated histidine residue and initiating the allosteric changes seen in the globulins. Early postulates by bio-inorganic chemists claimed that possibility #1 (above) was correct and that iron should exist in oxidation state II. This seemed particularly likely since the iron oxidation state III as methemoglobin, when not accompanied by superoxide .O2- to "hold" the oxidation electron, was known to render hemoglobin incapable of binding normal triplet O2 as it occurs in the air. It was thus assumed that iron remained as Fe(II) when oxygen gas was bound in the lungs. The iron chemistry in this previous classical model was elegant, but the required presence of the required diamagnetic high-energy singlet oxygen was never explained. It was classically argued that the binding of an oxygen molecule placed high-spin iron(II) in an octahedral field of strong-field ligands; this change in field would increase the crystal field splitting energy, causing iron's electrons to pair into the low-spin configuration, which would be diamagnetic in Fe(II). This forced low-spin pairing is indeed thought to happen in iron when oxygen binds, but is not enough to explain iron's change in size. Extraction of an additional electron from iron by oxygen is required to explain both iron's smaller size and observed increased oxidation state, and oxygen's weaker bond. It should be noted that the assignment of a whole-number oxidation state is a formalism, as the covalent bonds are not required to have perfect bond orders involving whole electron-transfer. Thus, all three models for paramagnetic Hb-O2 may contribute to some small degree (by resonance) to the actual electronic configuration of Hb-O2. However, the model of iron in Hb-O2 being Fe(III) is more correct than the classical idea that it remains Fe(II).

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Binding for ligands other than oxygen

Besides the oxygen ligand, which binds to hemoglobin in a cooperative manner, hemoglobin ligands also include competitive inhibitors such as carbon monoxide (CO) and allosteric ligands such as carbon dioxide (CO2) and nitric oxide (NO). The carbon dioxide is bound to amino groups of the globin proteins as carbaminohemoglobin, and is thought to account for about 10% of carbon dioxide transport in mammals. Nitric oxide is bound to specific thiol groups in the globin protein to form an S-nitrosothiol which dissociates into free nitric oxide and thiol again, as the hemoglobin releases oxygen from its heme site. This nitric oxide transport to peripheral tissues is hypothesized to assist oxygen transport in tissues, by releasing vasodilatory nitric oxide to tissues in which oxygen levels are low.[39]

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Cooperative
When oxygen binds to the iron complex, it causes the iron atom to move back toward the center of the plane of the porphyrin ring (see moving diagram). At the same time, the imidazole side-chain of the histidine residue interacting at the other pole of the iron is pulled toward the porphyrin ring. This interaction forces the plane of the ring sideways toward the outside of the tetramer, and also induces a strain in the protein helix containing the histidine as it moves nearer to the iron atom. This strain is transmitted to the remaining three monomers in the tetramer, where it induces a similar conformational change in the other heme sites such that binding of oxygen to these sites becomes easier. In the tetrameric form of normal adult hemoglobin, the binding of oxygen is, thus, a cooperative process. The binding affinity of hemoglobin for oxygen is increased by the oxygen saturation of the molecule, with the first oxygens bound influencing the shape of the binding sites for the next oxygens, in a way favorable for binding. This positive cooperative binding is achieved through steric conformational changes of the hemoglobin protein complex as discussed above; i.e., when one subunit protein in hemoglobin becomes oxygenated, a conformational or structural change in the whole complex is initiated, causing the other subunits to gain an increased affinity for oxygen. As a consequence, the oxygen binding curve of hemoglobin is sigmoidal, or S-shaped, as opposed to the normal hyperbolic curve associated with noncooperative binding.

A schematic visual model of oxygen-binding process, showing all four monomers and hemes, and protein chains only as diagramatic coils, to facilitate visualization into the molecule. Oxygen is not shown in this model, but, for each of the iron atoms, it binds to the iron (red sphere) in the flat heme. For example, in the upper left of the four hemes shown, oxygen binds at the left of the iron atom shown in the upper left of diagram. This causes the iron atom to move backward into the heme which holds it (the iron moves upward as it binds oxygen, in this illustration), tugging the histidine residue (modeled as a red pentagon on the right of the iron) closer, as it does. This, in turn, pulls on the protein chain holding the histidine.

The dynamic mechanism of the cooperativity in hemoglobin and its relation with the low-frequency resonance has been discussed.[40]

Competitive
Hemoglobin's oxygen-binding capacity is decreased in the presence of carbon monoxide because both gases compete for the same binding sites on hemoglobin, carbon monoxide binding preferentially in place of oxygen. The binding of oxygen is affected by molecules such as carbon monoxide (CO) (for example, from tobacco smoking, car exhaust, and incomplete combustion in furnaces). CO competes with oxygen at the heme binding site. Hemoglobin binding affinity for CO is 250 times greater than its affinity for oxygen,[41] meaning that small amounts of CO dramatically reduce hemoglobin's ability to transport oxygen. Since Carbon Monoxide is a colorless, odorless and tasteless gas, and poses a potentially fatal threat detectors have become commercially available to warn of dangerous levels in residences. When hemoglobin combines with CO, it forms a very bright red compound called carboxyhemoglobin, which may cause the skin of CO poisoning victims to appear pink in death, instead of white or blue. When inspired air contains CO levels as low as 0.02%, headache and nausea occur; if the CO concentration is increased to 0.1%, unconsciousness will follow. In heavy smokers, up to 20% of the oxygen-active sites can be blocked by CO. In similar fashion, hemoglobin also has competitive binding affinity for cyanide (CN-), sulfur monoxide (SO), nitric oxide (NO), and sulfide(S2-), including hydrogen sulfide (H2S). All of these bind to iron in heme without changing its oxidation state, but they nevertheless inhibit oxygen-binding, causing grave toxicity.

Hemoglobin The iron atom in the heme group must initially be in the ferrous (Fe2+) oxidation state to support oxygen and other gases' binding and transport (it temporarily switches to ferric during the time oxygen is bound, as explained above). Initial oxidation to the ferric (Fe3+) state without oxygen converts hemoglobin into "hemiglobin" or methemoglobin (pronounced "MET-hemoglobin"), which cannot bind oxygen. Hemoglobin in normal red blood cells is protected by a reduction system to keep this from happening. Nitric oxide is capable of converting a small fraction of hemoglobin to methemoglobin in red blood cells. The latter reaction is a remnant activity of the more ancient nitric oxide dioxygenase function of globins.

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Allosteric
Further information: Oxygen-hemoglobin dissociation curve Carbon dioxide occupies a different binding site on the hemoglobin. Carbon dioxide is more readily dissolved in deoxygenated blood, facilitating its removal from the body after the oxygen has been released to tissues undergoing metabolism. This increased affinity for carbon dioxide by the venous blood is known as the Haldane effect. Through the enzyme carbonic anhydrase, carbon dioxide reacts with water to give carbonic acid, which decomposes into bicarbonate and protons: CO2 + H2O H2CO3 HCO3- + H+ Hence blood with high carbon dioxide levels is also lower in pH (more acidic). Hemoglobin can bind protons and carbon dioxide, which causes a conformational change in the protein and facilitates the release of oxygen. Protons bind at various places on the protein, while carbon dioxide binds at the -amino group.[42] Carbon dioxide binds to hemoglobin and forms carbaminohemoglobin.[43] This decrease in hemoglobin's affinity for oxygen by the binding of carbon dioxide and acid is known as the Bohr effect (shifts the O2-saturation curve to the right). Conversely, when the carbon dioxide levels in the blood decrease (i.e., in the lung capillaries), carbon dioxide and protons are released from hemoglobin, increasing the oxygen The sigmoidal shape of hemoglobin's oxygen-dissociation curve results affinity of the protein. A reduction in the total from cooperative binding of oxygen to hemoglobin. binding capacity of hemoglobin to oxygen (i.e. shifting the curve down, not just to the right) due to reduced pH is called the root effect. This is seen in bony fish. It is necessary for hemoglobin to release the oxygen that it binds; if not, there is no point in binding it. The sigmoidal curve of hemoglobin makes it efficient in binding (taking up O2 in lungs), and efficient in unloading (unloading O2 in tissues).[44] In people acclimated to high altitudes, the concentration of 2,3-Bisphosphoglycerate (2,3-BPG) in the blood is increased, which allows these individuals to deliver a larger amount of oxygen to tissues under conditions of lower oxygen tension. This phenomenon, where molecule Y affects the binding of molecule X to a transport molecule Z, is called a heterotropic allosteric effect. A variant hemoglobin, called fetal hemoglobin (HbF, 22), is found in the developing fetus, and binds oxygen with greater affinity than adult hemoglobin. This means that the oxygen binding curve for fetal hemoglobin is left-shifted (i.e., a higher percentage of hemoglobin has oxygen bound to it at lower oxygen tension), in comparison to that of adult hemoglobin. As a result, fetal blood in the placenta is able to take oxygen from maternal blood.

Hemoglobin Hemoglobin also carries nitric oxide in the globin part of the molecule. This improves oxygen delivery in the periphery and contributes to the control of respiration. NO binds reversibly to a specific cysteine residue in globin; the binding depends on the state (R or T) of the hemoglobin. The resulting S-nitrosylated hemoglobin influences various NO-related activities such as the control of vascular resistance, blood pressure and respiration. NO is not released in the cytoplasm of erythrocytes but transported by an anion exchanger called AE1 out of them.[45] A study was performed to examine the influence of the form of hemoglobin (Hb) on the partitioning of inhaled volatile organic compounds (VOCs) into [human and animal] blood. Benzene was the prototypic VOC used in the investigations for this research due to the similar properties it shares with many other VOCs. To be specific, this study analyses the influence of the water solubility of Hb on the partitioning coefficient (PC) of a VOC as compared to the influence of the species or form of Hb. The different forms of blood used include: human hemoglobin (HbA), rat Hb, and sickle-cell hemoglobin (HbS). Rat Hb contains little water and is in a quasi-crystalline form, found inside the red blood cells (RBC), meaning they are more hydrophobic than human Hb, which are water-soluble. Sickle-cell hemoglobin (HbS) is water-soluble, however it can become water-insoluble, forming hydrophobic polymers, when deoxygenated. The findings state that the benzene PC for rat Hb was much higher than human that for Hb; however, the tests that measured the PCs of the oxygenated and deoxygenated forms of HbA and HbS did not differ, indicating that the affinity of benzene was not affected by the water solubility of Hb.[46]

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Types in humans
Hemoglobin variants are a part of the normal embryonic and fetal development, but may also be pathologic mutant forms of hemoglobin in a population, caused by variations in genetics. Some well-known hemoglobin variants such as sickle-cell anemia are responsible for diseases, and are considered hemoglobinopathies. Other variants cause no detectable pathology, and are thus considered non-pathological variants.[47] [48] In the embryo: Gower 1 (22) Gower 2 (22) (PDB 1A9W [49]) Hemoglobin Portland (22) In the fetus: Hemoglobin F (22) (PDB 1FDH [50]) In adults: Hemoglobin A (22) (PDB 1BZ0 [51]) - The most common with a normal amount over 95% Hemoglobin A2 (22) - chain synthesis begins late in the third trimester and in adults, it has a normal range of 1.5-3.5% Hemoglobin F (22) - In adults Hemoglobin F is restricted to a limited population of red cells called F-cells. However, the level of Hb F can be elevated in persons with sickle-cell disease and beta-thalassemia.

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Variant forms that cause disease: Hemoglobin H (4) - A variant form of hemoglobin, formed by a tetramer of chains, which may be present in variants of thalassemia. Hemoglobin Barts (4) - A variant form of hemoglobin, formed by a tetramer of chains, which may be present in variants of thalassemia. Hemoglobin S (2S2) - A variant form of hemoglobin found in people with sickle cell disease. There is a variation in the -chain gene, causing a change in the properties of hemoglobin, which results in sickling of red blood cells.

Hemoglobin C (2C2) - Another variant due to a variation in the -chain gene. This variant causes a mild chronic hemolytic anemia.

Gene expression of hemoglobin before and after birth. Also identifies the types of cells and organs in which the gene expression (data on Wood W.G., (1976). Br. Med. Bull. 32, 282.)

Hemoglobin E (2E2) - Another variant due to a variation in the -chain gene. This variant causes a mild chronic hemolytic anemia. Hemoglobin AS - A heterozygous form causing Sickle cell trait with one adult gene and one sickle cell disease gene Hemoglobin SC disease - Another heterozygous form with one sickle gene and another encoding Hemoglobin C.

Degradation in vertebrate animals

When red cells reach the end of their life due to aging or defects, they are broken down, the hemoglobin molecule is broken up and the iron gets recycled. When the porphyrin ring is broken up, the fragments are normally secreted in the bile by the liver. This process also produces one molecule of carbon monoxide for every molecule of heme degraded.[52] This is one of the few natural sources of carbon monoxide production in the human body, and is responsible for the normal blood levels of carbon monoxide even in people breathing pure air. The other major final product of heme degradation is bilirubin. Increased levels of this chemical are detected in the blood if red cells are being destroyed more rapidly than usual. Improperly degraded hemoglobin protein or hemoglobin that has been released from the blood cells too rapidly can clog small blood vessels, especially the delicate blood filtering vessels of the kidneys, causing kidney damage.

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Role in disease
Hemoglobin deficiency can be caused either by decreased amount of hemoglobin molecules, as in anemia, or by decreased ability of each molecule to bind oxygen at the same partial pressure of oxygen. Hemoglobinopathies (genetic defects resulting in abnormal structure of the hemoglobin molecule)[53] may cause both. In any case, hemoglobin deficiency decreases blood oxygen-carrying capacity. Hemoglobin deficiency is, in general, strictly distinguished from hypoxemia, defined as decreased partial pressure of oxygen in blood,[54] [55] [56] [57] although both are causes of hypoxia (insufficient oxygen supply to tissues). Other common causes of low hemoglobin include loss of blood, nutritional deficiency, bone marrow problems, chemotherapy, kidney failure, or abnormal hemoglobin (such as that of sickle-cell disease). High hemoglobin levels may be caused by exposure to high altitudes, smoking, dehydration, or tumors.[31] The ability of each hemoglobin molecule to carry oxygen is normally modified by altered blood pH or CO2, causing an altered oxygenhemoglobin dissociation curve. However, it can also be pathologically altered in, e.g., carbon monoxide poisoning.

Decrease of hemoglobin, with or without an absolute decrease of red blood cells, leads to symptoms of anemia. Anemia has many different causes, although iron deficiency and its resultant iron deficiency anemia are the most common causes in the Western world. As absence of iron decreases heme synthesis, red blood cells in iron deficiency anemia are hypochromic (lacking the red hemoglobin pigment) and microcytic (smaller than normal). Other anemias are rarer. In hemolysis (accelerated breakdown of red blood cells), associated jaundice is caused by the hemoglobin metabolite bilirubin, and the circulating hemoglobin can cause renal failure. Some mutations in the globin chain are associated with the hemoglobinopathies, such as sickle-cell disease and thalassemia. Other mutations, as discussed at the beginning of the article, are benign and are referred to merely as hemoglobin variants. There is a group of genetic disorders, known as the porphyrias that are characterized by errors in metabolic pathways of heme synthesis. King George III of the United Kingdom was probably the most famous porphyria sufferer. To a small extent, hemoglobin A slowly combines with glucose at the terminal valine (an alpha aminoacid) of each chain. The resulting molecule is often referred to as Hb A1c. As the concentration of glucose in the blood increases, the percentage of Hb A that turns into Hb A1c increases. In diabetics whose glucose usually runs high, the percent Hb A1c also runs high. Because of the slow rate of Hb A combination with glucose, the Hb A1c percentage is representative of glucose level in the blood averaged over a longer time (the half-life of red blood cells, which is typically 5055 days). Glycosylated hemoglobin is the form of hemoglobin to which glucose is bound. The binding of glucose to amino acids in the hemoglobin takes place spontaneously (without the help of an enzyme) in many proteins, and is not known to serve a useful purpose. However, the binding to hemoglobin does serve as a record for average blood glucose levels over the lifetime of red cells, which is approximately 120 days. The levels of glycosylated hemoglobin are therefore measured in order to monitor the long-term control of the chronic disease of type 2 diabetes mellitus (T2DM). Poor control of T2DM results in high levels of glycosylated hemoglobin in the red blood cells. The normal

In sickle cell hemoglobin (HbS) glutamic acid in position 6 (in beta chain) is mutated to valine. This change allows the deoxygenated form of the hemoglobin to stick to itself.

Hemoglobin reference range is approximately 45.9 %. Though difficult to obtain, values less than 7 % are recommended for people with T2DM. Levels greater than 9 % are associated with poor control of the glycosylated hemoglobin, and levels greater than 12 % are associated with very poor control. Diabetics who keep their glycosylated hemoglobin levels close to 7 % have a much better chance of avoiding the complications that may accompany diabetes (than those whose levels are 8 % or higher).[58] Elevated levels of hemoglobin are associated with increased numbers or sizes of red blood cells, called polycythemia. This elevation may be caused by congenital heart disease, cor pulmonale, pulmonary fibrosis, too much erythropoietin, or polycythemia vera.[59] Elevation in levels of hemoglobin were found in one study of the yogic practice of Yoga Nidra (yogic sleep) for half an hour daily.[60] A recent study done in Pondicherry, India, shows its importance in coronary artery disease.[61]

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Diagnostic uses
Hemoglobin concentration measurement is among the most commonly performed blood tests, usually as part of a complete blood count. For example it is typically tested before or after blood donation. Results are reported in g/L, g/dL or mol/L. 1 g/dL equals about 0.6206 mmol/L.[62] Normal levels are: Men: 13.8 to 18.0 g/dL (138 to 182 g/L, or 8.56 to 11.3mmol/L) Women: 12.1 to 15.1 g/dL (121 to 151 g/L, or 7.51 to 9.37mmol/L) Children: 11 to 16 g/dL (111 to 160 g/L, or 6.83 to 9.93mmol/L) Pregnant women: 11 to 12 g/dL (110 to 120 g/L, or 6.83 to 7.45mmol/L) [63] [64]

Normal values of hemoglobin in the 1st and 3rd trimesters of pregnant women must be at least 11 g/dL and at least 10.5 g/dL during the 2nd trimester.[65] Dehydration or hyperhydration can greatly influence measured hemoglobin levels. Albumin can indicate hydration status. If the concentration is below normal, this is called anemia. Anemias are classified by the size of red blood cells, the cells that contain hemoglobin in vertebrates. The anemia is called "microcytic" if red cells are small, "macrocytic" if they are large, and "normocytic" otherwise. Hematocrit, the proportion of blood volume occupied by red blood cells, is typically about three times the hemoglobin level. For example, if the hemoglobin is measured at 17, that compares with a hematocrit of 51.[66] Laboratory hemoglobin test methods require a blood sample (arterial, venous, or capillary) and analysis on hematology analyzer and CO-oximeter. Additionally, a new noninvasive hemoglobin (SpHb) test method called Pulse CO-Oximetry is also available with comparable accuracy to invasive methods.[67] Long-term control of blood sugar concentration can be measured by the concentration of Hb A1c. Measuring it directly would require many samples because blood sugar levels vary widely through the day. Hb A1c is the product of the irreversible reaction of hemoglobin A with glucose. A higher glucose concentration results in more Hb A1c. Because the reaction is slow, the Hb A1c proportion represents glucose level in blood averaged over the half-life of red blood cells, is typically 5055 days. An Hb A1c proportion of 6.0% or less show good long-term glucose control, while values above 7.0% are elevated. This test is especially useful for diabetics.[68] The functional magnetic resonance imaging (fMRI) machine uses the signal from deoxyhemoglobin, which is sensitive to magnetic fields since it is paramagnetic.

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Analogues in non-vertebrate organisms

A variety of oxygen-transport and -binding proteins exist in organisms throughout the animal and plant kingdoms. Organisms including bacteria, protozoans, and fungi all have hemoglobin-like proteins whose known and predicted roles include the reversible binding of gaseous ligands. Since many of these proteins contain globins and the heme moiety (iron in a flat porphyrin support), they are often called hemoglobins, even if their overall tertiary structure is very different from that of vertebrate hemoglobin. In particular, the distinction of myoglobin and hemoglobin in lower animals is often impossible, because some of these organisms do not contain muscles. Or, they may have a recognizable separate circulatory system but not one that deals with oxygen transport (for example, many insects and other arthropods). In all these groups, heme/globin-containing molecules (even monomeric globin ones) that deal with gas-binding are referred to as oxyhemoglobins. In addition to dealing with transport and sensing of oxygen, they may also deal with NO, CO2, sulfide compounds, and even O2 scavenging in environments that must be anaerobic. They may even deal with detoxification of chlorinated materials in a way analogous to heme-containing P450 enzymes and peroxidases. The structure of hemoglobins varies across species. Hemoglobin occurs in all kingdoms of organisms, but not in all organisms. Primitive species such as bacteria, protozoa, algae, and plants often have single-globin hemoglobins. Many nematode worms, molluscs, and crustaceans contain very large multisubunit molecules, much larger than those in vertebrates. In particular, chimeric hemoglobins found in fungi and giant annelids may contain both globin and other types of proteins.[69]
The giant tube worm Riftia pachyptila showing

One of the most striking occurrences and uses of hemoglobin in red hemoglobin-containing plumes organisms is in the giant tube worm (Riftia pachyptila, also called Vestimentifera), which can reach 2.4 meters length and populates ocean volcanic vents. Instead of a digestive tract, these worms contain a population of bacteria constituting half the organism's weight. The bacteria react with H2S from the vent and O2 from the water to produce energy to make food from H2O and CO2. The worms end with a deep red fan-like structure ("plume"), which extends into the water and absorbs H2S and O2 for the bacteria, and CO2 for use as synthetic raw material similar to photosynthetic plants. The structures are bright-red due to their containing several extraordinarily complex hemoglobins that have up to 144 globin chains, each including associated heme structures. These hemoglobins are remarkable for being able to carry oxygen in the presence of sulfide, and even to carry sulfide, without being completely "poisoned" or inhibited by it as hemoglobins in most other species are.[70] [71]

Other oxygen-binding proteins

Myoglobin Found in the muscle tissue of many vertebrates, including humans, it gives muscle tissue a distinct red or dark gray color. It is very similar to hemoglobin in structure and sequence, but is not a tetramer; instead, it is a monomer that lacks cooperative binding. It is used to store oxygen rather than transport it. Hemocyanin The second most common oxygen-transporting protein found in nature, it is found in the blood of many arthropods and molluscs. Uses copper prosthetic groups instead of iron heme groups and is blue in color when oxygenated. Hemerythrin

Hemoglobin Some marine invertebrates and a few species of annelid use this iron-containing non-heme protein to carry oxygen in their blood. Appears pink/violet when oxygenated, clear when not. Chlorocruorin Found in many annelids, it is very similar to erythrocruorin, but the heme group is significantly different in structure. Appears green when deoxygenated and red when oxygenated. Vanabins Also known as vanadium chromagens, they are found in the blood of sea squirts. There were once hypothesized to use the rare metal vanadium as an oxygen binding prosthetic group. However, although they do contain vanadium by preference, they apparently bind little oxygen, and thus have some other function, which has not been elucidated (sea squirts also contain some hemoglobin). They may act as toxins. Erythrocruorin Found in many annelids, including earthworms, it is a giant free-floating blood protein containing many dozenspossibly hundredsof iron- and heme-bearing protein subunits bound together into a single protein complex with a molecular mass greater than 3.5 million daltons. Pinnaglobin Only seen in the mollusc Pinna squamosa. Brown manganese-based porphyrin protein. Leghemoglobin In leguminous plants, such as alfalfa or soybeans, the nitrogen fixing bacteria in the roots are protected from oxygen by this iron heme containing oxygen-binding protein. The specific enzyme protected is nitrogenase, which is unable to reduce nitrogen gas in the presence of free oxygen. Coboglobin A synthetic cobalt-based porphyrin. Coboprotein would appear colorless when oxygenated, but yellow when in veins.

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Presence in nonerythroid cells

Some nonerythroid cells (i.e., cells other than the red blood cell line) contain hemoglobin. In the brain, these include the A9 dopaminergic neurons in the substantia nigra, astrocytes in the cerebral cortex and hippocampus, and in all mature oligodendrocytes.[10] It has been suggested that brain hemoglobin in these cell may enable the "storage of oxygen to provide a homeostatic mechanism in anoxic conditions, which is especially important for A9 DA neurons that have an elevated metabolism with a high requirement for energy production".[10] It has been noted further that "A9 dopaminergic neurons may be at particular risk since in addition to their high mitochondrial activity they are under intense oxidative stress caused by the production of hydrogen peroxide via autoxidation and/or monoamine oxidase (MAO)-mediated deamination of dopamine and the subsequent reaction of accessible ferrous iron to generate highly toxic hydroxyl radicals".[10] This may explain the risk of these cells for degeneration in Parkinson's disease.[10] The presence of iron from hemoglobin in these cells also results in the post-mortem darkness of these cells, which is the origin of the Latin name, substantia nigra. Outside the brain, hemoglobin has non-oxygen-carrying functions as an antioxidant and a regulator of iron metabolism in macrophages,[72] alveolar cells,[73] and mesangial cells in the kidney.[74]

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137

In history, art and music

Historically, the color of blood was associated with rust, as ancient Romans associated the planet Mars with the god of war since Mars is orange-red. The color of Mars is due to the iron oxide in the Martian soil, but the red in blood is not due to the iron in hemoglobin and its oxides, which is a common misconception. The red is due to the porphyrin moiety of hemoglobin to which the iron is bound, not the iron itself,[75] although the ligation and redox state of the iron can influence the pi to pi* or n to pi* electronic transitions of the porphyrin and hence its optical characteristics.

The planet Mars

Artist Julian Voss-Andreae created a sculpture called "Heart of Steel (Hemoglobin)" in 2005, based on the protein's backbone. The sculpture was made from glass and weathering steel. The intentional rusting of the initially shiny work of art mirrors hemoglobin's fundamental chemical reaction of oxygen binding to iron.[76] [77] Rock band Placebo recorded a song Heart of Steel (Hemoglobin) (2005) by Julian Voss-Andreae. The images show the 5' called "Haemoglobin" with the lyrics (1.60m) tall sculpture right after installation, after 10 days, and after several months of exposure to the elements. "Haemoglobin is the key to a healthy heartbeat". French rap artist MC Solaar also had a successful single titled "La Concubine de L'Hemoglobin" in 1994.

References
[1] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1GZX [2] [3] [4] [5] http:/ / www. proteopedia. org/ wiki/ index. php/ Hemoglobin http:/ / www. ncbi. nlm. nih. gov/ Omim/ getmap. cgi?chromosome=16p13. 3 http:/ / www. ncbi. nlm. nih. gov/ Omim/ getmap. cgi?chromosome=11p15. 5 Maton, Anthea; Jean Hopkins, Charles William McLaughlin, Susan Johnson, Maryanna Quon Warner, David LaHart, Jill D. Wright (1993). Human Biology and Health. Englewood Cliffs, New Jersey, USA: Prentice Hall. ISBN0-13-981176-1. [6] Sidell, Bruce; Kristin O'Brien (2006). "When bad things happen to good fish: the loss of hemoglobin and myglobin expression in Antarctic icefishes". The Journal of Experimental Biology 209 (Pt 10): 17911802. doi:10.1242/jeb.02091. PMID16651546. [7] Dominguez de Villota ED, Ruiz Carmona MT, Rubio JJ, de Andrs S (December 1981). "Equality of the in vivo and in vitro oxygen-binding capacity of haemoglobin in patients with severe respiratory disease". Br J Anaesth 53 (12): 13258. doi:10.1093/bja/53.12.1325. ISSN0007-0912. PMID7317251. [8] Costanzo, Linda S. (2007). Physiology. Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-7311-3. [9] Respiratory Function of Hemoglobin. Connie C.W. Hsia, M.D. N Engl J Med 1998; 338:239-248 January 22, 1998 [10] Biagioli M, Pinto M, Cesselli D, Zaninello M, Lazarevic D, Roncaglia P, Simone R, Vlachouli C, Plessy C, Bertin N, Beltrami A, Kobayashi K, Gallo V, Santoro C, Ferrer I, Rivella S, Beltrami CA, Carninci P, Raviola E, Gustincich S. (2009). Unexpected expression of {alpha}- and {beta}-globin in mesencephalic dopaminergic neurons and glial cells. (http:/ / www. pubmedcentral. nih. gov/ picrender. fcgi?artid=2732704& blobtype=pdf) Proc Natl Acad Sci U S A. 106:1545415459. PMID 19717439 doi:10.1073/pnas.0813216106 [11] Hnefeld F.L. (1840). Die Chemismus in der thierischen Organization. Leipzig. [12] Funke O (1851). "ber das milzvenenblut". Z Rat Med 1: 172218.

Hemoglobin
[13] "A NASA Recipe For Protein Crystallography" (http:/ / www. okcareertech. org/ cimc/ special/ nochild/ downloads/ science/ Protein. Crystallography. pdf) (PDF). Educational Brief. National Aeronautics and Space Administration. . Retrieved 2008-10-12. [14] Hoppe-Seyler F (1866). "ber die oxydation in lebendem blute". Med-chem Untersuch Lab 1: 133140. [15] Perutz, M.F.; Rossmann, M.G.; Cullis, A.F.; Muirhead, H.; Will, G.; North, A.C.T. (1960). "Structure of H". Nature 185 (4711): 416422. doi:10.1038/185416a0. PMID18990801. [16] Perutz MF (November 1960). "Structure of haemoglobin". Brookhaven symposia in biology 13: 16583. ISSN0068-2799. PMID13734651. [17] A Syllabus of Human Hemoglobin Variants (1996) (http:/ / globin. cse. psu. edu/ html/ huisman/ variants/ ) [18] Hemoglobin Variants (http:/ / www. labtestsonline. org/ understanding/ analytes/ hemoglobin_var/ glance-3. html) [19] Uthman, MD, Ed. "Hemoglobinopathies and Thalassemias" (http:/ / web2. airmail. net/ uthman/ hemoglobinopathy/ hemoglobinopathy. html). . Retrieved 2007-12-26. [20] Reed, Leslie. "Adaptation found in mouse genes." Omaha World-Herald 11 Aug. 2009: EBSCO. Web. 30 Oct. 2009. [21] "Mammoths had anti-freeze blood" (http:/ / news. bbc. co. uk/ 2/ hi/ science/ nature/ 8657464. stm). BBC. 2010-05-02. . Retrieved 2010-05-02. [22] "Hemoglobin Synthesis" (http:/ / sickle. bwh. harvard. edu/ hbsynthesis. html). April 14, 2002. . Retrieved 2007-12-26. [23] van Kessel et al. "2.4 Proteins - Natural Polyamides." Chemistry 12. Toronto: Nelson, 2003. 122. Print. [24] "Hemoglobin Tutorial." University of Massachusetts Amherst. N.p., n.d. Web. 23 Oct. 2009. <http://www.umass.edu/molvis/tutorials/hemoglobin/index.htm>. [25] Steinberg 2001, p.95 [26] Hardison 1996, p.1 [27] "Hemoglobin." School of Chemistry - Bristol University - UK. N.p., n.d. Web. 12 Oct. 2009. <http://www.chm.bris.ac.uk/motm/hemoglobin/hemoglobjm.htm>. [28] WikiPremed > Coordination Chemistry (http:/ / wikipremed. com/ interdisciplinary_course. php?code=0213000100000000) Retrieved on July 2, 2009 [29] Linberg R, Conover CD, Shum KL, Shorr RG (Mar 1998). "Hemoglobin based oxygen carriers: how much methemoglobin is too much?". Artif Cells Blood Substit Immobil Biotechnol 26 (2): 13348. doi:10.3109/10731199809119772. ISSN1073-1199. PMID9564432. [30] Van Beekvelt MC, Colier WN, Wevers RA, Van Engelen BG (Feb 2001). "Performance of near-infrared spectroscopy in measuring local O2 consumption and blood flow in skeletal muscle" (http:/ / jap. physiology. org/ cgi/ pmidlookup?view=long& pmid=11160049). J Appl Physiol 90 (2): 511519. ISSN8750-7587. PMID11160049. . [31] "Hemoglobin." MedicineNet. N.p., n.d. Web. 12 Oct. 2009. <www.medicinenet.com/hemoglobin/article.htm>. [32] "Hemoglobin Home." Biology @ Davidson. N.p., n.d. Web. 12 Oct. 2009. <http://www.bio.davidson.edu/Courses/Molbio/MolStudents/spring2005/Heiner/hemoglobin.html>. [33] Baillie/Simpson. "Online model of the hemoglobin binding and the effects of hyperventilation" (http:/ / www. altitude. org/ hemoglobin_saturation. php). . Retrieved 2006-08-10. [34] Pin S, Alpert B, Michalowicz A. (Oct 1982). "Oxygen bonding in human hemoglobin and its isolated subunits: A XANES study". FEBS Lett. 147 (1): 10610. doi:10.1016/0014-5793(82)81021-1. PMID7140986. [35] S Pin, P Valat, R Cortes, A Michalowicz, and B Alpert (December 1985). "Ligand binding processes in hemoglobin. Chemical reactivity of iron studied by XANES spectroscopy". Biophys J. 48(6): 99710. [36] A. Bianconi, A. Congiu-Castellanoa, M. Dell'Aricciaa, A. Giovannellia, E. Burattinib and P. J. Durhamc (August 1985). "Increase of the Fe effective charge in hemoproteins during oxygenation process". Biochemical and Biophysical Research Communications 30 (1): 98102. doi:10.1016/0006-291X(85)91775-9. PMID4038310. [37] Childs PE (2001). "Haemoglobin - a molecular lung: 2" (http:/ / www. ul. ie/ ~childsp/ CinA/ Issue65/ TOC28_Haemoglobin. htm). Chemistry in Action (65). ISSN0332-2637. . [38] Chen H, Ikeda-Saito M, Shaik S (11 2008). "Nature of the Fe-O2 bonding in oxy-myoglobin: effect of the protein" (http:/ / pubs. acs. org/ doi/ abs/ 10. 1021/ ja805434m). Journal of the American Chemical Society 130 (44): 1477814790. doi:10.1021/ja805434m. PMID18847206. . [39] Jensen, Frank B (4/9/2009). "The dual roles of red blood cells in tissue oxygen delivery: oxygen carriers and regulators of local blood flow". Journal of Experimental Biology (The Company of Biologists) 212 (Pt 21): 33873393. doi:10.1242/jeb.023697. PMID19837879. [40] Chou KC (June 1989). "Low-frequency resonance and cooperativity of hemoglobin". Trends Biochem. Sci. 14 (6): 2123. doi:10.1016/0968-0004(89)90026-1. PMID2763333. [41] Guyton A C: Medical Physiology 12ed. 2010, page 502 [42] Nelson, D. L.; Cox, M. M. (2000). Lehninger Principles of Biochemistry, 3rd ed. New York, NY: Worth Publishers. p 217 [43] Guyton, Arthur C.; John E. Hall (2006). Textbook of Medical Physiology (11 ed.). Philadelphia: Elsevier Saunders. p.511. ISBN0721602401. [44] "YouTube - Lecture - 12 Myoglobin and Hemoglobin." YouTube - Broadcast Yourself.. N.p., n.d. Web. 30 Oct. 2009. <http://www.youtube.com/watch?v=6AfRX6oh9-E>. [45] Rang, H.P.; Dale M.M., Ritter J.M., Moore P.K. (2003). Pharmacology, Fifth Edition. Elsevier. ISBN0443072027. [46] Wiester et al. "Partitioning of Benzene in Blood: Influence of Hemoglobin Type in Humans and Animals." Environmental Health Perspectives 110.3 (2002): p255-261. EBSCO. Web. 1 Nov. 2009.

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[47] "Hemoglobin Variants" (http:/ / www. labtestsonline. org/ understanding/ analytes/ hemoglobin_var/ glance-3. html). Lab Tests Online. American Association for Clinical Chemistry. 2007-11-10. . Retrieved 2008-10-12. [48] Huisman THJ (1996). "A Syllabus of Human Hemoglobin Variants" (http:/ / globin. cse. psu. edu/ html/ huisman/ variants/ ). Globin Gene Server. Pennsylvania State University. . Retrieved 2008-10-12. [49] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1A9W [50] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1FDH [51] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1BZ0 [52] Kikuchi, G.; Yoshida, T.; Noguchi, M. (2005). "Heme oxygenase and heme degradation". Biochemical and Biophysical Research Communications 338 (1): 558567. doi:10.1016/j.bbrc.2005.08.020. PMID16115609. [53] " hemoglobinopathy (http:/ / web. archive. org/ web/ 20090616022448/ http:/ / www. mercksource. com/ pp/ us/ cns/ cns_hl_dorlands_split. jsp?pg=/ ppdocs/ us/ common/ dorlands/ dorland/ four/ 000048231. htm)" at Dorland's Medical Dictionary [54] britannica.com --> blood disease (http:/ / www. britannica. com/ EBchecked/ topic/ 280141/ hypoxemia), stating hypoxemia (reduced oxygen tension in the blood). Retrieved on May 25, 2009 [55] Biology-Online.org --> Dictionary H Hypoxemia (http:/ / www. biology-online. org/ dictionary/ Hypoxemia) last modified 00:05, 29 December 2008 [56] Page 430 -> Pathophysiology of acute respiratory failure (http:/ / books. google. dk/ books?id=3H3AIEtvc8YC& pg=PA430& lpg=PA430& dq=hypoxemia+ definition+ "partial+ pressure"& source=bl& ots=p3N6uD-dVb& sig=UvR-_OjG_K-1y4yId6PIBw7owXg& hl=en& ei=QUAaSqL0OofU-QbNw-XLDg& sa=X& oi=book_result& ct=result& resnum=6) in Trauma By William C. Wilson, Christopher M. Grande, David B. Hoyt Edition: illustrated Published by CRC Press, 2007 ISBN 0-8247-2920-X, 9780824729202 1384 pages [57] Hazards of hypoxemia: How to protect your patient from low oxygen levels (http:/ / findarticles. com/ p/ articles/ mi_qa3689/ is_199605/ ai_n8735092/ ) In Nursing , May 1996 by McGaffigan, Patricia A [58] "Definition of Glycosylated Hemoglobin." Medicine Net. N.p., n.d. Web. 12 Oct. 2009. <www.medterms.com/script/main/art.asp?articlekey=16295>. [59] Hemoglobin (http:/ / www. nlm. nih. gov/ medlineplus/ ency/ article/ 003645. htm#What abnormal results mean) at Medline Plus [60] Kumar, Dr. Kamakhya; The Healing Sleep, Yoga, Mind Body Spirit; Yoga Magazine, 26 York Street London, Vol. 50 page 42-44. [61] Padmanaban P, Toora BD. Hemoglobin: Emerging marker in stable coronary artery disease. Chron Young Sci [serial online] 2011 [cited 2011 Jul 24];2:109-10. Available from: http:/ / www. cysonline. org/ text. asp?2011/ 2/ 2/ 109/ 82971. [62] http:/ / www. unc. edu/ ~rowlett/ units/ scales/ clinical_data. html [63] Hemoglobin Level Test (http:/ / ibdcrohns. about. com/ od/ diagnostictesting/ p/ testhemo. htm) [64] Although other sources can have slightly differing values, such as http:/ / www. gpnotebook. co. uk/ simplepage. cfm?ID=1026883654 [65] Murray S.S. & McKinney E.S.(2006). Foundations of Maternal-Newborn Nursing.(4th ed., p 919).Philadelphia: Saunders Elsevier [66] "Hematocrit (HCT) or Packed Cell Volume (PCV)" (http:/ / www. doctorslounge. com/ hematology/ labs/ hematocrit. htm). DoctorsLounge.com. . Retrieved 2007-12-26. [67] Frasca D., Dahyot-Fizelier C., Catherine K., Levrat Q., Debaene B., Mimoz O. Crit Care Med. 2011 Oct;39(10):2277-82. [68] This Hb A1c level is only useful in individuals who have red blood cells (RBCs) with normal survivals (i.e., normal half-life). In individuals with abnormal RBCs, whether due to abnormal hemoglobin molecules (such as Hemoglobin S in Sickle Cell Anemia) or RBC membrane defects - or other problems, the RBC half-life is frequently shortened. In these individuals, an alternative test called "fructosamine level" can be used. It measures the degree of glycation (glucose binding) to albumin, the most common blood protein, and reflects average blood glucose levels over the previous 18-21 days, which is the half-life of albumin molecules in the circulation. [69] Weber RE, Vinogradov SN (Apr 2001). "Nonvertebrate hemoglobins: functions and molecular adaptations" (http:/ / physrev. physiology. org/ cgi/ pmidlookup?view=long& pmid=11274340). Physiol. Rev. 81 (2): 569628. ISSN0031-9333. PMID11274340. . [70] Zal F, Lallier FH, Green BN, Vinogradov SN, Toulmond A (Apr 1996). "The multi-hemoglobin system of the hydrothermal vent tube worm Riftia pachyptila. II. Complete polypeptide chain composition investigated by maximum entropy analysis of mass spectra" (http:/ / www. jbc. org/ cgi/ pmidlookup?view=long& pmid=8621529). J. Biol. Chem. 271 (15): 887581. doi:10.1074/jbc.271.15.8875. ISSN0021-9258. PMID8621529. . [71] Minic Z, Herv G (Aug 2004). "Biochemical and enzymological aspects of the symbiosis between the deep-sea tubeworm Riftia pachyptila and its bacterial endosymbiont". Eur. J. Biochem. 271 (15): 3093102. doi:10.1111/j.1432-1033.2004.04248.x. ISSN0014-2956. PMID15265029. [72] Liu L, Zeng M, Stamler JS (June 1999). "Hemoglobin induction in mouse macrophages". Proceedings of the National Academy of Sciences of the United States of America 96 (12): 66437. doi:10.1073/pnas.96.12.6643. PMC21968. PMID10359765. [73] Newton DA, Rao KM, Dluhy RA, Baatz JE (March 2006). "Hemoglobin Is Expressed by Alveolar Epithelial Cells" (http:/ / www. jbc. org/ cgi/ reprint/ 281/ 9/ 5668). Journal of Biological Chemistry 281 (9): 566876. doi:10.1074/jbc.M509314200. PMID16407281. . [74] Nishi H, Inagi R, Kato H, et al. (August 2008). "Hemoglobin Is Expressed by Mesangial Cells and Reduces Oxidant Stress" (http:/ / www. pubmedcentral. nih. gov/ articlerender. fcgi?tool=pubmed& pubmedid=18448584). Journal of the American Society of Nephrology 19 (8): 15008. doi:10.1681/ASN.2007101085. PMC2488266. PMID18448584. . [75] Boh, Larry (2001). Pharmacy Practice Manual: A Guide to the Clinical Experience. Lippincott Williams & Wilkins. ISBN0781725410. [76] Holden, Constance (30 September 2005). "Blood and Steel" (http:/ / www. sciencemag. org/ cgi/ reprint/ 309/ 5744/ 2160d. pdf) (pdf). Science 309 (5744): 2160. doi:10.1126/science.309.5744.2160d. . [77] Moran L, Horton RA, Scrimgeour G, Perry M (2011). Principles of Biochemistry. Boston, MA: Pearson. pp.127. ISBN0-321-70733-8.

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Further reading
Campbell, MK (1999). Biochemistry (Third Edition). Harcourt. ISBN0-03-024426-9 Eshaghian, S; Horwich, TB; Fonarow, GC (January 2006). "An unexpected inverse relationship between HbA1c levels and mortality in patients with diabetes and advanced systolic heart failure". Am Heart J 151 (1): 91. doi:10.1016/j.ahj.2005.10.008. PMID16368297. Ganong, WF (2003). Review of Medical Physiology (Twenty-First Edition). Lange. ISBN0-07-140236-5. Hager, T (1995). Force of Nature: The Life of Linus Pauling. Simon and Schuster. ISBN0-684-80909-5. Hardison, RC (June 11, 1996). "A brief history of hemoglobins: plant, animal, protist, and bacteria" (http:/ / www. pubmedcentral. gov/ articlerender. fcgi?tool=pubmed& pubmedid=8650150). Proc Natl Acad Sci USA 93 (12): 56759. doi:10.1073/pnas.93.12.5675. PMC39118. PMID8650150. PMID 8650150. Kneipp, J; Balakrishnan, G; Chen, R, Shen TJ, Sahu SC, Ho NT, Giovannelli JL, Simplaceanu V, Ho C, Spiro TG (November 22, 2005). "Dynamics of allostery in hemoglobin: roles of the penultimate tyrosine H bonds". J Mol Biol. PMID 16368110. Steinberg, MH (2001). Disorders of Hemoglobin: Genetics, Pathophysiology, and Clinical Management (http:/ / books. google. com/ books?vid=ISBN0521632668). Cambridge University Press. ISBN0-521-63266-8.

External links
Interactive hemoglobin saturation curves (http://www.altitude.org/hemoglobin_saturation.php) Interactive models of hemoglobin (http://www.ufp.pt/~pedros/anim/2frame-hben.htm) (Requires MDL Chime (http://www.mdl.com/products/framework/chime/)) Hemoglobin. . (http://medzeit.ru/analizy/krov/ norma-gemoglobina-v-krovi.html) National Anemia Action Council (http://www.anemia.org/) - anemia.org New hemoglobin type causes mock diagnosis with pulse oxymeters (http://www.life-of-science.net/medicine/ news/new-hemoglobin-type-discovered-causing-mock-diagnosis-of-cardiac-insufficiency.html)

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Enzyme mechanisms
Enzyme catalysis
Enzyme catalysis is the catalysis of chemical reactions by specialized proteins known as enzymes. Catalysis of biochemical reactions in the cell is vital due to the very low reaction rates of the uncatalysed reactions. The mechanism of enzyme catalysis is similar in principle to other types of chemical catalysis. By providing an alternative reaction route and by stabilizing intermediates the enzyme reduces the energy required to reach the highest energy transition state of the reaction. The reduction of activation energy (Ea) increases the number of reactant molecules with enough energy to reach the activation energy and form the product.

Stabilization of the transition state by an enzyme.

Induced fit
The favored model for the enzyme-substrate interaction is the induced fit model.[1] This model proposes that the initial interaction between enzyme and substrate is relatively weak, but that these weak interactions rapidly induce conformational changes in the enzyme that strengthen binding.

Diagrams to show the induced fit hypothesis of enzyme action.

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Catalysis by induced fit

The advantages of the induced fit mechanism arise due to the stabilizing effect of strong enzyme binding. There are two different mechanisms of substrate binding: uniform binding, which has strong substrate binding, and differential binding, which has strong transition state binding. The stabilizing effect of uniform binding increases both substrate and transition state binding affinity, while differential binding increases only transition state binding affinity. Both are used by enzymes and have been evolutionarily chosen to minimize the Ea of the reaction. Enzymes which are saturated, that is, have a high affinity substrate binding, require differential binding to reduce the Ea, whereas small substrate unbound enzymes may use either differential or uniform binding. These effects have led to most proteins using the differential binding mechanism to reduce the Ea, so most proteins have high affinity of the enzyme to the transition state. Differential binding is carried out by the induced fit mechanism - the substrate first binds weakly, then the enzyme changes conformation increasing the affinity to the transition state and stabilizing it, so reducing the activation energy to reach it.

The different mechanisms of substrate binding

It is important to clarify, however, that the induced fit concept cannot be used to rationalize catalysis. That is, the chemical catalysis is defined as the reduction of Ea (when the system is already in the ES) relative to Ea in the uncatalyzed reaction in water (without the enzyme). The induced fit only suggests that the barrier is lower in the closed form of the enzyme but does not tell us what the reason for the barrier reduction is. Induced fit may be beneficial to the fidelity of molecular recognition in the presence of competition and noise via the conformational proofreading mechanism .[2]

Mechanisms of transition state stabilization

These conformational changes also bring catalytic residues in the active site close to the chemical bonds in the substrate that will be altered in the reaction. After binding takes place, one or more mechanisms of catalysis lowers the energy of the reaction's transition state, by providing an alternative chemical pathway for the reaction. There are six possible mechanisms of "over the barrier" catalysis as well as a "through the barrier" mechanism:

Catalysis by bond strain

This is the principal effect of induced fit binding, where the affinity of the enzyme to the transition state is greater than to the substrate itself. This induces structural rearrangements which strain substrate bonds into a position closer to the conformation of the transition state, so lowering the energy difference between the substrate and transition state and helping catalyze the reaction. However, the strain effect is, in fact, a ground state destabilization effect, rather than transition state stabilization effect.[3] [4] Furthermore, enzymes are very flexible and they cannot apply large strain effect.[5] In addition to bond strain in the substrate, bond strain may also be induced within the enzyme itself to activate residues in the active site.

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143

For example: Substrate, bound substrate, and transition state conformations of lysozyme.

The substrate, on binding, is distorted from the typical 'chair' hexose ring into the 'sofa' conformation, which is similar in shape to the transition state.

Catalysis by proximity and orientation

This increases the rate of the reaction as enzyme-substrate interactions align reactive chemical groups and hold them close together. This reduces the entropy of the reactants and thus makes reactions such as ligations or addition reactions more favorable, there is a reduction in the overall loss of entropy when two reactants become a single product. This effect is analogous to an effective increase in concentration of the reagents. The binding of the reagents to the enzyme gives the reaction intramolecular character, which gives a massive rate increase.
For example: Similar reactions will occur far faster if the reaction is intramolecular.

The effective concentration of acetate in the intramolecular reaction can be estimated as k2/k1 = 2 x 105 Molar.

However, the situation might be more complex, since modern computational studies have established that traditional examples of proximity effects cannot be related directly to enzyme entropic effects.[6] [7] [8] Also, the original entropic proposal[9] has been found to largely overestimate the contribution of orientation entropy to catalysis.[10]

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Catalysis involving proton donors or acceptors (Acid/Base Catalysis)

Proton donors and acceptors, i.e. acids and bases, may donate and accept protons in order to stabilize developing charges in the transition state. This typically has the effect of activating nucleophile and electrophile groups, or stabilizing leaving groups. Histidine is often the residue involved in these acid/base reactions, since it has a pKa close to neutral pH and can therefore both accept and donate protons. Many reaction mechanisms involving acid/base catalysis assume a substantially altered pKa. This alteration of pKa is possible through the local environment of the residue.
Conditions Hydrophobic environment Adjacent residues of like charge Salt bridge (and hydrogen bond) formation Acids Increase pKa Increase pKa Bases Decrease pKa Decrease pKa

Decrease pKa Increase pKa

The pKa is can be modified significantly by the environment, to the extent that residues which are basic in solution may act as proton donors, and vice versa.
For example: Serine protease catalytic mechanism

The initial step of the serine protease catalytic mechanism involves the histidine of the active site accepting a proton from the serine residue. This prepares the serine as a nucleophile to attack the amide bond of the substrate. This mechanism includes donation of a proton from serine (a base, pKa 14) to histidine (an acid, pKa 6), made possible due to the local environment of the bases.

It is important to clarify that the modification of the pKas is a pure part of the electrostatic mechanism.[4] Furthermore, the catalytic effect of the above example is mainly associated with the reduction of the pKa of the oxy anion and the increase in the pKa of the histidine, while the proton transfer from the serine to the histidine is not catalyzed significantly, since it is not the rate determining barrier.[11]

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Electrostatic catalysis
Stabilization of charged transition states can also be by residues in the active site forming ionic bonds (or partial ionic charge interactions) with the intermediate. These bonds can either come from acidic or basic side chains found on amino acids such as lysine, arginine, aspartic acid or glutamic acid or come from metal cofactors such as zinc. Metal ions are particularly effective and can reduce the pKa of water enough to make it an effective nucleophile. Systematic computer simulation studies established that electrostatic effects give, by far, the largest contribution to catalysis.[4] In particular, it has been found that enzyme provides an environment which is more polar than water, and that the ionic transition states are stabilized by fixed dipoles. This is very different from transition state stabilization in water, where the water molecules must pay with "reorganization energy".[12] in order to stabilize ionic and charged states. Thus, the catalysis is associated with the fact that the enzyme polar groups are preorganized
[13]

For example: Carboxypeptidase catalytic mechanism

The tetrahedral intermediate is stabilised by a partial ionic bond between the Zn2+ ion and the negative charge on the oxygen.

Covalent catalysis
Covalent catalysis involves the substrate forming a transient covalent bond with residues in the active site or with a cofactor. This adds an additional covalent intermediate to the reaction, and helps to reduce the energy of later transition states of the reaction. The covalent bond must, at a later stage in the reaction, be broken to regenerate the enzyme. This mechanism is found in enzymes such as proteases like chymotrypsin and trypsin, where an acyl-enzyme intermediate is formed. Schiff base formation using the free amine from a lysine residue is another mechanism, as seen in the enzyme aldolase during glycolysis. Some enzymes utilize non-amino acid cofactors such as pyridoxal phosphate (PLP) or thiamine pyrophosphate (TPP) to form covalent intermediates with reactant molecules.[14] [15] Such covalent intermediates function to reduce the energy of later transition states, similar to how covalent intermediates formed with active site amino acid residues allow stabilization, but the capabilities of cofactors allow enzymes to carryout reactions that amino acid side residues alone could not. Enzymes utilizing such cofactors include the PLP-dependent enzyme aspartate transaminase and the TPP-dependent enzyme pyruvate dehydrogenase.[16] [17] It is important to clarify that covalent catalysis does correspond in most cases to simply the use of a specific mechanism rather than to true catalysis.[4] For example, the energetics of the covalent bond to the serine molecule in chymotrypsin should be compared to the well-understood covalent bond to the nucleophile in the uncatalyzed solution reaction. A true proposal of a covalent catalysis (where the barrier is lower than the corresponding barrier in

Enzyme catalysis solution) would require, for example, a partial covalent bond to the transition state by an enzyme group (e.g., a very strong hydrogen bond), and such effects do not contribute significantly to catalysis.

146

Quantum tunneling
These traditional "over the barrier" mechanisms have been challenged in some cases by models and observations of "through the barrier" mechanisms (quantum tunneling). Some enzymes operate with kinetics which are faster than what would be predicted by the classical G. In "through the barrier" models, a proton or an electron can tunnel through activation barriers.[18] [19] Quantum tunneling for protons has been observed in tryptamine oxidation by aromatic amine dehydrogenase.[20] Interestingly, quantum tunneling does not appear to provide a major catalytic advantage, since the tunneling contributions are similar in the catalyzed and the uncatalyzed reactions in solution.[19] [21] [22] [23] However, the tunneling contribution (typically enhancing rate constants by a factor of ~1000[20] compared to the rate of reaction for the classical 'over the barrier' route) is likely crucial to the viability of biological organisms. This emphasizes the general importance of tunneling reactions in biology. In 1971-1972 the first quantum-mechanical model of enzyme catalysis was formulated.[24] [25]

Examples of catalytic mechanisms

In reality, most enzyme mechanisms involve a combination of several different types of catalysis.

Triose phosphate isomerase

Triose phosphate isomerase (EC 5.3.1.1 [26]) catalyses the reversible interconvertion of the two triose phosphates isomers dihydroxyacetone phosphate and D-glyceraldehyde 3-phosphate.

Trypsin
Trypsin (EC 3.4.21.4 residues.
[27]

) is a serine protease that cleaves protein substrates at lysine and arginine amino acid

Aldolase
Aldolase (EC 4.1.2.13 [28]) catalyses the breakdown of fructose 1,6-bisphosphate (F-1,6-BP) into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate (DHAP).

References
[1] Koshland DE (February 1958). "Application of a Theory of Enzyme Specificity to Protein Synthesis". Proc. Natl. Acad. Sci. U.S.A. 44 (2): 98104. doi:10.1073/pnas.44.2.98. PMC335371. PMID16590179. [2] Savir Y & Tlusty T (2007). Scalas, Enrico. ed. "Conformational Proofreading: The Impact of Conformational Changes on the Specificity of Molecular Recognition" (http:/ / www. weizmann. ac. il/ complex/ tlusty/ papers/ PLoSONE2007. pdf). PLoS ONE 2 (5): e468. doi:10.1371/journal.pone.0000468. PMC1868595. PMID17520027. . [3] Jencks W.P. "Catalysis in Chemistry and Enzymology" 1987, Dover, New York [4] Warshel, A.; Sharma, P.K.; Kato, M.; Xiang, Y.; Liu, H.; Olsson, M.H.M. (2006). "Electrostatic Basis of Enzyme Catalysis". Chem. Rev. 106 (8): 32103235. doi:10.1021/cr0503106. PMID16895325. [5] Warshel, A.; Levitt, M. (1976). "Theoretical Studies of Enzymatic Reactions: Dielectric Electrostatic and Steric Stabilization of the Carbonium Ion in the Reaction of Lysozyme". J. Mol. Biol. 103 (2): 22749. doi:10.1016/0022-2836(76)90311-9. PMID985660. [6] Stanton, R.V.; Perakyla, M.; Bakowies, D.; Kollman, P.A. (1998). "Combined ab initio and Free Energy Calculations To Study Reactions in Enzymes and Solution: Amide Hydrolysis in Trypsin and Aqueous Solution". J. Am. Chem. Soc 120 (14): 34483457. doi:10.1021/ja972723x. [7] Kuhn, B.; Kollman, P.A. (2000). "QM-FE and Molecular Dynamics Calculations on Catechol O-Methyltransferase: Free Energy of Activation in the Enzyme and in Aqueous Solution and Regioselectivity of the Enzyme-Catalyzed Reaction". J. Am. Chem. Soc 122 (11): 25862596. doi:10.1021/ja992218v.

Enzyme catalysis
[8] Bruice, T.C.; Lightstone, F.C. (1999). "Ground State and Transition State Contributions to the Rates of Intramolecular and Enzymatic Reactions". Acc. Chem. Res. 32 (2): 127136. doi:10.1021/ar960131y. [9] Page, M.I.; Jencks, W.P.. "Entropic Contributions to Rate Accelerations in Enzymic and Intramolecular Reactions and the Chelate Effect". Proc. Natl. Acad. Sci. USA 1971 (68): 16781683. doi:10.1073/pnas.68.8.1678. PMC389269. PMID5288752. [10] Warshel, A.; Parson, W.W. (2001). "Dynamics of Biochemical and Biophysical Reactions: Insight from Computer Simulations". Quart. Rev. Biophys 34: 563679. [11] Warshel, A.; Naray-Szabo, G.; Sussman, F.; Hwang, J.-K. (1989). "How do Serine Proteases Really Work?". Biochemistry 1989 (28): 362937. doi:10.1021/bi00435a001. PMID2665806. [12] Marcus R. A. "On the Theory of Electron-Transfer Reactions. VI. Unified Treatment for Homogeneous and Electrode Reactions" J. Chem. Phys. 1965 43:679-701 [13] Warshel A. "Energetics of Enzyme Catalysis", Proc. Natl. Acad. Sci. USA, 1978 75: 5250 [14] Toney, M. D. "Reaction specificity in pyridoxal enzymes." Archives of biochemistry and biophysics (2005) 433: 279-287 [15] Micronutrient Information Center, Oregon State University (http:/ / lpi. oregonstate. edu/ infocenter/ vitamins/ thiamin/ ) [16] Voet, Donald; Judith Voet (2004). Biochemistry. John Wiley & Sons Inc. pp.986989. ISBN0-471-25090-2. [17] Voet, Donald; Judith Voet (2004). Biochemistry. John Wiley & Sons Inc. pp.604606. ISBN0-471-25090-2. [18] Garcia-Viloca, M; Gao, J; Karplus, M; Truhlar, DG (2004). "How enzymes work: analysis by modern rate theory and computer simulations". Science 303 (5655): 18695. doi:10.1126/science.1088172. PMID14716003. [19] Olsson, MH; Siegbahn, PE; Warshel, A (2004). "Simulations of the large kinetic isotope effect and the temperature dependence of the hydrogen atom transfer in lipoxygenase". Journal of the American Chemical Society 126 (9): 28208. doi:10.1021/ja037233l. PMID14995199. [20] Masgrau, L; Roujeinikova, A; Johannissen, LO; Hothi, P; Basran, J; Ranaghan, KE; Mulholland, AJ; Sutcliffe, MJ et al. (2006). "Atomic description of an enzyme reaction dominated by proton tunneling". Science 312 (5771): 23741. doi:10.1126/science.1126002. PMID16614214. [21] Hwang, J.-K.; Warshel, A.. "How important are quantum mechanical nuclear motions in enzyme catalysis". J. Am. Chem. Soc 1996 (118): 1174511751. [22] Ball, P. (2004). "Enzymes: By chance, or by design?". Nature 431 (7007): 396. Bibcode2004Natur.431..396B. doi:10.1038/431396a. [23] Olsson, M.H.M.; Parson, W.W.; Warshel, A.. "Dynamical Contributions to Enzyme Catalysis: Critical Tests of A Popular Hypothesis". Chem. Rev. 2006 (105): 17371756. [24] Volkenshtein M.V., Dogonadze R.R., Madumarov A.K., Urushadze Z.D., Kharkats Yu.I. Theory of Enzyme Catalysis.- Molekuliarnaya Biologia, Moscow, 6, 1972, 431-439 [25] Volkenshtein M.V., Dogonadze R.R., Madumarov A.K., Urushadze Z.D., Kharkats Yu.I. Electronic and Conformational Interactions in Enzyme Catalysis. In: E.L. Andronikashvili (Ed.), Konformatsionnie Izmenenia Biopolimerov v Rastvorakh, Publishing House "Nauka", Moscow, 1973, 153-157 [26] http:/ / enzyme. expasy. org/ EC/ 5. 3. 1. 1 [27] http:/ / enzyme. expasy. org/ EC/ 3. 4. 21. 4 [28] http:/ / enzyme. expasy. org/ EC/ 4. 1. 2. 13

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Further reading
Alan Fersht, Structure and Mechanism in Protein Science : A Guide to Enzyme Catalysis and Protein Folding. W. H. Freeman, 1998. ISBN 0-7167-3268-8 Dedicated issue of Philosophical Transactions B on Quantum catalysis in enzymes freely available. (http:// publishing.royalsociety.org/quantum-catalysis)

148

Enzyme kinetics
Enzyme kinetics
Enzyme kinetics is the study of the chemical reactions that are catalysed by enzymes. In enzyme kinetics, the reaction rate is measured and the effects of varying the conditions of the reaction investigated. Studying an enzyme's kinetics in this way can reveal the catalytic mechanism of this enzyme, its role in metabolism, how its activity is controlled, and how a drug or an agonist might inhibit the enzyme. Enzymes are usually protein molecules that manipulate other molecules the enzymes' substrates. These target molecules bind to an enzyme's active site and are transformed into products through a series of steps known as the enzymatic mechanism. These mechanisms can be divided into single-substrate and multiple-substrate mechanisms. Kinetic studies on enzymes that only bind one substrate, such as triosephosphate isomerase, aim to measure the affinity with which the enzyme binds this substrate and the turnover rate. When enzymes bind multiple substrates, such as (right) and NADPH (left), bound in the active site. The protein is shown as a dihydrofolate reductase (shown right), enzyme ribbon diagram, with alpha helices in red, beta sheets in yellow and loops in [1] blue. Generated from 7DFR . kinetics can also show the sequence in which these substrates bind and the sequence in which products are released. An example of enzymes that bind a single substrate and release multiple products are proteases, which cleave one protein substrate into two polypeptide products. Others join two substrates together, such as DNA polymerase linking a nucleotide to DNA. Although these mechanisms are often a complex series of steps, there is typically one rate-determining step that determines the overall kinetics. This rate-determining step may be a chemical reaction or a conformational change of the enzyme or substrates, such as those involved in the release of product(s) from the enzyme. Knowledge of the enzyme's structure is helpful in interpreting kinetic data. For example, the structure can suggest how substrates and products bind during catalysis; what changes occur during the reaction; and even the role of particular amino acid residues in the mechanism. Some enzymes change shape significantly during the mechanism; in such cases, it is helpful to determine the enzyme structure with and without bound substrate analogues that do not undergo the enzymatic reaction.
Dihydrofolate reductase from E. coli with its two substrates, dihydrofolate

Enzyme kinetics Not all biological catalysts are protein enzymes; RNA-based catalysts such as ribozymes and ribosomes are essential to many cellular functions, such as RNA splicing and translation. The main difference between ribozymes and enzymes is that RNA catalysts are composed of nucleotides, whereas enzymes are composed of amino acids. Ribozymes also perform a more limited set of reactions, although their reaction mechanisms and kinetics can be analysed and classified by the same methods.

149

General principles
The reaction catalysed by an enzyme uses exactly the same reactants and produces exactly the same products as the uncatalysed reaction. Like other catalysts, enzymes do not alter the position of equilibrium between substrates and products.[2] However, unlike uncatalysed chemical reactions, enzyme-catalysed reactions display saturation kinetics. For a given enzyme concentration and for relatively low substrate concentrations, the reaction rate As larger amounts of substrate are added to a reaction, the available enzyme increases linearly with substrate binding sites become filled to the limit of . Beyond this limit the enzyme is concentration; the enzyme molecules are saturated with substrate and the reaction rate ceases to increase. largely free to catalyse the reaction, and increasing substrate concentration means an increasing rate at which the enzyme and substrate molecules encounter one another. However, at relatively high substrate concentrations, the reaction rate asymptotically approaches the theoretical maximum; the enzyme active sites are almost all occupied and the reaction rate is determined by the intrinsic turnover rate of the enzyme. The substrate concentration midway between these two limiting cases is denoted by KM. The two most important kinetic properties of an enzyme are how quickly the enzyme becomes saturated with a particular substrate, and the maximum rate it can achieve. Knowing these properties suggests what an enzyme might do in the cell and can show how the enzyme will respond to changes in these conditions.

Enzyme kinetics

150

Enzyme assays
Enzyme assays are laboratory procedures that measure the rate of enzyme reactions. Because enzymes are not consumed by the reactions they catalyse, enzyme assays usually follow changes in the concentration of either substrates or products to measure the rate of reaction. There are many methods of measurement. Spectrophotometric assays observe change in the absorbance of light between products and reactants; radiometric assays involve the incorporation or release of radioactivity to measure the amount of product made over time. Spectrophotometric assays are most convenient since they allow the rate of the reaction to be measured continuously. Although radiometric assays Progress curve for an enzyme reaction. The slope in the initial rate period is the initial rate of reaction v. The MichaelisMenten require the removal and counting of samples (i.e., they equation describes how this slope varies with the concentration of are discontinuous assays) they are usually extremely substrate. sensitive and can measure very low levels of enzyme activity.[3] An analogous approach is to use mass spectrometry to monitor the incorporation or release of stable isotopes as substrate is converted into product. The most sensitive enzyme assays use lasers focused through a microscope to observe changes in single enzyme molecules as they catalyse their reactions. These measurements either use changes in the fluorescence of cofactors during an enzyme's reaction mechanism, or of fluorescent dyes added onto specific sites of the protein to report movements that occur during catalysis.[4] These studies are providing a new view of the kinetics and dynamics of single enzymes, as opposed to traditional enzyme kinetics, which observes the average behaviour of populations of millions of enzyme molecules.[5] [6] An example progress curve for an enzyme assay is shown above. The enzyme produces product at an initial rate that is approximately linear for a short period after the start of the reaction. As the reaction proceeds and substrate is consumed, the rate continuously slows (so long as substrate is not still at saturating levels). To measure the initial (and maximal) rate, enzyme assays are typically carried out while the reaction has progressed only a few percent towards total completion. The length of the initial rate period depends on the assay conditions and can range from milliseconds to hours. However, equipment for rapidly mixing liquids allows fast kinetic measurements on initial rates of less than one second.[7] These very rapid assays are essential for measuring pre-steady-state kinetics, which are discussed below. Most enzyme kinetics studies concentrate on this initial, approximately linear part of enzyme reactions. However, it is also possible to measure the complete reaction curve and fit this data to a non-linear rate equation. This way of measuring enzyme reactions is called progress-curve analysis.[8] This approach is useful as an alternative to rapid kinetics when the initial rate is too fast to measure accurately.

Single-substrate reactions
Enzymes with single-substrate mechanisms include isomerases such as triosephosphateisomerase or bisphosphoglycerate mutase, intramolecular lyases such as adenylate cyclase and the hammerhead ribozyme, a RNA lyase.[9] However, some enzymes that only have a single substrate do not fall into this category of mechanisms. Catalase is an example of this, as the enzyme reacts with a first molecule of hydrogen peroxide substrate, becomes oxidised and is then reduced by a second molecule of substrate. Although a single substrate is involved, the existence of a modified enzyme intermediate means that the mechanism of catalase is actually a pingpong mechanism, a type

Enzyme kinetics of mechanism that is discussed in the Multi-substrate reactions section below.

151

MichaelisMenten kinetics
As enzyme-catalysed reactions are saturable, their rate of catalysis does not show a linear response to increasing substrate. If the initial rate of the reaction is measured over a range of substrate concentrations (denoted as [S]), the reaction rate (v) increases as [S] increases, as shown on the right. However, as [S] gets higher, the enzyme becomes saturated with substrate and the rate reaches Vmax, the enzyme's maximum rate.

Saturation curve for an enzyme showing the relation between the concentration of substrate and rate.

Single-substrate mechanism for an enzyme reaction. k1, k-1 and k2 are the rate constants for the individual steps.

The MichaelisMenten kinetic model of a single-substrate reaction is shown on the right. There is an initial bimolecular reaction between the enzyme E and substrate S to form the enzymesubstrate complex ES. Although the enzymatic mechanism for the unimolecular reaction can be quite complex, there is typically one rate-determining enzymatic step that allows this reaction to be modelled as a single catalytic step with an apparent unimolecular rate constant kcat. If the reaction path proceeds over one or several intermediates, kcat will be a function of several elementary rate constants, whereas in the simplest case of a single elementary reaction (e.g. no intermediates) it will be identical to the elementary unimolecular rate constant k2. The apparent unimolecular rate constant kcat is also called turnover number and denotes the maximum number of enzymatic reactions catalysed per second. The MichaelisMenten equation[10] describes how the (initial) reaction rate v0 depends on the position of the substrate-binding equilibrium and the rate constant k2. (MichaelisMenten equation) with the constants

Enzyme kinetics

152

This MichaelisMenten equation is the basis for most single-substrate enzyme kinetics. Two crucial assumptions underlie this equation (apart from the general assumption about the mechanism only involving no intermediate or product inhibition, and there is no allostericity or cooperativity). The first assumption is the so called quasi-steady-state assumption (or pseudo-steady-state hypothesis), namely that the concentration of the substrate-bound enzyme (and hence also the unbound enzyme) changes much more slowly than those of the product and substrate and thus the change over time of the complex can be set to zero . The second assumption is that the total enzyme concentration does not . A complete derivation can be found here. change over time, thus

The Michaelis constant KM is experimentally defined as the concentration at which the rate of the enzyme reaction is half Vmax, which can be verified by substituting [S] = Km into the MichaelisMenten equation and can also be seen graphically. If the rate-determining enzymatic step is slow compared to substrate dissociation ( ), the Michaelis constant KM is roughly the dissociation constant KD of the ES complex. If is small compared to then the term and also very little ES complex is formed, thus . Therefore, the rate of product formation is

Thus the product formation rate depends on the enzyme concentration as well as on the substrate concentration, the equation resembles a bimolecular reaction with a corresponding pseudo-second order rate constant . This constant is a measure of catalytic efficiency. The most efficient enzymes reach a in the range of 108 1010M1s1. These enzymes are so efficient they effectively catalyse a reaction each time they encounter a substrate molecule and have thus reached an upper theoretical limit for efficiency (diffusion limit); these enzymes have often been termed perfect enzymes.[11]

Direct use of the MichaelisMenten equation for time course kinetic analysis
Further information: Rate equation The observed velocities predicted by the MichaelisMenten equation can be used to directly model the time course disappearance of substrate and the production of product through incorporation of the MichaelisMenten equation into the equation for first order chemical kinetics. This can only be achieved however if one recognises the problem associated with the use of Euler's number in the description of first order chemical kinetics. i.e. e-k is a split constant that introduces a systematic error into calculations and can be rewritten as a single constant which represents the remaining substrate after each time period.[12]

In 1997, Santiago Schnell and Claudio Mendoza derived a closed form solution for the time course kinetics analysis of the Michaelis-Menten mechanism.[13] The solution has the form: where W[] is the Lambert-W function.[14] [15]

Enzyme kinetics

153

Linear plots of the MichaelisMenten equation

Using an interactive MichaelisMenten kinetics tutorial at the University of Virginia,[] the effects on the behaviour of an enzyme of varying kinetic constants can be explored. The plot of v versus [S] above is not linear; although initially linear at low [S], it bends over to saturate at high [S]. Before the modern era of nonlinear curve-fitting on computers, this nonlinearity could make it difficult to estimate KM and Vmax accurately. LineweaverBurk or double-reciprocal plot of kinetic data, showing the significance of Therefore, several researchers the axis intercepts and gradient. developed linearisations of the MichaelisMenten equation, such as the LineweaverBurk plot, the EadieHofstee diagram and the HanesWoolf plot. All of these linear representations can be useful for visualising data, but none should be used to determine kinetic parameters, as computer software is readily available that allows for more accurate determination by nonlinear regression methods.[16] The LineweaverBurk plot or double reciprocal plot is a common way of illustrating kinetic data. This is produced by taking the reciprocal of both sides of the MichaelisMenten equation. As shown on the right, this is a linear form of the MichaelisMenten equation and produces a straight line with the equation y = mx + c with a y-intercept equivalent to 1/Vmax and an x-intercept of the graph representing -1/KM.

Naturally, no experimental values can be taken at negative 1/[S]; the lower limiting value 1/[S] = 0 (the y-intercept) corresponds to an infinite substrate concentration, where 1/v=1/Vmax as shown at the right; thus, the x-intercept is an extrapolation of the experimental data taken at positive concentrations. More generally, the LineweaverBurk plot skews the importance of measurements taken at low substrate concentrations and, thus, can yield inaccurate estimates of Vmax and KM.[17] A more accurate linear plotting method is the Eadie-Hofstee plot. In this case, v is plotted against v/[S]. In the third common linear representation, the Hanes-Woolf plot, [S]/v is plotted against [S]. In general, data normalisation can help diminish the amount of experimental work and can increase the reliability of the output, and is suitable for both graphical and numerical analysis.[18]

Practical significance of kinetic constants

The study of enzyme kinetics is important for two basic reasons. Firstly, it helps explain how enzymes work, and secondly, it helps predict how enzymes behave in living organisms. The kinetic constants defined above, KM and Vmax, are critical to attempts to understand how enzymes work together to control metabolism. Making these predictions is not trivial, even for simple systems. For example, oxaloacetate is formed by malate dehydrogenase within the mitochondrion. Oxaloacetate can then be consumed by citrate synthase, phosphoenolpyruvate carboxykinase or aspartate aminotransferase, feeding into the citric acid cycle, gluconeogenesis or aspartic acid biosynthesis, respectively. Being able to predict how much oxaloacetate goes into which pathway requires knowledge of the concentration of oxaloacetate as well as the concentration and kinetics of each of these enzymes. This aim of predicting the behaviour of metabolic pathways reaches its most complex

Enzyme kinetics expression in the synthesis of huge amounts of kinetic and gene expression data into mathematical models of entire organisms. Alternatively, one useful simplification of the metabolic modelling problem is to ignore the underlying enzyme kinetics and only rely on information about the reaction network's stoichiometry, a technique called flux balance analysis.[19] [20]

154

MichaelisMenten kinetics with intermediate

One could also consider the less simple case

where a complex with the enzyme and an intermediate exists and the intermediate is converted into product in a second step. In this case we have a very similar equation[21]

but the constants are different

We see that for the limiting case

, thus when the last step from EI to E + P is much faster than the and .

previous step, we get again the original equation. Mathematically we have then

Multi-substrate reactions
Multi-substrate reactions follow complex rate equations that describe how the substrates bind and in what sequence. The analysis of these reactions is much simpler if the concentration of substrate A is kept constant and substrate B varied. Under these conditions, the enzyme behaves just like a single-substrate enzyme and a plot of v by [S] gives apparent KM and Vmax constants for substrate B. If a set of these measurements is performed at different fixed concentrations of A, these data can be used to work out what the mechanism of the reaction is. For an enzyme that takes two substrates A and B and turns them into two products P and Q, there are two types of mechanism: ternary complex and pingpong.

Ternary-complex mechanisms
In these enzymes, both substrates bind to the enzyme at the same time to produce an EAB ternary complex. The order of binding can either be random (in a random mechanism) or substrates have to bind in a particular sequence (in an ordered mechanism). When a set of v by [S] curves (fixed A, varying B) from an enzyme with a ternary-complex mechanism are plotted in a LineweaverBurk plot, the set of lines produced will intersect.

Random-order ternary-complex mechanism for an enzyme reaction. The reaction path is shown as a line and enzyme intermediates containing substrates A and B or products P and Q are written below the line.

Enzyme kinetics Enzymes with ternary-complex mechanisms include glutathione S-transferase,[22] dihydrofolate reductase[23] and DNA polymerase.[24] The following links show short animations of the ternary-complex mechanisms of the enzymes dihydrofolate reductase[] and DNA polymerase[].

155

Pingpong mechanisms
As shown on the right, enzymes with a ping-pong mechanism can exist in two states, E and a chemically modified form of the enzyme E*; this modified enzyme Pingpong mechanism for an enzyme reaction. Intermediates contain substrates A is known as an intermediate. In such and B or products P and Q. mechanisms, substrate A binds, changes the enzyme to E* by, for example, transferring a chemical group to the active site, and is then released. Only after the first substrate is released can substrate B bind and react with the modified enzyme, regenerating the unmodified E form. When a set of v by [S] curves (fixed A, varying B) from an enzyme with a pingpong mechanism are plotted in a LineweaverBurk plot, a set of parallel lines will be produced. This is called a secondary plot. Enzymes with pingpong mechanisms include some oxidoreductases such as thioredoxin peroxidase,[25] transferases such as acylneuraminate cytydilyltransferase[26] and serine proteases such as trypsin and chymotrypsin.[27] Serine proteases are a very common and diverse family of enzymes, including digestive enzymes (trypsin, chymotrypsin, and elastase), several enzymes of the blood clotting cascade and many others. In these serine proteases, the E* intermediate is an acyl-enzyme species formed by the attack of an active site serine residue on a peptide bond in a protein substrate. A short animation showing the mechanism of chymotrypsin is linked here.[]

Non-MichaelisMenten kinetics
Some enzymes produce a sigmoid v by [S] plot, which often indicates cooperative binding of substrate to the active site. This means that the binding of one substrate molecule affects the binding of subsequent substrate molecules. This behavior is most common in multimeric enzymes with several interacting active sites.[28] Here, the mechanism of cooperation is similar to that of hemoglobin, with binding of substrate to one active site altering the affinity of the other active sites for substrate molecules. Positive cooperativity occurs when binding of the first substrate molecule increases the affinity of the other active sites for substrate. Negative cooperativity occurs when binding of the first substrate decreases the affinity of

Saturation curve for an enzyme reaction showing sigmoid kinetics.

the enzyme for other substrate molecules. Allosteric enzymes include mammalian tyrosyl tRNA-synthetase, which shows negative cooperativity,[29] and bacterial aspartate transcarbamoylase[30] and phosphofructokinase,[31] which show positive cooperativity. Cooperativity is surprisingly common and can help regulate the responses of enzymes to changes in the concentrations of their substrates. Positive cooperativity makes enzymes much more sensitive to [S] and their

Enzyme kinetics activities can show large changes over a narrow range of substrate concentration. Conversely, negative cooperativity makes enzymes insensitive to small changes in [S]. The Hill equation (biochemistry)[32] is often used to describe the degree of cooperativity quantitatively in non-MichaelisMenten kinetics. The derived Hill coefficient n measures how much the binding of substrate to one active site affects the binding of substrate to the other active sites. A Hill coefficient of <1 indicates negative cooperativity and a coefficient of >1 indicates positive cooperativity.

156

Pre-steady-state kinetics
In the first moment after an enzyme is mixed with substrate, no product has been formed and no intermediates exist. The study of the next few milliseconds of the reaction is called Pre-steady-state kinetics also referred to as Burst kinetics. Pre-steady-state kinetics is therefore concerned with the formation and consumption of enzymesubstrate intermediates (such as ES or E*) until their steady-state concentrations are reached. This approach was first applied to the hydrolysis reaction catalysed by Pre-steady state progress curve, showing the burst phase of an enzyme reaction. [33] chymotrypsin. Often, the detection of an intermediate is a vital piece of evidence in investigations of what mechanism an enzyme follows. For example, in the pingpong mechanisms that are shown above, rapid kinetic measurements can follow the release of product P and measure the formation of the modified enzyme intermediate E*.[34] In the case of chymotrypsin, this intermediate is formed by an attack on the substrate by the nucleophilic serine in the active site and the formation of the acyl-enzyme intermediate. In the figure to the right, the enzyme produces E* rapidly in the first few seconds of the reaction. The rate then slows as steady state is reached. This rapid burst phase of the reaction measures a single turnover of the enzyme. Consequently, the amount of product released in this burst, shown as the intercept on the y-axis of the graph, also gives the amount of functional enzyme which is present in the assay.[35]

Chemical mechanism
An important goal of measuring enzyme kinetics is to determine the chemical mechanism of an enzyme reaction, i.e., the sequence of chemical steps that transform substrate into product. The kinetic approaches discussed above will show at what rates intermediates are formed and inter-converted, but they cannot identify exactly what these intermediates are. Kinetic measurements taken under various solution conditions or on slightly modified enzymes or substrates often shed light on this chemical mechanism, as they reveal the rate-determining step or intermediates in the reaction. For example, the breaking of a covalent bond to a hydrogen atom is a common rate-determining step. Which of the possible hydrogen transfers is rate determining can be shown by measuring the kinetic effects of substituting each hydrogen by deuterium, its stable isotope. The rate will change when the critical hydrogen is replaced, due to a primary kinetic isotope effect, which occurs because bonds to deuterium are harder to break than bonds to hydrogen.[36] It is also possible to measure similar effects with other isotope substitutions, such as 13C/12C and 18 16 O/ O, but these effects are more subtle.[37]

Enzyme kinetics Isotopes can also be used to reveal the fate of various parts of the substrate molecules in the final products. For example, it is sometimes difficult to discern the origin of an oxygen atom in the final product; since it may have come from water or from part of the substrate. This may be determined by systematically substituting oxygen's stable isotope 18O into the various molecules that participate in the reaction and checking for the isotope in the product.[38] The chemical mechanism can also be elucidated by examining the kinetics and isotope effects under different pH conditions,[39] by altering the metal ions or other bound cofactors,[40] by site-directed mutagenesis of conserved amino acid residues, or by studying the behaviour of the enzyme in the presence of analogues of the substrate(s).[41]

157

Enzyme inhibition and activation

Enzyme inhibitors are molecules that reduce or abolish enzyme activity, while enzyme activators are molecules that increase the catalytic rate of enzymes. These interactions can be either reversible (i.e., removal of the inhibitor restores enzyme activity) or irreversible (i.e., the inhibitor permanently inactivates the enzyme).

Reversible inhibitors

Kinetic scheme for reversible enzyme inhibitors.

Traditionally reversible enzyme inhibitors have been classified as competitive, uncompetitive, or non-competitive, according to their effects on Km and Vmax. These different effects result from the inhibitor binding to the enzyme E, to the enzymesubstrate complex ES, or to both, respectively. The particular type of an inhibitor can be discerned by studying the enzyme kinetics as a function of the inhibitor concentration. The three types of inhibition produce LineweaverBurke and EadieHofstee plots[17] that vary in distinctive ways with inhibitor concentration. For a straightforward qualitative explanation, see[42] Non-linear regression fits of the enzyme kinetics data to rate equations[43] can yield accurate estimates of dissociation constants and yield important information relating to the mechanism of action.

Adding zero to the bottom ([I]-[I])

Dividing by [I]+Ki

Enzyme kinetics

158

This notation demonstrates that similar to the MichaelisMenten equation,where the rate of reaction depends on the percent of the enzyme population interacting with substrate fraction of the enzyme population bound by substrate

fraction of the enzyme population bound by inhibitor

the effect of the inhibitor is a result of the percent of the enzyme population interacting with inhibitor. The only problem with this equation in its present form is that it assumes absolute inhibition of the enzyme with inhibitor binding, when in fact there can be a wide range of affects anywhere from 100% inhibition of substrate turn over to just >0%. To account for this the equation can be easily modified to allow for different degrees of inhibition by including a delta Vmax term.

or

This term can then define the residual enzymatic activity present when the inhibitor is interacting with individual enzymes in the population. However the inclusion of this term has the added value of allowing for the possibility of activation if the secondary Vmax term turns out to be higher than the initial term. To account for the possibly of activation as well the notation can then be rewritten replacing the inhibitor "I" with a modifier term denoted here as "X".

While this terminology results in a simplified way of dealing with kinetic effects relating to the maximum velocity of the MichaelisMenten equation, it highlights potential problems with the term used to describe effects relating to the Km. The Km relating to the affinity of the enzyme for the substrate should in most cases relate to potential changes in the binding site of the enzyme which would directly result from enzyme inhibitor interactions. As such a term similar to the one proposed above to modulate Vmax should be appropriate in most situations.:[44]

Enzyme kinetics

159

Irreversible inhibitors
Enzyme inhibitors can also irreversibly inactivate enzymes, usually by covalently modifying active site residues. These reactions, which may be called suicide substrates, follow exponential decay functions and are usually saturable. Below saturation, they follow first order kinetics with respect to inhibitor.

Mechanisms of catalysis
The favoured model for the enzymesubstrate interaction is the induced fit model.[45] This model proposes that the initial interaction between enzyme and substrate is relatively weak, but that these weak interactions rapidly induce conformational changes in the enzyme that strengthen binding. These conformational changes also bring catalytic residues in the active site close to the chemical bonds in the substrate that will be altered in the reaction.[46] Conformational changes can be measured using circular dichroism or dual polarisation interferometry. After binding takes place, one or more mechanisms of The energy variation as a function of reaction coordinate shows the stabilisation of catalysis lower the energy of the reaction's the transition state by an enzyme. transition state by providing an alternative chemical pathway for the reaction. Mechanisms of catalysis include catalysis by bond strain; by proximity and orientation; by active-site proton donors or acceptors; covalent catalysis and quantum tunnelling.[34] [47] Enzyme kinetics cannot prove which modes of catalysis are used by an enzyme. However, some kinetic data can suggest possibilities to be examined by other techniques. For example, a pingpong mechanism with burst-phase pre-steady-state kinetics would suggest covalent catalysis might be important in this enzyme's mechanism. Alternatively, the observation of a strong pH effect on Vmax but not Km might indicate that a residue in the active site needs to be in a particular ionisation state for catalysis to occur.

Footnotes
. Link: Interactive MichaelisMenten kinetics tutorial (Java required) [48] . Link: dihydrofolate reductase mechanism (Gif) [49] . Link: DNA polymerase mechanism (Gif) [50] . Link: Chymotrypsin mechanism (Flash required) [51]

References
[1] http:/ / www. rcsb. org/ pdb/ explore. do?structureId=7DFR [2] Wrighton, Mark S.; Ebbing, Darrell D. (1993). General chemistry (4th ed.). Boston: Houghton Mifflin. ISBN0-395-63696-5. [3] Danson, Michael; Eisenthal, Robert (2002). Enzyme assays: a practical approach. Oxford [Oxfordshire]: Oxford University Press. ISBN0-19-963820-9. [4] Xie XS, Lu HP (June 1999). "Single-molecule enzymology" (http:/ / www. jbc. org/ cgi/ content/ full/ 274/ 23/ 15967). J. Biol. Chem. 274 (23): 1596770. doi:10.1074/jbc.274.23.15967. PMID10347141. .

Enzyme kinetics
[5] Lu H (2004). "Single-molecule spectroscopy studies of conformational change dynamics in enzymatic reactions". Current pharmaceutical biotechnology 5 (3): 2619. doi:10.2174/1389201043376887. PMID15180547. [6] Schnell J, Dyson H, Wright P (2004). "Structure, dynamics, and catalytic function of dihydrofolate reductase". Annual review of biophysics and biomolecular structure 33: 11940. doi:10.1146/annurev.biophys.33.110502.133613. PMID15139807. [7] Gibson QH (1969). "Rapid mixing: Stopped flow". Methods Enzymol. Methods in Enzymology 16: 187228. doi:10.1016/S0076-6879(69)16009-7. ISBN9780121818739. [8] Duggleby RG (1995). "Analysis of enzyme progress curves by non-linear regression". Methods Enzymol. Methods in Enzymology 249: 6190. doi:10.1016/0076-6879(95)49031-0. ISBN9780121821500. PMID7791628. [9] Murray JB, Dunham CM, Scott WG (January 2002). "A pH-dependent conformational change, rather than the chemical step, appears to be rate-limiting in the hammerhead ribozyme cleavage reaction". J. Mol. Biol. 315 (2): 12130. doi:10.1006/jmbi.2001.5145. PMID11779233. [10] Michaelis L. and Menten M.L. Kinetik der Invertinwirkung Biochem. Z. 1913; 49:333369 English translation (http:/ / web. lemoyne. edu/ ~giunta/ menten. html) Accessed 6 April 2007 [11] Stroppolo ME, Falconi M, Caccuri AM, Desideri A (2001). "Superefficient enzymes". Cell. Mol. Life Sci. 58 (10): 145160. doi:10.1007/PL00000788. PMID11693526. [12] Walsh R, Martin E, Darvesh S. A method to describe enzyme-catalyzed reactions by combining steady state and time course enzyme kinetic parameters. Biochim Biophys Acta. 2010 Jan;1800:1-5 [13] Schnell S, Mendoza C. A closed form solution for time-dependent enzyme kinetics. Journal of theoretical Biology, 187 (1997): 207-212 [14] C.T. Goudar, J.R. Sonnad, R.G. Duggleby (1999). Parameter estimation using a direct solution of the integrated Michaelis-Menten equation. Biochimica et Biophysica Acta - Protein Structure and Molecular Enzymology 1429, 377-383. [15] C.T. Goudar, S.K. Harris, M.J. McInerney, J.M Suflita (2004). Progress curve analysis for enzyme and microbial kinetic reactions using explicit solutions based on the Lambert W function. Journal of Microbiological Methods 59, 317-326 [16] Jones ME (1 January 1992). "Analysis of algebraic weighted least-squares estimators for enzyme parameters". Biochem. J. 288 (Pt 2): 5338. PMC1132043. PMID1463456. [17] Tseng SJ, Hsu JP (August 1990). "A comparison of the parameter estimating procedures for the Michaelis-Menten model". J. Theor. Biol. 145 (4): 45764. doi:10.1016/S0022-5193(05)80481-3. PMID2246896. [18] Bravo IG, Busto F, De Arriaga D, et al. (September 2001). "A normalized plot as a novel and time-saving tool in complex enzyme kinetic analysis" (http:/ / www. biochemj. org/ bj/ 358/ 0573/ bj3580573. htm). Biochem. J. 358 (Pt 3): 57383. PMC1222113. PMID11577687. . [19] Almaas E, Kovcs B, Vicsek T, Oltvai ZN, Barabsi AL (February 2004). "Global organization of metabolic fluxes in the bacterium Escherichia coli". Nature 427 (6977): 83943. doi:10.1038/nature02289. PMID14985762. [20] Reed JL, Vo TD, Schilling CH, Palsson BO (2003). "An expanded genome-scale model of Escherichia coli K-12 (iJR904 GSM/GPR)". Genome Biol. 4 (9): R54. doi:10.1186/gb-2003-4-9-r54. PMC193654. PMID12952533. [21] for a complete derivation, see here [22] Dirr H, Reinemer P, Huber R (March 1994). "X-ray crystal structures of cytosolic glutathione S-transferases. Implications for protein architecture, substrate recognition and catalytic function". Eur. J. Biochem. 220 (3): 64561. doi:10.1111/j.1432-1033.1994.tb18666.x. PMID8143720. [23] Stone SR, Morrison JF (July 1988). "Dihydrofolate reductase from Escherichia coli: the kinetic mechanism with NADPH and reduced acetylpyridine adenine dinucleotide phosphate as substrates". Biochemistry 27 (15): 54939. doi:10.1021/bi00415a016. PMID3052577. [24] Fisher PA (1994). "Enzymologic mechanism of replicative DNA polymerases in higher eukaryotes". Prog. Nucleic Acid Res. Mol. Biol.. Progress in Nucleic Acid Research and Molecular Biology 47: 37197. doi:10.1016/S0079-6603(08)60257-3. ISBN9780125400473. PMID8016325. [25] Akerman SE, Mller S (August 2003). "2-Cys peroxiredoxin PfTrx-Px1 is involved in the antioxidant defence of Plasmodium falciparum" (http:/ / www. sciencedirect. com/ science?_ob=ArticleURL& _udi=B6T29-49321S6-2& _coverDate=08/ 31/ 2003& _alid=466052639& _rdoc=1& _fmt=& _orig=search& _qd=1& _cdi=4913& _sort=d& view=c& _acct=C000050221& _version=1& _urlVersion=0& _userid=10& md5=965e18a4c2ad0af711ba05d1a46dc855). Mol. Biochem. Parasitol. 130 (2): 7581. doi:10.1016/S0166-6851(03)00161-0. PMID12946843. . [26] Bravo IG, Barrallo S, Ferrero MA, Rodrguez-Aparicio LB, Martnez-Blanco H, Reglero A (September 2001). "Kinetic properties of the acylneuraminate cytidylyltransferase from Pasteurella haemolytica A2" (http:/ / www. biochemj. org/ bj/ 358/ 0585/ bj3580585. htm). Biochem. J. 358 (Pt 3): 58598. PMC1222114. PMID11577688. . [27] Kraut J (1977). "Serine proteases: structure and mechanism of catalysis" (http:/ / arjournals. annualreviews. org/ doi/ abs/ 10. 1146/ annurev. bi. 46. 070177. 001555?url_ver=Z39. 88-2003& rfr_id=ori:rid:crossref. org& rfr_dat=cr_pub=ncbi. nlm. nih. gov). Annu. Rev. Biochem. 46: 33158. doi:10.1146/annurev.bi.46.070177.001555. PMID332063. . [28] Ricard J, Cornish-Bowden A (July 1987). "Co-operative and allosteric enzymes: 20 years on". Eur. J. Biochem. 166 (2): 25572. doi:10.1111/j.1432-1033.1987.tb13510.x. PMID3301336. [29] Ward WH, Fersht AR (July 1988). "Tyrosyl-tRNA synthetase acts as an asymmetric dimer in charging tRNA. A rationale for half-of-the-sites activity". Biochemistry 27 (15): 552530. doi:10.1021/bi00415a021. PMID3179266. [30] Helmstaedt K, Krappmann S, Braus GH (September 2001). "Allosteric Regulation of Catalytic Activity: Escherichia coli Aspartate Transcarbamoylase versus Yeast Chorismate Mutase" (http:/ / mmbr. asm. org/ cgi/ content/ full/ 65/ 3/ 404). Microbiol. Mol. Biol. Rev. 65 (3): 40421, table of contents. doi:10.1128/MMBR.65.3.404-421.2001. PMC99034. PMID11528003. .

160

Enzyme kinetics
[31] Schirmer T, Evans PR (January 1990). "Structural basis of the allosteric behaviour of phosphofructokinase". Nature 343 (6254): 1405. doi:10.1038/343140a0. PMID2136935. [32] Hill, A. V. The possible effects of the aggregation of the molecules of haemoglobin on its dissociation curves. J. Physiol. (Lond.), 1910 40, iv-vii. [33] Hartley BS, Kilby BA (February 1954). "The reaction of p-nitrophenyl esters with chymotrypsin and insulin". Biochem. J. 56 (2): 28897. PMC1269615. PMID13140189. [34] Fersht, Alan (1999). Structure and mechanism in protein science: a guide to enzyme catalysis and protein folding. San Francisco: W.H. Freeman. ISBN0-7167-3268-8. [35] Bender ML, Begu-Cantn ML, Blakeley RL, et al. (December 1966). "The determination of the concentration of hydrolytic enzyme solutions: alpha-chymotrypsin, trypsin, papain, elastase, subtilisin, and acetylcholinesterase". J. Am. Chem. Soc. 88 (24): 5890913. doi:10.1021/ja00976a034. PMID5980876. [36] Cleland WW (January 2005). "The use of isotope effects to determine enzyme mechanisms" (http:/ / www. sciencedirect. com/ science?_ob=ArticleURL& _udi=B6WB5-4DD8DXJ-8& _coverDate=01/ 01/ 2005& _alid=466049795& _rdoc=1& _fmt=& _orig=search& _qd=1& _cdi=6701& _sort=d& view=c& _acct=C000050221& _version=1& _urlVersion=0& _userid=10& md5=cf322c4a1c7db6b9f89551a8469a1a2d). Arch. Biochem. Biophys. 433 (1): 212. doi:10.1016/j.abb.2004.08.027. PMID15581561. . [37] Northrop D (1981). "The expression of isotope effects on enzyme-catalyzed reactions". Annu. Rev. Biochem. 50: 10331. doi:10.1146/annurev.bi.50.070181.000535. PMID7023356. [38] Baillie T, Rettenmeier A (1986). "Drug biotransformation: mechanistic studies with stable isotopes". Journal of clinical pharmacology 26 (6): 44851. PMID3734135. [39] Cleland WW (1982). "Use of isotope effects to elucidate enzyme mechanisms". CRC Crit. Rev. Biochem. 13 (4): 385428. doi:10.3109/10409238209108715. PMID6759038. [40] Christianson DW, Cox JD (1999). "Catalysis by metal-activated hydroxide in zinc and manganese metalloenzymes". Annu. Rev. Biochem. 68: 3357. doi:10.1146/annurev.biochem.68.1.33. PMID10872443. [41] Kraut D, Carroll K, Herschlag D (2003). "Challenges in enzyme mechanism and energetics". Annu. Rev. Biochem. 72: 51771. doi:10.1146/annurev.biochem.72.121801.161617. PMID12704087. [42] Waldrop GL (January 2009). "A qualitative approach to enzyme inhibition". Biochemistry and Molecular Biology Education 37 (1): 1115. doi:10.1002/bmb.20243. PMID21567682. [43] Leatherbarrow RJ (December 1990). "Using linear and non-linear regression to fit biochemical data". Trends Biochem. Sci. 15 (12): 4558. doi:10.1016/0968-0004(90)90295-M. PMID2077683. [44] Walsh R, Martin E, Darvesh S. A versatile equation to describe reversible enzyme inhibition and activation kinetics: modeling beta-galactosidase and butyrylcholinesterase. Biochim Biophys Acta. 2007 1770:733-46. [45] Koshland DE (February 1958). "Application of a Theory of Enzyme Specificity to Protein Synthesis". Proc. Natl. Acad. Sci. U.S.A. 44 (2): 98104. doi:10.1073/pnas.44.2.98. PMC335371. PMID16590179. [46] Hammes G (2002). "Multiple conformational changes in enzyme catalysis". Biochemistry 41 (26): 82218. doi:10.1021/bi0260839. PMID12081470. [47] Sutcliffe M, Scrutton N (2002). "A new conceptual framework for enzyme catalysis. Hydrogen tunnelling coupled to enzyme dynamics in flavoprotein and quinoprotein enzymes" (http:/ / content. febsjournal. org/ cgi/ content/ full/ 269/ 13/ 3096). Eur. J. Biochem. 269 (13): 3096102. doi:10.1046/j.1432-1033.2002.03020.x. PMID12084049. . [48] http:/ / cti. itc. virginia. edu/ ~cmg/ Demo/ scriptFrame. html [49] http:/ / chem-faculty. ucsd. edu/ kraut/ dhfr. html [50] http:/ / chem-faculty. ucsd. edu/ kraut/ dNTP. html [51] http:/ / courses. cm. utexas. edu/ jrobertus/ ch339k/ overheads-2/ 06_21_chymotrypsin. html

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Further reading
Introductory Cornish-Bowden, Athel (2004). Fundamentals of enzyme kinetics (3rd ed.). London: Portland Press. ISBN1-85578-158-1. Stevens, Lewis; Price, Nicholas C. (1999). Fundamentals of enzymology: the cell and molecular biology of catalytic proteins. Oxford [Oxfordshire]: Oxford University Press. ISBN0-19-850229-X. Bugg, Tim (2004). Introduction to Enzyme and Coenzyme Chemistry. Cambridge, MA: Blackwell Publishers. ISBN1-4051-1452-5. Advanced Segel, Irwin H. (1993). Enzyme kinetics: behavior and analysis of rapid equilibrium and steady state enzyme systems (New ed.). New York: Wiley. ISBN0-471-30309-7.

Enzyme kinetics Fersht, Alan (1999). Structure and mechanism in protein science: a guide to enzyme catalysis and protein folding. San Francisco: W.H. Freeman. ISBN0-7167-3268-8. Santiago Schnell, Philip K. Maini (2004). "A century of enzyme kinetics: Reliability of the KM and vmax estimates" (http://www.informatics.indiana.edu/schnell/papers/ctb8_169.pdf). Comments on Theoretical Biology 8 (23): 16987. doi:10.1080/08948550302453. Walsh, Christopher (1979). Enzymatic reaction mechanisms. San Francisco: W. H. Freeman. ISBN0-7167-0070-0. Cleland, William Wallace; Cook, Paul (2007). Enzyme kinetics and mechanism. New York: Garland Science. ISBN0-8153-4140-7.

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External links
Animation of an enzyme assay (http://www.kscience.co.uk/animations/model.swf) Shows effects of manipulating assay conditions MACiE (http://www.ebi.ac.uk/thornton-srv/databases/MACiE/) A database of enzyme reaction mechanisms ENZYME (http://us.expasy.org/enzyme/) Expasy enzyme nomenclature database ENZO (http://enzo.cmm.ki.si) Web application for easy construction and quick testing of kinetic models of enzyme catalyzed reactions. ExCatDB (http://mbs.cbrc.jp/EzCatDB/) A database of enzyme catalytic mechanisms BRENDA (http://www.brenda-enzymes.info/) Comprehensive enzyme database, giving substrates, inhibitors and reaction diagrams SABIO-RK (http://sabio.villa-bosch.de/SABIORK/) A database of reaction kinetics Joseph Kraut's Research Group, University of California San Diego (http://chem-faculty.ucsd.edu/kraut/dhfr. html) Animations of several enzyme reaction mechanisms Symbolism and Terminology in Enzyme Kinetics (http://www.chem.qmul.ac.uk/iubmb/kinetics/) A comprehensive explanation of concepts and terminology in enzyme kinetics An introduction to enzyme kinetics (http://orion1.paisley.ac.uk/kinetics/contents.html) An accessible set of on-line tutorials on enzyme kinetics Enzyme kinetics animated tutorial (http://www.wiley.com/college/pratt/0471393878/student/animations/ enzyme_kinetics/index.html) An animated tutorial with audio

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Lipids and membranes

Lipid
Lipids constitute a broad group of naturally occurring molecules that include fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E, and K), monoglycerides, diglycerides, triglycerides, phospholipids, and others. The main biological functions of lipids include energy storage, as structural components of cell membranes, and as important signaling molecules.[4] Lipids may be broadly defined as hydrophobic or amphiphilic small molecules; the amphiphilic nature of some lipids allows them to form structures such as vesicles, liposomes, or membranes in an aqueous environment. Biological lipids originate entirely or in part from two distinct types of biochemical subunits [1] [2] Structures of some common lipids. At the top are oleic acid and cholesterol. The or "building-blocks": ketoacyl and middle structure is a triglyceride composed of oleoyl, stearoyl, and palmitoyl chains attached to a glycerol backbone. At the bottom is the common phospholipid, isoprene groups.[4] Using this [3] phosphatidylcholine. approach, lipids may be divided into eight categories: fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, and polyketides (derived from condensation of ketoacyl subunits); and sterol lipids and prenol lipids (derived from condensation of isoprene subunits).[4] Although the term lipid is sometimes used as a synonym for fats, fats are a subgroup of lipids called triglycerides. Lipids also encompass molecules such as fatty acids and their derivatives (including tri-, di-, monoglycerides, and phospholipids), as well as other sterol-containing metabolites such as cholesterol.[5] Although humans and other mammals use various biosynthetic pathways to both break down and synthesize lipids, some essential lipids cannot be made this way and must be obtained from the diet.

Categories of lipids

Lipid

164

Fatty acyls
Fatty acyls, a generic term for describing fatty acids, their conjugates, and derivatives, are a diverse group of molecules synthesized by chain-elongation of an acetyl-CoA primer with malonyl-CoA or methylmalonyl-CoA groups in a process called fatty acid synthesis.[6] [7] They are made of a hydrocarbon chain that terminates with a carboxylic acid group; this arrangement confers the molecule with a polar, hydrophilic end, and a nonpolar, hydrophobic end that is insoluble in water. The fatty acid structure is one of the most fundamental categories of biological lipids, and is commonly used as a building-block of more structurally complex lipids. The carbon chain, typically between four and 24 carbons long,[8] may be saturated or unsaturated, and may be attached to functional groups containing oxygen, halogens, nitrogen, and sulfur. Where a double bond exists, there is the possibility of either a cis or a trans geometric isomerism, which significantly affects the molecule's molecular configuration. Cis-double bonds cause the fatty acid chain to bend, an effect that is more pronounced the more double bonds there are in a chain. This in turn plays an important role in the structure and function of cell membranes.[9] Most naturally occurring fatty acids are of the cis configuration, although the trans form does exist in some natural and partially hydrogenated fats and oils.[10] Examples of biologically important fatty acids are the eicosanoids, derived primarily from arachidonic acid and eicosapentaenoic acid, which include prostaglandins, leukotrienes, and thromboxanes. Other major lipid classes in the fatty acid category are the fatty esters and fatty amides. Fatty esters include important biochemical intermediates such as wax esters, fatty acid thioester coenzyme A derivatives, fatty acid thioester ACP derivatives and fatty acid carnitines. The fatty amides include N-acyl ethanolamines, such as the cannabinoid neurotransmitter anandamide.[11]

Glycerolipids
Glycerolipids are composed mainly of mono-, di-, and tri-substituted glycerols,[12] the most well-known being the fatty acid triesters of glycerol (triacylglycerols), also known as triglycerides. In these compounds, the three hydroxyl groups of glycerol are each esterified, usually by different fatty acids. Because they function as a food store, these lipids comprise the bulk of storage fat in animal tissues. The hydrolysis of the ester bonds of triacylglycerols and the release of glycerol and fatty acids from adipose tissue is called fat mobilization.[13] Additional subclasses of glycerolipids are represented by glycosylglycerols, which are characterized by the presence of one or more sugar residues attached to glycerol via a glycosidic linkage. Examples of structures in this category are the digalactosyldiacylglycerols found in plant membranes[14] and seminolipid from mammalian sperm cells.[15]

Glycerophospholipids
Glycerophospholipids, also referred to as phospholipids, are ubiquitous in nature and are key components of the lipid bilayer of cells, as well as being involved in metabolism and cell signaling. Neural tissue (including the brain) contains relatively high amounts of glycerophospholipids, and alterations in their composition has been implicated in various neurological disorders.[16] Glycerophospholipids may be subdivided into distinct classes, based on the nature of the polar headgroup at the sn-3 position of the glycerol backbone in eukaryotes and eubacteria, or the sn-1 position in the case of archaebacteria.[17] Examples of glycerophospholipids found in biological membranes are phosphatidylcholine (also known as PC, GPCho or lecithin), phosphatidylethanolamine (PE or GPEtn) and phosphatidylserine (PS or GPSer). In addition to serving as a primary component of cellular membranes and binding sites for intra- and intercellular proteins, some glycerophospholipids in eukaryotic cells, such as phosphatidylinositols and phosphatidic acids are either precursors of or, themselves,

Phosphatidylethanolamine

Lipid membrane-derived second messengers.[18] Typically, one or both of these hydroxyl groups are acylated with long-chain fatty acids, but there are also alkyl-linked and 1Z-alkenyl-linked (plasmalogen) glycerophospholipids, as well as dialkylether variants in archaebacteria.[19]

165

Sphingolipids
Sphingolipids are a complex family of compounds[20] that share a common structural feature, a sphingoid base backbone that is synthesized de novo from the amino acid serine and a long-chain fatty acyl CoA, then converted into ceramides, phosphosphingolipids, glycosphingolipids and other compounds. The major sphingoid base of mammals is commonly referred to as sphingosine. Ceramides (N-acyl-sphingoid bases) are a major subclass of sphingoid base derivatives with an amide-linked fatty acid. The fatty acids are typically saturated or mono-unsaturated with chain lengths from 16 to 26 carbon atoms.[21] The major phosphosphingolipids of mammals are sphingomyelins (ceramide phosphocholines),[22] whereas insects contain mainly ceramide phosphoethanolamines[23] and fungi have phytoceramide phosphoinositols and mannose-containing headgroups.[24] The Sphingomyelin glycosphingolipids are a diverse family of molecules composed of one or more sugar residues linked via a glycosidic bond to the sphingoid base. Examples of these are the simple and complex glycosphingolipids such as cerebrosides and gangliosides.

Sterol lipids
Sterol lipids, such as cholesterol and its derivatives, are an important component of membrane lipids,[25] along with the glycerophospholipids and sphingomyelins. The steroids, all derived from the same fused four-ring core structure, have different biological roles as hormones and signaling molecules. The eighteen-carbon (C18) steroids include the estrogen family whereas the C19 steroids comprise the androgens such as testosterone and androsterone. The C21 subclass includes the progestogens as well as the glucocorticoids and mineralocorticoids.[26] The secosteroids, comprising various forms of vitamin D, are characterized by cleavage of the B ring of the core structure.[27] Other examples of sterols are the bile acids and their conjugates,[28] which in mammals are oxidized derivatives of cholesterol and are synthesized in the liver. The plant equivalents are the phytosterols, such as -sitosterol, stigmasterol, and brassicasterol; the latter compound is also used as a biomarker for algal growth.[29] The predominant sterol in fungal cell membranes is ergosterol.[30]

Prenol lipids
Prenol lipids are synthesized from the 5 carbon precursors isopentenyl diphosphate and dimethylallyl diphosphate that are produced mainly via the mevalonic acid (MVA) pathway.[31] The simple isoprenoids (linear alcohols, diphosphates, etc.) are formed by the successive addition of C5 units, and are classified according to number of these terpene units. Structures containing greater than 40 carbons are known as polyterpenes. Carotenoids are important simple isoprenoids that function as antioxidants and as precursors of vitamin A.[32] Another biologically important class of molecules is exemplified by the quinones and hydroquinones, which contain an isoprenoid tail attached to a quinonoid core of non-isoprenoid origin.[33] Vitamin E and vitamin K, as well as the ubiquinones, are examples of this class. Prokaryotes synthesize polyprenols (called bactoprenols) in which the terminal isoprenoid unit attached to oxygen remains unsaturated, whereas in animal polyprenols (dolichols) the terminal isoprenoid is reduced.[34]

Lipid

166

Saccharolipids
Saccharolipids describe compounds in which fatty acids are linked directly to a sugar backbone, forming structures that are compatible with membrane bilayers. In the saccharolipids, a monosaccharide substitutes for the glycerol backbone present in glycerolipids and glycerophospholipids. The most familiar saccharolipids are the acylated glucosamine precursors of the LipidA component of the lipopolysaccharides in Gram-negative bacteria. Typical lipidA molecules are disaccharides of glucosamine, which are derivatized with as many as seven fatty-acyl chains. The minimal lipopolysaccharide required for growth in E. coli is Kdo2-Lipid A, a hexa-acylated disaccharide of glucosamine that is glycosylated with two 3-deoxy-D-manno-octulosonic acid (Kdo) residues.[35]

[35] Structure of the saccharolipid Kdo2-Lipid A. Glucosamine residues in blue, Kdo residues in red, acyl chains in black and phosphate groups in green.

Polyketides
Polyketides are synthesized by polymerization of acetyl and propionyl subunits by classic enzymes as well as iterative and multimodular enzymes that share mechanistic features with the fatty acid synthases. They comprise a large number of secondary metabolites and natural products from animal, plant, bacterial, fungal and marine sources, and have great structural diversity.[36] [37] Many polyketides are cyclic molecules whose backbones are often further modified by glycosylation, methylation, hydroxylation, oxidation, and/or other processes. Many commonly used anti-microbial, anti-parasitic, and anti-cancer agents are polyketides or polyketide derivatives, such as erythromycins, tetracyclines, avermectins, and antitumor epothilones.[38]

Biological functions
Membranes
Eukaryotic cells are compartmentalized into membrane-bound organelles that carry out different biological functions. The glycerophospholipids are the main structural component of biological membranes, such as the cellular plasma membrane and the intracellular membranes of organelles; in animal cells the plasma membrane physically separates the intracellular components from the extracellular environment. The glycerophospholipids are amphipathic molecules (containing both hydrophobic and hydrophilic regions) that contain a glycerol core linked to two fatty acid-derived "tails" by ester linkages and to one "head" group by a phosphate ester linkage. While glycerophospholipids are the major component of biological membranes, other non-glyceride lipid components such as sphingomyelin and sterols (mainly cholesterol in animal cell membranes) are also found in biological membranes.[39] In plants and algae, the galactosyldiacylglycerols,[40] and sulfoquinovosyldiacylglycerol,[14] which lack a phosphate group, are important components of membranes of chloroplasts and related organelles and are the most abundant lipids in photosynthetic tissues, including those of higher plants, algae and certain bacteria.

Lipid Bilayers have been found to exhibit high levels of birefringence, which can be used to probe the degree of order (or disruption) within the bilayer using techniques such as dual polarization interferometry A biological membrane is a form of lipid bilayer. The formation of lipid bilayers is an energetically preferred process when the glycerophospholipids described above are in an aqueous environment.[41] In an aqueous system, the polar heads of lipids align towards the polar, aqueous environment, while the hydrophobic tails minimize their contact with water and tend to cluster together, forming a vesicle; depending on the concentration of the lipid, this biophysical interaction may result in the formation of micelles, liposomes, or lipid bilayers. Other aggregations are also observed and form part of the polymorphism of amphiphile (lipid) behavior. Phase behavior is an area of study within biophysics and is the subject of current academic research.[42] [43] Micelles and bilayers form in the polar medium by a process known as the hydrophobic effect.[44] When dissolving a lipophilic or amphiphilic substance in a polar environment, the polar molecules (i.e., water in an aqueous solution) become more Self-organization of phospholipids: a spherical liposome, a micelle, ordered around the dissolved lipophilic substance, since and a lipid bilayer. the polar molecules cannot form hydrogen bonds to the lipophilic areas of the amphiphile. So in an aqueous environment, the water molecules form an ordered "clathrate" cage around the dissolved lipophilic molecule.[45]

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Energy storage
Triacylglycerols, stored in adipose tissue, are a major form of energy storage in animals. The adipocyte, or fat cell, is designed for continuous synthesis and breakdown of triacylglycerols, with breakdown controlled mainly by the activation of hormone-sensitive enzyme lipase.[46] The complete oxidation of fatty acids provides high caloric content, about 9kcal/g, compared with 4kcal/g for the breakdown of carbohydrates and proteins. Migratory birds that must fly long distances without eating use stored energy of triacylglycerols to fuel their flights.[47]

Signaling
In recent years, evidence has emerged showing that lipid signaling is a vital part of the cell signaling.[48] Lipid signaling may occur via activation of G protein-coupled or nuclear receptors, and members of several different lipid categories have been identified as signaling molecules and cellular messengers.[49] These include sphingosine-1-phosphate, a sphingolipid derived from ceramide that is a potent messenger molecule involved in regulating calcium mobilization,[50] cell growth, and apoptosis;[51] diacylglycerol (DAG) and the phosphatidylinositol phosphates (PIPs), involved in calcium-mediated activation of protein kinase C;[52] the prostaglandins, which are one type of fatty-acid derived eicosanoid involved in inflammation and immunity;[53] the steroid hormones such as estrogen, testosterone and cortisol, which modulate a host of functions such as reproduction, metabolism and blood pressure; and the oxysterols such as 25-hydroxy-cholesterol that are liver X receptor agonists.[54]

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Other functions
The "fat-soluble" vitamins (A, D, E and K) which are isoprene-based lipids are essential nutrients stored in the liver and fatty tissues, with a diverse range of functions. Acyl-carnitines are involved in the transport and metabolism of fatty acids in and out of mitochondria, where they undergo beta oxidation.[55] Polyprenols and their phosphorylated derivatives also play important transport roles, in this case the transport of oligosaccharides across membranes. Polyprenol phosphate sugars and polyprenol diphosphate sugars function in extra-cytoplasmic glycosylation reactions, in extracellular polysaccharide biosynthesis (for instance, peptidoglycan polymerization in bacteria), and in eukaryotic protein N-glycosylation.[56] [57] Cardiolipins are a subclass of glycerophospholipids containing four acyl chains and three glycerol groups that are particularly abundant in the inner mitochondrial membrane.[58] [59] [60] They are believed to activate enzymes involved with oxidative phosphorylation.[61] Lipids also form the basis of steroid hormones. [62]

Metabolism
The major dietary lipids for humans and other animals are animal and plant triglycerides, sterols, and membrane phospholipids. The process of lipid metabolism synthesizes and degrades the lipid stores and produces the structural and functional lipids characteristic of individual tissues.

Biosynthesis
In animals, when there is an oversupply of dietary carbohydrate, the excess carbohydrate is converted to triacylglycerol. This involves the synthesis of fatty acids from acetyl-CoA and the esterification of fatty acids in the production of triacylglycerol, a process called lipogenesis.[63] Fatty acids are made by fatty acid synthases that polymerize and then reduce acetyl-CoA units. The acyl chains in the fatty acids are extended by a cycle of reactions that add the acetyl group, reduce it to an alcohol, dehydrate it to an alkene group and then reduce it again to an alkane group. The enzymes of fatty acid biosynthesis are divided into two groups, in animals and fungi all these fatty acid synthase reactions are carried out by a single multifunctional protein,[64] while in plant plastids and bacteria separate enzymes perform each step in the pathway.[65] [66] The fatty acids may be subsequently converted to triacylglycerols that are packaged in lipoproteins and secreted from the liver. The synthesis of unsaturated fatty acids involves a desaturation reaction, whereby a double bond is introduced into the fatty acyl chain. For example, in humans, the desaturation of stearic acid by stearoyl-CoA desaturase-1 produces oleic acid. The doubly unsaturated fatty acid linoleic acid as well as the triply unsaturated -linolenic acid cannot be synthesized in mammalian tissues, and are therefore essential fatty acids and must be obtained from the diet.[67] Triacylglycerol synthesis takes place in the endoplasmic reticulum by metabolic pathways in which acyl groups in fatty acyl-CoAs are transferred to the hydroxyl groups of glycerol-3-phosphate and diacylglycerol.[68] Terpenes and isoprenoids, including the carotenoids, are made by the assembly and modification of isoprene units donated from the reactive precursors isopentenyl pyrophosphate and dimethylallyl pyrophosphate.[69] These precursors can be made in different ways. In animals and archaea, the mevalonate pathway produces these compounds from acetyl-CoA,[70] while in plants and bacteria the non-mevalonate pathway uses pyruvate and glyceraldehyde 3-phosphate as substrates.[69] [71] One important reaction that uses these activated isoprene donors is steroid biosynthesis. Here, the isoprene units are joined together to make squalene and then folded up and formed into a set of rings to make lanosterol.[72] Lanosterol can then be converted into other steroids such as cholesterol and ergosterol.[72] [73]

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169

Degradation
Beta oxidation is the metabolic process by which fatty acids are broken down in the mitochondria and/or in peroxisomes to generate acetyl-CoA. For the most part, fatty acids are oxidized by a mechanism that is similar to, but not identical with, a reversal of the process of fatty acid synthesis. That is, two-carbon fragments are removed sequentially from the carboxyl end of the acid after steps of dehydrogenation, hydration, and oxidation to form a beta-keto acid, which is split by thiolysis. The acetyl-CoA is then ultimately converted into ATP, CO2, and H2O using the citric acid cycle and the electron transport chain. Hence the Krebs Cycle can start at acetyl-CoA when fat is being broken down for energy if there is little or no glucose available. The energy yield of the complete oxidation of the fatty acid palmitate is 106 ATP.[74] Unsaturated and odd-chain fatty acids require additional enzymatic steps for degradation.

Nutrition and health

Most of the lipid found in food is in the form of triacylglycerols, cholesterol, and phospholipids. A minimum amount of dietary fat is necessary to facilitate absorption of fat-soluble vitamins (A, D, E, and K) and carotenoids.[75] Humans and other mammals have a dietary requirement for certain essential fatty acids, such as linoleic acid (an omega-6 fatty acid) and alpha-linolenic acid (an omega-3 fatty acid) because they cannot be synthesized from simple precursors in the diet.[67] Both of these fatty acids are 18-carbon polyunsaturated fatty acids differing in the number and position of the double bonds. Most vegetable oils are rich in linoleic acid (safflower, sunflower, and corn oils). Alpha-linolenic acid is found in the green leaves of plants, and in selected seeds, nuts, and legumes (in particular flax, rapeseed, walnut, and soy).[76] Fish oils are particularly rich in the longer-chain omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA).[77] A large number of studies have shown positive health benefits associated with consumption of omega-3 fatty acids on infant development, cancer, cardiovascular diseases, and various mental illnesses, such as depression, attention-deficit hyperactivity disorder, and dementia.[78] [79] In contrast, it is now well-established that consumption of trans fats, such as those present in partially hydrogenated vegetable oils, are a risk factor for cardiovascular disease.[80] [81] [82] A few studies have suggested that total dietary fat intake is linked to an increased risk of obesity[83] [84] and diabetes.[85] [86] However, a number of very large studies, including the Women's Health Initiative Dietary Modification Trial, an eight year study of 49,000 women, the Nurses' Health Study and the Health Professionals Follow-up Study, revealed no such links.[87] [88] [89] None of these studies suggested any connection between percentage of calories from fat and risk of cancer, heart disease, or weight gain. The Nutrition Source, a website maintained by the Department of Nutrition at the Harvard School of Public Health, summarizes the current evidence on the impact of dietary fat: "Detailed researchmuch of it done at Harvardshows that the total amount of fat in the diet isn't really linked with weight or disease."[90]

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References
[1] [2] [3] [4] Stryer et al., p. 328. Maitland, Jr Jones (1998). Organic Chemistry. W W Norton & Co Inc (Np). p.139. ISBN0-393-97378-6. Stryer et al., p. 330. Fahy E, Subramaniam S, Murphy R, Nishijima M, Raetz C, Shimizu T, Spener F, Van Meer G, Wakelam M and Dennis E.A (2009). "Update of the LIPID MAPS comprehensive classification system for lipids". Journal of Lipid Research 50: S9S14. doi:10.1194/jlr.R800095-JLR200. PMID19098281. [5] Michelle A, Hopkins J, McLaughlin CW, Johnson S, Warner MQ, LaHart D, Wright JD (1993). Human Biology and Health. Englewood Cliffs, New Jersey, USA: Prentice Hall. ISBN0-13-981176-1. OCLC32308337. [6] Vance JE, Vance DE (2002). Biochemistry of Lipids, Lipoproteins and Membranes. Amsterdam: Elsevier. ISBN0444511393. OCLC51001207. [7] Brown HA, ed (2007). Lipodomics and Bioactive Lipids: Mass Spectrometry Based Lipid Analysis, Volume 432 (Methods in Enzymology). Boston: Academic Press. ISBN0123738954. OCLC166624879. [8] Hunt SM, Groff JL, Gropper SAS (1995). Advanced Nutrition and Human Metabolism. Belmont, CA: West Pub. Co. p.98. ISBN0-314-04467-1. [9] Devlin, pp. 19395. [10] Hunter JE (November 2006). "Dietary trans fatty acids: review of recent human studies and food industry responses". Lipids 41 (11): 96792. doi:10.1007/s11745-006-5049-y. PMID17263298. [11] Fezza F, De Simone C, Amadio D, Maccarrone M (2008). "Fatty acid amide hydrolase: a gate-keeper of the endocannabinoid system". Subcellular Biochemistry 49: 10132. doi:10.1007/978-1-4020-8831-5_4. PMID18751909. [12] Coleman RA, Lee DP (2004). "Enzymes of triacylglycerol synthesis and their regulation". Progress in Lipid Research 43 (2): 13476. doi:10.1016/S0163-7827(03)00051-1. PMID14654091. [13] van Holde and Mathews, p. 63031. [14] Hlzl G, Drmann P (2007). "Structure and function of glycoglycerolipids in plants and bacteria" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0163-7827(07)00015-X). Progress in Lipid Research 46 (5): 22543. doi:10.1016/j.plipres.2007.05.001. PMID17599463. . [15] Honke K, Zhang Y, Cheng X, Kotani N, Taniguchi N (2004). "Biological roles of sulfoglycolipids and pathophysiology of their deficiency" (http:/ / www. kluweronline. com/ art. pdf?issn=0282-0080& volume=21& page=59). Glycoconjugates Journal 21 (12): 5962. doi:10.1023/B:GLYC.0000043749.06556.3d. PMID15467400. . [16] Farooqui AA, Horrocks LA, Farooqui T (2000). "Glycerophospholipids in brain: their metabolism, incorporation into membranes, functions, and involvement in neurological disorders" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0009-3084(00)00128-6). Chemistry and Physics of Lipids 106 (1): 129. doi:10.1016/S0009-3084(00)00128-6. PMID10878232. . Retrieved 2009-04-12. [17] Ivanova PT, Milne SB, Byrne MO, Xiang Y, Brown HA (2007). "Glycerophospholipid identification and quantitation by electrospray ionization mass spectrometry". Methods in Enzymology 432: 2157. doi:10.1016/S0076-6879(07)32002-8. PMID17954212. [18] van Holde and Mathews, p. 844. [19] Paltauf F (1994). "Ether lipids in biomembranes". Chemistry and Physics of Lipids 74 (2): 10139. doi:10.1016/0009-3084(94)90054-X. PMID7859340. [20] Merrill AH, Sandhoff K. (2002). "Sphingolipids: metabolism and cell signaling",in New Comprehensive Biochemistry: Biochemistry of Lipids, Lipoproteins,and Membranes, Vance, D.E. and Vance, J.E., eds. Elsevier Science, NY. Ch. 14. [21] Devlin, pp. 42122. [22] Hori T, Sugita M (1993). "Sphingolipids in lower animals". Prog. Lipid Res 32 (1): 2545. doi:10.1016/0163-7827(93)90003-F. PMID8415797. [23] Wiegandt H (1992). "Insect glycolipids". Biochimica et Biophysica Acta 1123 (2): 11726. PMID1739742. [24] Guan X, Wenk MR (2008). "Biochemistry of inositol lipids". Frontiers in Bioscience 13: 323951. doi:10.2741/2923. PMID18508430. [25] Bach D, Wachtel E (2003). "Phospholipid/cholesterol model membranes: formation of cholesterol crystallites". Biochim Biophys Acta 1610 (2): 18797. doi:10.1016/S0005-2736(03)00017-8. PMID12648773. [26] Stryer et al., p. 749. [27] Bouillon R, Verstuyf A, Mathieu C, Van Cromphaut S, Masuyama R, Dehaes P, Carmeliet G (2006). "Vitamin D resistance" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S1521-690X(06)00079-0). Best Practice & Research. Clinical Endocrinology & Metabolism 20 (4): 62745. doi:10.1016/j.beem.2006.09.008. PMID17161336. . [28] Russell DW (2003). "The enzymes, regulation, and genetics of bile acid synthesis". Annual Review of Biochemistry 72: 13774. doi:10.1146/annurev.biochem.72.121801.161712. PMID12543708. [29] Villinski JC, Hayes JM, Brassell SC, Riggert VL, Dunbar RB (2008). "Sedimentary sterols as biogeochemical indicators in the Southern Ocean". Organic Geochemistry 39 (5): 56788. doi:10.1016/j.orggeochem.2008.01.009. [30] Deacon J (2005). Fungal Biology. Cambridge, MA: Blackwell Publishers. p.342. ISBN1-4051-3066-0. [31] Kuzuyama T, Seto H (2003). "Diversity of the biosynthesis of the isoprene units". Natural Product Reports 20 (2): 17183. doi:10.1039/b109860h. PMID12735695. [32] Rao AV, Rao LG (2007). "Carotenoids and human health" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S1043-6618(07)00035-7). Pharmacological Research : the Official Journal of the Italian Pharmacological Society 55 (3): 20716. doi:10.1016/j.phrs.2007.01.012.

Lipid
PMID17349800. . [33] Brunmark A, Cadenas E (1989). "Redox and addition chemistry of quinoid compounds and its biological implications". Free Radical Biology & Medicine 7 (4): 43577. doi:10.1016/0891-5849(89)90126-3. PMID2691341. [34] Swiezewska E, Danikiewicz W (2005). "Polyisoprenoids: structure, biosynthesis and function" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0163-7827(05)00022-6). Progress in Lipid Research 44 (4): 23558. doi:10.1016/j.plipres.2005.05.002. PMID16019076. . [35] Raetz CR, Garrett TA, Reynolds CM, Shaw WA, Moore JD, Smith DC Jr, Ribeiro AA, Murphy RC,Ulevitch RJ, Fearns C, Reichart D, Glass CK, Benner C, Subramaniam S, Harkewicz R, Bowers-Gentry RC, Buczynski MW, Cooper JA, Deems RA, Dennis EA (2006). "Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4". Journal of Lipid Research 47 (5): 1097111. doi:10.1194/jlr.M600027-JLR200. PMID16479018. [36] Walsh CT (2004). "Polyketide and nonribosomal peptide antibiotics: modularity and versatility". Science 303 (5665): 180510. doi:10.1126/science.1094318. PMID15031493. [37] Caffrey P, Aparicio JF, Malpartida F, Zotchev SB (2008). "Biosynthetic engineering of polyene macrolides towards generation of improved antifungal and antiparasitic agents" (http:/ / www. bentham-direct. org/ pages/ content. php?CTMC/ 2008/ 00000008/ 00000008/ 0003R. SGM). Current Topics in Medicinal Chemistry 8 (8): 63953. doi:10.2174/156802608784221479. PMID18473889. . Retrieved 2009-04-12. [38] Minto RE, Blacklock BJ (2008). "Biosynthesis and function of polyacetylenes and allied natural products" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0163-7827(08)00015-5). Progress in Lipid Research 47 (4): 233306. doi:10.1016/j.plipres.2008.02.002. PMC2515280. PMID18387369. . Retrieved 2009-04-12. [39] Stryer et al., pp. 329331 [40] Heinz E.(1996). Plant glycolipids: structure, isolation and analysis. in Advances in Lipid Methodology - 3, pp. 211332 (ed. W.W. Christie, Oily Press, Dundee) [41] Stryer et al., pp. 33334. [42] van Meer G, Voelker DR, Feigenson GW (2008). "Membrane lipids: where they are and how they behave". Nature Reviews. Molecular Cell Biology 9 (2): 11224. doi:10.1038/nrm2330. PMC2642958. PMID18216768. [43] Feigenson GW (2006). "Phase behavior of lipid mixtures". Nature Chemical Biology 2 (11): 56063. doi:10.1038/nchembio1106-560. PMC2685072. PMID17051225. [44] Wiggins PM (1990). "Role of water in some biological processes". Microbiological Reviews 54 (4): 43249. PMC372788. PMID2087221. [45] Raschke TM, Levitt M (2005). "Nonpolar solutes enhance water structure within hydration shells while reducing interactions between them". Proceedings of the National Academy of Sciences U.S.A 102 (19): 677782. doi:10.1073/pnas.0500225102. PMC1100774. PMID15867152. [46] Brasaemle DL (December 2007). "Thematic review series: adipocyte biology. The perilipin family of structural lipid droplet proteins: stabilization of lipid droplets and control of lipolysis" (http:/ / www. jlr. org/ cgi/ content/ full/ 48/ 12/ 2547). J. Lipid Res 48 (12): 254759. doi:10.1194/jlr.R700014-JLR200. PMID17878492. . [47] Stryer et al., p. 619. [48] Wang X (2004). "Lipid signaling". Current Opinions in Plant Biology 7 (3): 32936. doi:10.1016/j.pbi.2004.03.012. PMID15134755. [49] Eyster KM (2007). "The membrane and lipids as integral participants in signal transduction". Advances in Physiology Education 31 (1): 516. doi:10.1152/advan.00088.2006. PMID17327576. [50] Hinkovska-Galcheva V, VanWay SM, Shanley TP, Kunkel RG (2008). "The role of sphingosine-1-phosphate and ceramide-1-phosphate in calcium homeostasis". Current Opinion in Investigational Drugs 9 (11): 1192205. PMID18951299. [51] Saddoughi SA, Song P, Ogretmen B (2008). "Roles of bioactive sphingolipids in cancer biology and therapeutics". Subcellular Biochemistry 49: 41340. doi:10.1007/978-1-4020-8831-5_16. PMC2636716. PMID18751921. [52] Klein C, Malviya AN (2008). "Mechanism of nuclear calcium signaling by inositol 1,4,5-trisphosphate produced in the nucleus, nuclear located protein kinase C and cyclic AMP-dependent protein kinase" (http:/ / www. bioscience. org/ 2008/ v13/ af/ 2756/ fulltext. htm). Frontiers in Bioscience 13: 120626. doi:10.2741/2756. PMID17981624. . [53] Boyce JA (2008). "Eicosanoids in asthma, allergic inflammation, and host defense" (http:/ / www. bentham-direct. org/ pages/ content. php?CMM/ 2008/ 00000008/ 00000005/ 0003M. SGM). Current Molecular Medicine 8 (5): 33549. doi:10.2174/156652408785160989. PMID18691060. . [54] Betowski J (2008). "Liver X receptors (LXR) as therapeutic targets in dyslipidemia". Cardiovascular Therapy 26 (4): 297316. doi:10.1111/j.1755-5922.2008.00062.x. PMID19035881. [55] Indiveri C, Tonazzi A, Palmieri F (October 1991). "Characterization of the unidirectional transport of carnitine catalyzed by the reconstituted carnitine carrier from rat liver mitochondria". Biochim. Biophys. Acta 1069 (1): 1106. doi:10.1016/0005-2736(91)90110-T. PMID1932043. [56] Parodi AJ, Leloir LF (April 1979). "The role of lipid intermediates in the glycosylation of proteins in the eucaryotic cell". Biochim. Biophys. Acta 559 (1): 137. doi:10.1016/0304-4157(79)90006-6. PMID375981. [57] Helenius A, Aebi M (2001). "Intracellular functions of N-linked glycans". Science 291 (5512): 236469. doi:10.1126/science.291.5512.2364. PMID11269317. [58] M. Nowicki and M. Frentzen (2005). "Cardiolipin synthase of Arabidopsis thaliana". FEBS Letters 579 (10): 21612165. doi:10.1016/j.febslet.2005.03.007. PMID15811335. [59] M. Nowicki (2006). "Characterization of the Cardiolipin Synthase from Arabidopsis thaliana" (http:/ / darwin. bth. rwth-aachen. de/ opus/ volltexte/ 2006/ 1629/ ). Ph.D. thesis, RWTH-Aachen University. .

171

Lipid
[60] Gohil VM, Greenberg ML (2009). "Mitochondrial membrane biogenesis: phospholipids and proteins go hand in hand" (http:/ / jcb. rupress. org/ cgi/ content/ full/ 184/ 4/ 469). Journal of Cell Biology 184 (4): 46972. doi:10.1083/jcb.200901127. PMC2654137. PMID19237595. . [61] Hoch FL (1992). "Cardiolipins and biomembrane function". Biochimica et Biophysica Acta 1113 (1): 71133. PMID10206472. [62] http:/ / www. elmhurst. edu/ ~chm/ vchembook/ 556steroids. html [63] Stryer et al., p. 634. [64] Chirala S, Wakil S (2004). "Structure and function of animal fatty acid synthase". Lipids 39 (11): 104553. doi:10.1007/s11745-004-1329-9. PMID15726818. [65] White S, Zheng J, Zhang Y (2005). "The structural biology of type II fatty acid biosynthesis". Annual Review of Biochemistry 74: 791831. doi:10.1146/annurev.biochem.74.082803.133524. PMID15952903. [66] Ohlrogge J, Jaworski J (1997). "Regulation of fatty acid synthesis". Annual Review of Plant Physiology and Plant Molecular Biology 48: 109136. doi:10.1146/annurev.arplant.48.1.109. PMID15012259. [67] Stryer et al., p. 643. [68] Stryer et al., pp. 73339. [69] Kuzuyama T, Seto H (2003). "Diversity of the biosynthesis of the isoprene units". Natural Product Reports 20 (2): 17183. doi:10.1039/b109860h. PMID12735695. [70] Grochowski L, Xu H, White R (2006). "Methanocaldococcus jannaschii uses a modified mevalonate pathway for biosynthesis of isopentenyl diphosphate". Journal of Bacteriology 188 (9): 319298. doi:10.1128/JB.188.9.3192-3198.2006. PMC1447442. PMID16621811. [71] Lichtenthaler H (1999). "The 1-Dideoxy-D-xylulose-5-phosphate pathway of isoprenoid biosynthesis in plants". Annual Review of Plant Physiology and Plant Molecular Biology 50: 4765. doi:10.1146/annurev.arplant.50.1.47. PMID15012203. [72] Schroepfer G (1981). "Sterol biosynthesis". Annual Review of Biochemistry 50: 585621. doi:10.1146/annurev.bi.50.070181.003101. PMID7023367. [73] Lees N, Skaggs B, Kirsch D, Bard M (1995). "Cloning of the late genes in the ergosterol biosynthetic pathway of Saccharomyces cerevisiaea review". Lipids 30 (3): 22126. doi:10.1007/BF02537824. PMID7791529. [74] Stryer et al., pp. 62526. [75] Bhagavan, p. 903. [76] Russo GL (2009). "Dietary n-6 and n-3 polyunsaturated fatty acids: from biochemistry to clinical implications in cardiovascular prevention" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S0006-2952(08)00777-6). Biochemical Pharmacology 77 (6): 93746. doi:10.1016/j.bcp.2008.10.020. PMID19022225. . [77] Bhagavan, p. 388. [78] Riediger ND, Othman RA, Suh M, Moghadasian MH (2009). "A systemic review of the roles of n-3 fatty acids in health and disease". Journal of the American Dietetic Association 109 (4): 66879. doi:10.1016/j.jada.2008.12.022. PMID19328262. [79] Galli C, Ris P (2009). "Fish consumption, omega 3 fatty acids and cardiovascular disease. The science and the clinical trials". Nutrition and Health (Berkhamsted, Hertfordshire) 20 (1): 1120. PMID19326716. [80] Micha R, Mozaffarian D (2008). "Trans fatty acids: effects on cardiometabolic health and implications for policy". Prostaglandins, Leukotrienes, and Essential Fatty Acids 79 (35): 14752. doi:10.1016/j.plefa.2008.09.008. PMC2639783. PMID18996687. [81] Dalainas I, Ioannou HP (2008). "The role of trans fatty acids in atherosclerosis, cardiovascular disease and infant development". International Angiology: a Journal of the International Union of Angiology 27 (2): 14656. PMID18427401. [82] Mozaffarian D, Willett WC (2007). "Trans fatty acids and cardiovascular risk: a unique cardiometabolic imprint?". Current Atherosclerosis Reports 9 (6): 48693. doi:10.1007/s11883-007-0065-9. PMID18377789. [83] Astrup A, Dyerberg J, Selleck M, Stender S (2008). "Nutrition transition and its relationship to the development of obesity and related chronic diseases". Obesity Review 9 Suppl 1: 4852. doi:10.1111/j.1467-789X.2007.00438.x. PMID18307699. [84] Astrup A (2005). "The role of dietary fat in obesity". Seminars in Vascular Medicine 5 (1): 4047. doi:10.1055/s-2005-871740. PMID15968579. [85] Ma Y. et al (2006). "Low-carbohydrate and high-fat intake among adult patients with poorly controlled type 2 diabetes mellitus". Nutrition 22 (11-12): 11291136. doi:10.1016/j.nut.2006.08.006. PMC2039705. PMID17027229. [86] Astrup A (2008). "Dietary management of obesity". JPEN Journal of Parenteral and Enteral Nutrition 32 (5): 57577. doi:10.1177/0148607108321707. PMID18753397. [87] Beresford SA, Johnson KC, Ritenbaugh C, et al. (2006). "Low-fat dietary pattern and risk of colorectal cancer: the Women's Health Initiative Randomized Controlled Dietary Modification Trial". JAMA: the Journal of the American Medical Association 295 (6): 64354. doi:10.1001/jama.295.6.643. PMID16467233. [88] Howard BV, Manson JE, Stefanick ML, et al. (2006). "Low-fat dietary pattern and weight change over 7 years: the Women's Health Initiative Dietary Modification Trial" (http:/ / jama. ama-assn. org/ cgi/ pmidlookup?view=long& pmid=16391215). JAMA: the Journal of the American Medical Association 295 (1): 3949. doi:10.1001/jama.295.1.39. PMID16391215. . [89] Howard BV, Van Horn L, Hsia J, et al. (2006). "Low-fat dietary pattern and risk of cardiovascular disease: the Women's Health Initiative Randomized Controlled Dietary Modification Trial". JAMA : the Journal of the American Medical Association 295 (6): 65566. doi:10.1001/jama.295.6.655. PMID16467234. [90] "Fats and Cholesterol: Out with the Bad, In with the Good - What Should You Eat? - The Nutrition Source - Harvard School of Public Health" (http:/ / www. hsph. harvard. edu/ nutritionsource/ what-should-you-eat/ fats-full-story/ index. html). . Retrieved 2009-05-12.

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Bibliography
Bhagavan NV (2002). Medical Biochemistry (http://books.google.com/?id=vT9YttFTPi0C& printsec=frontcover). San Diego: Harcourt/Academic Press. ISBN0-12-095440-0. Devlin TM (1997). Textbook of Biochemistry: With Clinical Correlations (4th ed.). Chichester: John Wiley & Sons. ISBN0-471-17053-4. Stryer L, Berg JM, Tymoczko JL (2007). Biochemistry (6th ed.). San Francisco: W.H. Freeman. ISBN0-7167-8724-5. Van Holde KE, Mathews CK (1996). Biochemistry (2nd ed.). Menlo Park, Calif: Benjamin/Cummings Pub. Co. ISBN0-8053-3931-0.

External links
Introductory List of lipid-related web sites (http://www.cyberlipid.org/cyberlip/link0041.htm) Nature Lipidomics Gateway (http://www.lipidmaps.org/) - Round-up and summaries of recent lipid research Lipid Library (http://www.lipidlibrary.co.uk/) - General reference on lipid chemistry and biochemistry Cyberlipid.org (http://www.cyberlipid.org/) - Resources and history for lipids.

Molecular Computer Simulations (http://www.fos.su.se/~sasha/Lipid_membranes.html) - Modeling of Lipid Membranes Lipids, Membranes and Vesicle Trafficking (http://www.biochemweb.org/lipids_membranes.shtml) - The Virtual Library of Biochemistry and Cell Biology Nomenclature IUPAC nomenclature of lipids (http://www.chem.qmul.ac.uk/iupac/lipid/) IUPAC glossary entry for the lipid class of molecules (http://www.chem.qmul.ac.uk/iupac/class/lipid.html) Databases LIPID MAPS (http://www.lipidmaps.org/data/databases.html) - Comprehensive lipid and lipid-associated gene/protein databases. LipidBank (http://lipidbank.jp/) - Japanese database of lipids and related properties, spectral data and references. LIPIDAT (http://www.lipidat.tcd.ie/) - Database composed mainly of phospholipids and associated thermodynamic data. General ApolloLipids (http://www.apollolipids.org/) - Provides dyslipidemia and cardiovascular disease prevention and treatment information as well as continuing medical education programs National Lipid Association (http://www.lipid.org/) - Professional medical education organization for health care professionals who seek to prevent morbidity and mortality stemming from dyslipidemias and other cholesterol-related disorders.

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Biological membrane
A biological membrane or biomembrane is an enclosing or separating membrane that acts as a selective barrier, within or around a cell. It consists of a lipid bilayer with embedded proteins that may constitute close to 50% of membrane content.[1] The cellular membranes should not be confused with isolating tissues formed by layers of cells, such as mucous and basement membranes.

Function
Membranes in cells typically define enclosed spaces or compartments in which cells may maintain a chemical or biochemical environment that differs from the outside. For example, the membrane around peroxisomes shields the rest of the cell from peroxides, and the cell membrane separates a cell from its surrounding medium. Most organelles are defined by such membranes, and are called "membrane-bound" organelles.

Cross section view of the structures that can be formed by phospholipids in aqueous solutions

Probably the most important feature of a biomembrane is that it is a selectively permeable structure. This means that the size, charge, and other chemical properties of the atoms and molecules attempting to cross it will determine whether they succeed in doing so. Selective permeability is essential for effective separation of a cell or organelle from its surroundings. Biological membranes also have certain mechanical or elastic properties. Particles that are required for cellular function but are unable to diffuse freely across a membrane enter through a membrane transport protein or are taken in by means of endocytosis.

Diversity of biological membranes

Many types of specialized plasma membranes can separate cell from external environment: apical, basolateral, presynaptic and postsynaptic ones, membranes of flagella, cilia, microvillus, filopodia and lamellipodia, the sarcolemma of muscle cells, as well as specialized myelin and dendritic spine membranes of neurons. Plasma membranes can also form different types of "supramembrane" structures such as caveola, postsynaptic density, podosome, invadopodium, desmosome, hemidesmosome, focal adhesion, and cell junctions. These types of membranes differ in lipid and protein composition. Distinct types of membranes also create intracellular organelles: endosome; smooth and rough endoplasmic reticulum; sarcoplasmic reticulum; Golgi apparatus; lysosome; mitochondrion (inner and outer membranes); nucleus (inner and outer membranes); peroxisome; vacuole; cytoplasmic granules; cell vesicles (phagosome, autophagosome, clathrin-coated vesicles, COPI-coated and COPII-coated vesicles) and secretory vesicles (including synaptosome, acrosomes, melanosomes, and chromaffin granules). Different types of biological membranes have diverse lipid and protein compositions. The content of membranes defines their physical and biological properties. Some components of membranes play a key role in medicine, such as the efflux pumps that pump drugs out of a cell.

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References
[1] Mark L. Latash (2007). Neurophysiological basis of movement. Human Kinetics Publishers. ISBN978-0-7360-6367-8.

von Heijne G, Rees D (August 2008). "Membranes: reading between the lines" (http://linkinghub.elsevier.com/ retrieve/pii/S0959-440X(08)00091-2). Curr. Opin. Struct. Biol. 18 (4): 4035. doi:10.1016/j.sbi.2008.06.003. PMID18634876.

External links
MeSH Membranes (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=&term=Membranes)

Membrane protein
A membrane protein is a protein molecule that is attached to, or associated with the membrane of a cell or an organelle. More than half of all proteins interact with membranes.

Function
Biological membranes consist of a phospholipid bilayer and a variety of proteins that accomplish vital biological functions. Structural proteins are attached to microfilaments in the cytoskeleton which ensures stability of the cell. Cell adhesion molecules allow cells to identify each other and interact. Such proteins are involved in immune response, for example. Membrane enzymes produce a variety of substances essential for cell function. Membrane receptor proteins serve as connection between the cell's internal and external environments. Transport proteins play an important role in the maintenance of concentrations of ions. These transport proteins come in two forms: carrier proteins and channel proteins.
Crystal structure of Potassium channel KvAP. Calculated hydrocarbon boundaries of the lipid bilayer are indicated by red and blue dots.

[ [membrane cells]] are the is a biological membrane that separates the interior of all cells from the outside environment

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Main categories
Membrane proteins can be divided into several categories:[1] Integral membrane proteins which are permanently bound to the lipid bilayer Peripheral membrane proteins that are temporarily associated with lipid bilayer or with integral membrane proteins Lipid-anchored proteins bound to lipid bilayer bound through lipidated amino acid residues In addition, pore-forming toxins and many antibacterial peptides are water-soluble molecules, but undergo a conformational transition upon association with lipid bilayer and become reversibly or irreversibly membrane-associated. A slightly different classification is to divide all membrane proteins to integral and amphitropic.[2] The amphitropic are proteins that can exist in two alternative states: a water-soluble and a lipid bilayer-bound. The amphitropic protein category includes water-soluble channel-forming polypeptide toxins, which associate irreversibly with membranes, but excludes peripheral proteins that interact with other membrane proteins rather than with lipid bilayer.

Integral membrane proteins

Integral membrane proteins are permanently attached to the membrane. They can be defined as those proteins which require a detergent (such as SDS or Triton X-100) or some other apolar solvent to be displaced. They can be classified according to their relationship with the bilayer: Integral polytopic proteins, also known as "transmembrane proteins," are proteins that are permanently attached to the lipid membrane and span across the membrane (at least once). The transmembrane regions of the proteins are either beta-barrels or alpha-helical. The alpha-helical domains are present in all types of biological membranes including outer membranes. The beta-barrels were found only in outer membranes of Gram-negative bacteria, lipid-rich cell walls of a few Gram-positive bacteria, and outer membranes of mitochondria and chloroplasts. Integral monotopic proteins are proteins that are permanently attached to the lipid membrane from only one side and do not span across the membrane.

Peripheral membrane proteins

Peripheral membrane proteins are also known as extrinsic proteins, they do not interact with the hydrophobic core of the lipid bilayer. Some peripheral membrane proteins are located at the outer part of the plasma membrane (exoplasmic). They interact with the membrane indirectly by binding to the integral membrane proteins.
[3]

Peripheral membrane proteins are temporarily attached either to the lipid bilayer or to integral proteins by a combination of hydrophobic, electrostatic, and other non-covalent interactions. Peripheral proteins dissociate following treatment with a polar reagent, such as a solution with an elevated pH or high salt concentrations. Integral and peripheral proteins may be post-translationally modified, with added fatty acid or prenyl chains, or GPI (glycosylphosphatidylinositol), which may be anchored in the lipid bilayer.

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Polypeptide toxins
Polypeptide toxins, such as colicins or hemolysins, and certain proteins involved in apoptosis, are sometimes considered a separate category. These proteins are water-soluble but can aggregate and associate irreversibly with the lipid bilayer and form alpha-helical or beta-barrel transmembrane channels.

Intracellular localization
Proteins are specifically targeted to many different types of biological membranes [4]

Membrane protein complexes

Membrane proteins commonly function as complexes. These complexes are vital to cellular function. Understanding how these complexes are assembled, degraded, and their composition are crucial to understanding their function and regulation. Reoccurring in recent literature are the ideas that: membrane protein complexes assemble in an orderly fashion, chaperones aid assembly by preventing unfavorable interactions, and membrane proteins can be interchanged in existing complexes. Membrane protein complexes assemble through the orderly assembly of intermediates. For example, the simple membrane-embedded four subunit complex, cytochrome bo3 of Escherichia coli, is assembled via two intermediate complexes. This suggests a linearly organized assembly pathway. Although interactions between other subunits could lead to the formation of many intermediates, they do not occur. Ordered assembly could be the cell's protection against harmful intermediates. Chaperones interact with membrane proteins guiding their assembly. They aid in preventing the assembly of dead-end and toxic intermediates, as well as unwanted aggregations. Via chaperones assembly can occur through inactive intermediates potentially preventing damaging interactions they could cause. Membrane protein complexes are not fixed entities. Though a process called dynamic exchange, membrane proteins are exchanged in and out of exsitisting protein complexes. This has its implications as a repair mechanism and in regulation. [5]

Assembly of membrane protein

There are two different ways a membrane protein could undergoes during construction are Constitutive membrane protein and non-constitutive membrane protein.

Constitutive membrane protein

The messengerRNA attaches to the translocon which is located to the cell membrane. The mRNA is translated into the translocon transmembrane tunnel. After the synthesis of protein the mRNA is released which closes the translocon and the protein is released in the bilayer membrane. When the protein is left in the membrane bilayer more protein folding occurs creating its final 3D structure (White).

Non-constitutive membrane protein

Examples of non-constitutive membrane proteins are toxins and antimicrobialpeptides. These membrane proteins inserts into specific target membranes by physicochemical processes. It inserts usually before, during, or after oligomerization into the target membrane (White).[6]

Membrane protein structures

In membrane proteins the two known structural classes of membrane proteins are alpha-helical bundle and beta-barrel porin. The portion of the membrane proteins that are attached to the lipid bilayer are consisting of hydophobic amino acids only. This is done so that the peptide bonds' carbonyl and amine will react with each other instead of the hydrophobic surrounding. The portion of the protein that is not touching the lipid bilayer and is

Membrane protein protruding out of the cell membrane are usually hydrophilic amino acids.[7] The structures of membrane proteins are stabilized by weak interactions and influenced by additional interactions with the solubilizing environment. The influence of the environment on membrane protein structures is especially significant. Despite the significant functional importance of membrane proteins, the structural biology has been particularly challenging as shown by the low number of membrane protein structures determined. Integral membrane proteins are present in a heterogeneous environment that poses major obstacles for existing structural methodologies. Many of the successful membrane protein structures are characterized by X-ray crystallography and are very large structures in which the interactions with the membrane mimetic environments can be anticipated to be small in comparison to those within the protein structures. The small domains are particularly sensitive to the influence of membrane mimetic environments, potentially leading to non-native structures. Fortunately, there are many sample preparation conditions that can be chosen for crystallization and for solution NMR. All membrane protein structural biology should be subjected to careful scrutiny; through a combination of structural methodologies it should be possible to achieve an understanding of the native functional state for membrane protein structures.[8]

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References
White, Stephen. General Principle of Membrane Protein Folding and Stability. Stephen White Laboratory Homepage. 10 Nov. 2009. web.
[1] Gerald Karp (2009). Cell and Molecular Biology: Concepts and Experiments (http:/ / books. google. com/ books?id=arRGYE0GxRQC& pg=PA128). John Wiley and Sons. pp.128. ISBN9780470483374. . Retrieved 13 November 2010. [2] Johnson JE, Cornell RB (1999). "Amphitropic proteins: regulation by reversible membrane interactions (review)". Mol. Membr. Biol. 16 (3): 217235. doi:10.1080/096876899294544. PMID10503244. [3] Lodish H, Berk A, Zipursky SL, et al. Molecular Cell Biology. 4th edition. New York: W. H. Freeman; 2000. [4] Classification of membrane proteins with known 3D structure to different membrane types (http:/ / opm. phar. umich. edu/ atlas. php) [5] Daley, Daniel. 2008,"The Assembly of Membrane Proteins into Complexes", Current Opinion in Structural Biology, 18:420-424. [6] White, Stephen. General Principle of Membrane Protein Folding and Stability. Stephen White Laboratory Homepage. 10 Nov. 2009. web. [7] White, Stephen. General Principle of Membrane Protein Folding and Stability. Stephen White Laboratory Homepage. 10 Nov. 2009. web. [8] Cross, Timothy, Mukesh Sharma, Myunggi Yi, Huan-Xiang Zhou (2010). "Influence of Solubilizing Environments on Membrane Protein Structures"

External links
TCDB (http://www.tcdb.org/) - Transporter Classification database Orientations of Proteins in Membranes (OPM) database (http://opm.phar.umich.edu/) 3D structures of integral and peripheral membrane proteins arranged in the lipid bilayer The Protein Data Bank of Transmembrane Proteins (http://pdbtm.enzim.hu/) 3D models of all transmembrane proteins currently in PDB. Approximate positions of membrane boundary planes were calculated for each PDB entry. List of transmembrane proteins of known 3D structure (http://blanco.biomol.uci.edu/ Membrane_Proteins_xtal.html) TransportDB (http://www.membranetransport.org/) Genomics-oriented database of transporters from TIGR Membrane PDB (http://www.mpdb.tcd.ie/) Database of 3D structures of integral membrane proteins and hydrophobic peptides with an emphasis on crystallization conditions Membrane targeting domains (MeTaDoR) (http://proteomics.bioengr.uic.edu/metador/MeTaDoR.html) Antimicrobial Peptide Database (http://aps.unmc.edu/AP/main.php) MeSH Membrane+proteins (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=&term=Membrane+ proteins) The Human Membrane Proteome (http://www.biomedcentral.com/1741-7007/7/50) - A comprehensive article covering the transmembrane protein component of the human proteome

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Cell membrane
The cell membrane or plasma membrane is a biological membrane that separates the interior of all cells from the outside environment.[1] The cell membrane is selectively permeable to ions and organic molecules and controls the movement of substances in and out of cells.[2] It basically protects the cell from outside forces. It consists of the lipid bilayer with embedded proteins. Cell membranes are involved in a variety of cellular processes such as cell adhesion, ion conductivity and cell signaling and serve as the attachment surface for several extracellular structures, including the cell wall, glycocalyx, and intracellular cytoskeleton.

Function
The cell membrane surrounds the cytoplasm of a cell and, in animal cells, physically separates the intracellular Illustration of a Eukaryotic cell membrane components from the extracellular environment. Fungi, bacteria and plants also have the cell wall which provides a mechanical support for the cell and precludes the passage of larger molecules. The cell membrane also plays a role in anchoring the cytoskeleton to provide shape to the cell, and in attaching to the extracellular matrix and other cells to help group cells together to form tissues. The membrane is differentially permeable and able to regulate what enters and exits the cell, thus facilitating the transport of materials needed for survival. The movement of substances across the membrane can be either passive, occurring without the input of cellular energy, or active, requiring the cell to expend energy in transporting it. The membrane also maintains the cell potential. The cell membrane thus works as a selective filter that allows only certain things to come inside or go outside the cell. To do so, the membrane employs a number of transport mechanisms: 1. Diffusion : Some substances (small molecules, ions) such as carbon dioxide (CO2), oxygen (O2), and water, can move across the plasma membrane by diffusion, which is a passive transport process. 2. Osmosis : Because the membrane acts as a barrier for certain molecules and ions, they can occur in different concentrations on the two sides of the membrane. Such a concentration difference across a semipermeable membrane can set up a osmotic flow for the solvent, in this case water. Water can thus be transported across the membrane by osmosis. 3. Mediated Transport : Nutrients such as sugars and materials of growth such as amino acid must enter the cell, and the waste of metabolism must leave. Such molecules are moved across the membrane by special proteins called transport proteins or permeases. Permeases form a small passageway through the membrane, enabling the solute

Cell membrane molecule to cross the phospholipid bilayer. Permeases are usually quite specific, recognizing and transporting only a limited group of chemical substances, often even only a single substance. 4. Endocytosis : Endocytosis is the process in which cells absorb molecules by engulfing them. The plasma membrane creates a small deformation inward, called an invagination, in which the substance to be transported is captured. The deformation then pinches off from the membrane on the inside of the cell, creating a vesicle containing the captured substance. Endocytosis is a pathway for internalizing solid particles (cell eating or phagocytosis), small molecules and ions (cell drinking or pinocytosis), and macromolecules. Endocytosis requires energy and is thus a form of active transport. 5. Exocytosis : Just as material can be brought into the cell by invagination and formation of a vesicle, the membrane of a vesicle can be fused with the plasma membrane, extruding its contents to the surrounding medium. This is the process of exocytosis. Exocytosis occurs in various cells to remove undigested residues of substances brought in by endocytosis, to secrete substances such as hormones and enzymes, and to transport a substance completely across a cellular barrier. In the process of exocytosis, the undigested waste-containing food vacuole or the secretory vesicle budded from Golgi apparatus, is first moved by cytoskeleton from the interior of the cell to the surface. The vesicle membrane comes in contact with the plasma membrane. The lipid molecules of the two bilayers rearrange themselves and the two membranes are, thus, fused. A passage is formed in the fused membrane and the vesicles discharges its contents outside the cell.

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Prokaryotes
Gram-negative bacteria have a plasma membrane and an outer membrane separated by a periplasmic space. Other prokaryotic species have only a plasma membrane. Prokaryotic cells are also surrounded by a cell wall composed of peptidoglycan (amino acid and sugar). Some eukaryotic cells also have cells walls, but none that are made of peptidoglycan.

Structure
Fluid mosaic model
According to the fluid mosaic model of S.J. Singer and G.L. Nicolson (1972), biological membranes can be considered as a two-dimensional liquid in which all lipid and protein molecules diffuse more or less easily.[3] Although the lipid bilayers that form the basis of the membranes do indeed form two-dimensional liquids by themselves, the plasma membrane also contains a large quantity of proteins, which provide more structure. Examples of such structures are protein-protein complexes, pickets and fences formed by the actin-based cytoskeleton, and potentially lipid rafts.

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Lipid bilayer
Lipid bilayers go through a self assembly process in the formation of membranes. The cell membrane consists primarily of a thin layer of amphipathic phospholipids which spontaneously arrange so that the hydrophobic "tail" regions are shielded from the surrounding polar fluid, causing the more hydrophilic "head" regions to associate with the cytosolic and extracellular faces of the resulting bilayer. This forms a continuous, spherical lipid bilayer. Forces such as Van der Waal, electrostatic, hyrdogen bonds, and noncovalent interactions, are all forces that contribute to the formation of the lipid bilayer. Overall, hydrophobic interactions are the major driving force in the formation of lipid bilayers.

Diagram of the arrangement of amphipathic lipid molecules to form a lipid bilayer. The yellow polar head groups separate the grey hydrophobic tails from the aqueous cytosolic and extracellular environments.

Lipid bilayers have very low permeability for ions and most polar molecules.The arrangement of hydrophilic heads and hydrophobic tails of the lipid bilayer prevent polar solutes (e.g. amino acids, nucleic acids, carbohydrates, proteins, and ions) from diffusing across the membrane, but generally allows for the passive diffusion of hydrophobic molecules. This affords the cell the ability to control the movement of these substances via transmembrane protein complexes such as pores and gates. Flippases and scramblases concentrate phosphatidyl serine, which carries a negative charge, on the inner membrane. Along with NANA, this creates an extra barrier to charged moieties moving through the membrane. Membranes serve diverse functions in eukaryotic and prokaryotic cells. One important role is to regulate the movement of materials into and out of cells. The phospholipid bilayer structure (fluid mosaic model) with specific membrane proteins accounts for the selective permeability of the membrane and passive and active transport mechanisms. In addition, membranes in prokaryotes and in the mitochondria and chloroplasts of eukaryotes facilitate the synthesis of ATP through chemiosmosis.

Membrane polarity
The apical membrane of a polarized cell is the surface of the plasma membrane that faces the lumen. This is particularly evident in epithelial and endothelial cells, but also describes other polarized cells, such as neurons. The basolateral membrane of a polarized cell is the surface of the plasma membrane that forms its basal and lateral surfaces. It faces towards the interstitium, and away from the lumen. "Basolateral membrane" is a compound phrase referring to the terms basal (base) membrane and lateral (side) membrane, Alpha intercalated cell which, especially in epithelial cells, are identical in composition and activity. Proteins (such as ion channels and pumps) are free to move from the basal to the lateral surface of the cell or vice versa in accordance with the fluid mosaic model.

Cell membrane Tight junctions that join epithelial cells near their apical surface prevent the migration of proteins from the basolateral membrane to the apical membrane. The basal and lateral surfaces thus remain roughly equivalent to one another, yet distinct from the apical surface.

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Integral membrane proteins

The cell membrane contains many integral membrane proteins, which pepper the entire surface. These structures, which can be visualized by electron microscopy or fluorescence microscopy, can be found on the inside of the membrane, the outside, or may span the entire membrane. These may include integrins, cadherins, desmosomes, clathrin-coated pits, caveolaes, and different structures involved in cell adhesion. Integral proteins are the most abundant type of protein to span the lipid bilayer. They interact widely with hydrocarbon chains of membrane lipids and can be released by agents that compete for the same nonpolar interactions.

Peripheral membrane proteins

Peripheral proteins are proteins that are bounded to the membrane by electrostatic interactions and hydrogen bonding with the hydrophilic phospholipid heads. Many of these proteins can be found bounded to the surfaces of integral proteins on either the cytoplasimic side of the cell or the extracellular side of the membrane. Some are anchored to the bilayer through covalent bond with a fatty acid.

Membrane skeleton
The cytoskeleton is found underlying the cell membrane in the cytoplasm and provides a scaffolding for membrane proteins to anchor to, as well as forming organelles that extend from the cell. Indeed, cytoskeletal elements interact extensively and intimately with the cell membrane.[4] Anchoring proteins restricts them to a particular cell surface for example, the apical surface of epithelial cells that line the vertebrate gut and limits how far they may diffuse within the bilayer. The cytoskeleton is able to form appendage-like organelles, such as cilia, which are microtubule-based extensions covered by the cell membrane, and filopodia, which are actin-based extensions. These extensions are ensheathed in membrane and project from the surface of the cell in order to sense the external environment and/or make contact with the substrate or other cells. The apical surfaces of epithelial cells are dense with actin-based finger-like projections known as microvilli, which increase cell surface area and thereby increase the absorption rate of nutrients. Localized decoupling of the cytoskeleton and cell membrane results in formation of a bleb.

Composition
Cell membranes contain a variety of biological molecules, notably lipids and proteins. Material is incorporated into the membrane, or deleted from it, by a variety of mechanisms: Fusion of intracellular vesicles with the membrane (exocytosis) not only excretes the contents of the vesicle but also incorporates the vesicle membrane's components into the cell membrane. The membrane may form blebs around extracellular material that pinch off to become vesicles (endocytosis). If a membrane is continuous with a tubular structure made of membrane material, then material from the tube can be drawn into the membrane continuously. Although the concentration of membrane components in the aqueous phase is low (stable membrane components have low solubility in water), there is an exchange of molecules between the lipid and aqueous phases.

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Lipids
The cell membrane consists of three classes of amphipathic lipids: phospholipids, glycolipids, and cholesterols. The amount of each depends upon the type of cell, but in the majority of cases phospholipids are the most abundant.[5] In RBC studies, 30% of the plasma membrane is lipid. The fatty chains in phospholipids and glycolipids usually contain an even number of carbon atoms, typically between 16 and 20. The 16- and 18-carbon fatty acids are the most common. Fatty acids may be saturated or unsaturated, with the configuration of the double bonds nearly always cis. The length and the degree of unsaturation of fatty acid chains have a profound effect on membrane fluidity[6] as unsaturated lipids create a kink, preventing the fatty Examples of the major membrane phospholipids and glycolipids: phosphatidylcholine acids from packing together as tightly, (PtdCho), phosphatidylethanolamine (PtdEtn), phosphatidylinositol (PtdIns), thus decreasing the melting temperature phosphatidylserine (PtdSer). (increasing the fluidity) of the membrane. The ability of some organisms to regulate the fluidity of their cell membranes by altering lipid composition is called homeoviscous adaptation. The entire membrane is held together via non-covalent interaction of hydrophobic tails, however the structure is quite fluid and not fixed rigidly in place. Under physiological conditions phospholipid molecules in the cell membrane are in the liquid crystalline state. It means the lipid molecules are free to diffuse and exhibit rapid lateral diffusion along the layer in which they are present. However, the exchange of phospholipid molecules between intracellular and extracellular leaflets of the bilayer is a very slow process. Lipid rafts and caveolae are examples of cholesterol-enriched microdomains in the cell membrane. In animal cells cholesterol is normally found dispersed in varying degrees throughout cell membranes, in the irregular spaces between the hydrophobic tails of the membrane lipids, where it confers a stiffening and strengthening effect on the membrane.[2]

Phospholipids forming lipid vesicles

Lipid vesicles or lisosomes are circular pockets that are enclosed by a lipid bilayer. These structures are used in laboratories to study the effects of chemicals in cells by delivering these chemicals directly to the cell, as well as getting more insight into cell membrane permeability. Lipid vesicles and liposomes are formed by first suspending a lipid in an aqueous solution then agitating the mixture through sonication, resulting in a vesicle. By measuring the rate of efflux from that of the inside of the vesicle to the ambient solution, allows researcher to better understand membrane permeability. Vesicles can be formed with molecules and ions inside the vesicle by forming the vesicle with the desired molecule or ion present in the solution. Proteins can also be embedded into the membrane through solubilizing the desired proteins in the presence of detergents and attaching them to the phospholipids in which the

Cell membrane liposome is formed. These provide researchers with a tool to examine various membrane protein functions.

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Carbohydrates
Plasma membranes also contain carbohydrates, predominantly glycoproteins, but with some glycolipids (cerebrosides and gangliosides). For the most part, no glycosylation occurs on membranes within the cell; rather generally glycosylation occurs on the extracellular surface of the plasma membrane. The glycocalyx is an important feature in all cells, especially epithelia with microvilli. Recent data suggest the glycocalyx participates in cell adhesion, lymphocyte homing, and many others. The penultimate sugar is galactose and the terminal sugar is sialic acid, as the sugar backbone is modified in the golgi apparatus. Sialic acid carries a negative charge, providing an external barrier to charged particles.

Proteins
Proteins within the membrane are key to the functioning of the overall membrane. These proteins mainly transport chemicals and information across the membrane. Every membrane has a varying degree of protein content. Proteins can be in the form of peripheral or integral.
Type Integral proteins or transmembrane proteins Description Span the membrane and have a hydrophilic cytosolic domain, which interacts with internal molecules, a hydrophobic membrane-spanning domain that anchors it within the cell membrane, and a hydrophilic extracellular domain that interacts with external molecules. The hydrophobic domain consists of one, multiple, or a combination of -helices and sheet protein motifs. Covalently bound to single or multiple lipid molecules; hydrophobically insert into the cell membrane and anchor the protein. The protein itself is not in contact with the membrane. Attached to integral membrane proteins, or associated with peripheral regions of the lipid bilayer. These proteins tend to have only temporary interactions with biological membranes, and, once reacted the molecule, dissociates to carry on its work in the cytoplasm. Examples Ion channels, proton pumps, G protein-coupled receptor

Lipid anchored proteins Peripheral proteins

G proteins

Some enzymes, some hormones

The cell membrane plays host to a large amount of protein that is responsible for its various activities. The amount of protein differs between species and according to function, however the typical amount in a cell membrane is 50%.[6] These proteins are undoubtedly important to a cell: Approximately a third of the genes in yeast code specifically for them, and this number is even higher in multicellular organisms.[5] The cell membrane, being exposed to the outside environment, is an important site of cell-cell communication. As such, a large variety of protein receptors and identification proteins, such as antigens, are present on the surface of the membrane. Functions of membrane proteins can also include cell-cell contact, surface recognition, cytoskeleton contact, signaling, enzymatic activity, or transporting substances across the membrane. Most membrane proteins must be inserted in some way into the membrane. For this to occur, an N-terminus "signal sequence" of amino acids directs proteins to the endoplasmic reticulum, which inserts the proteins into a lipid bilayer. Once inserted, the proteins are then transported to their final destination in vesicles, where the vesicle fuses with the target membrane.

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Variation
The cell membrane has different lipid and protein compositions in distinct types of cells and may have therefore specific names for certain cell types: Sarcolemma in myocytes Oolemma in oocytes Historically, the plasma membrane was also referred to as the plasmalemma.

Permeability
The permeability of a membrane is the ease with which molecules pass through it. These molecules are known as permeant molecules. Permeability depends mainly on the electric charge of the molecule and to a lesser extent the molar mass of the molecule. Due to the cell membrane's hydrophobic nature, electrically neutral and small molecules pass through the membrane easier than charged, large ones. The inability of charged molecules to pass through the cell membrane results in pH parturition of substances throughout the fluid compartments of the body.

References
[1] Kimball's Biology Pages (http:/ / users. rcn. com/ jkimball. ma. ultranet/ BiologyPages/ C/ CellMembranes. html), Cell Membranes [2] Alberts B, Johnson A, Lewis J, et al. (2002). Molecular Biology of the Cell (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. section. 1864) (4th ed.). New York: Garland Science. ISBN0-8153-3218-1. . [3] Singer SJ, Nicolson GL (Feb 1972). "The fluid mosaic model of the structure of cell membranes" (http:/ / www. sciencemag. org/ cgi/ content/ abstract/ 175/ 4023/ 720). Science 175 (4023): 72031. doi:10.1126/science.175.4023.720. PMID4333397. . [4] Doherty GJ and McMahon HT (2008). "Mediation, Modulation and Consequences of Membrane-Cytoskeleton Interactions" (http:/ / arjournals. annualreviews. org/ doi/ abs/ 10. 1146/ annurev. biophys. 37. 032807. 125912). Annual Review of Biophysics 37: 6595. doi:10.1146/annurev.biophys.37.032807.125912. PMID18573073. . [5] Lodish H, Berk A, Zipursky LS, et al. (2004). Molecular Cell Biology (4th ed.). New York: Scientific American Books. ISBN0716731363. [6] Jesse Gray, Shana Groeschler, Tony Le, Zara Gonzalez (2002). "Membrane Structure" (http:/ / www. bio. davidson. edu/ people/ macampbell/ 111/ memb-swf/ membranes. swf) (SWF). Davidson College. . Retrieved 2007-01-11.

External links
Lipids, Membranes and Vesicle Trafficking - The Virtual Library of Biochemistry and Cell Biology (http:// www.biochemweb.org/lipids_membranes.shtml) Cell membrane protein extraction protocol (http://www.westernblotting.org/protocol membrane extraction. htm) Membrane homeostasis, tension regulation, mechanosensitive membrane exchange and membrane traffic (http:// www.phys.unsw.edu.au/~jw/tension.html) 3D structures of proteins associated with plasma membrane of eukaryotic cells (http://opm.phar.umich.edu/ localization.php?localization=Eukaryotic plasma membrane) Lipid composition and proteins of some eukariotic membranes (http://opm.phar.umich.edu/atlas. php?membrane=Eukaryotic plasma membrane) (http://www.etap.org/demo/biology1/instruction3tutor.html)

186

Carbohydrate structure
Carbohydrate
A carbohydrate (pronounced/krbhadret/) is an organic compound with the empirical formula Cm(H2O)n (where m could be different from n); that is, consists only of carbon, hydrogen, and oxygen, with a hydrogen:oxygen atom ratio of 2:1 (as in water). However, there are exceptions to this. One common example would be Lactose is a disaccharide found in milk. It consists of a molecule of D-galactose and a deoxyribose, a component of DNA, which molecule of D-glucose bonded by beta-1-4 glycosidic linkage. It has a formula of C12H22O11. has the empirical formula C5H10O4. Carbohydrates can be viewed as hydrates of carbon, hence their name. Structurally however, it is more accurate to view them as polyhydroxy aldehydes and ketones. The term is most common in biochemistry, where it is a synonym of saccharide. The carbohydrates (saccharides) are divided into four chemical groupings: monosaccharides, disaccharides, oligosaccharides, and polysaccharides. In general, the monosaccharides and disaccharides, which are smaller (lower molecular weight) carbohydrates, are commonly referred to as sugars.[1] The word saccharide comes from the Greek word (skkharon), meaning "sugar". While the scientific nomenclature of carbohydrates is complex, the names of the monosaccharides and disaccharides very often end in the suffix -ose. For example, blood sugar is the monosaccharide glucose, table sugar is the disaccharide sucrose, and milk sugar is the disaccharide lactose (see illustration). Carbohydrates perform numerous roles in living things. Polysaccharides serve for the storage of energy (e.g., starch and glycogen), and as structural components (e.g., cellulose in plants and chitin in arthropods). The 5-carbon monosaccharide ribose is an important component of coenzymes (e.g., ATP, FAD, and NAD) and the backbone of the genetic molecule known as RNA. The related deoxyribose is a component of DNA. Saccharides and their derivatives include many other important biomolecules that play key roles in the immune system, fertilization, preventing pathogenesis, blood clotting, and development.[2] In food science and in many informal contexts, the term carbohydrate often means any food that is particularly rich in the complex carbohydrate starch (such as cereals, bread, and pasta) or simple carbohydrates, such as sugar (found in candy, jams, and desserts).

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Structure
Formerly the name "carbohydrate" was used in chemistry for any compound with the formula Cm (H2O) n. Following this definition, some chemists considered formaldehyde (CH2O) to be the simplest carbohydrate, [3] while others claimed that title for glycolaldehyde.[4] Today the term is generally understood in the biochemistry sense, which excludes compounds with only one or two carbons. Natural saccharides are generally built of simple carbohydrates called monosaccharides with general formula (CH2O)n where n is three or more. A typical monosaccharide has the structure H-(CHOH)x(C=O)-(CHOH)y-H, that is, an aldehyde or ketone with many hydroxyl groups added, usually one on each carbon atom that is not part of the aldehyde or ketone functional group. Examples of monosaccharides are glucose, fructose, and glyceraldehydes. However, some biological substances commonly called "monosaccharides" do not conform to this formula (e.g., uronic acids and deoxy-sugars such as fucose), and there are many chemicals that do conform to this formula but are not considered to be monosaccharides (e.g., formaldehyde CH2O and inositol (CH2O)6).[5] The open-chain form of a monosaccharide often coexists with a closed ring form where the aldehyde/ketone carbonyl group carbon (C=O) and hydroxyl group (-OH) react forming a hemiacetal with a new C-O-C bridge. Monosaccharides can be linked together into what are called polysaccharides (or oligosaccharides) in a large variety of ways. Many carbohydrates contain one or more modified monosaccharide units that have had one or more groups replaced or removed. For example, deoxyribose, a component of DNA, is a modified version of ribose; chitin is composed of repeating units of N-acetyl glucosamine, a nitrogen-containing form of glucose.

Monosaccharides
Monosaccharides are the simplest carbohydrates in that they cannot be hydrolyzed to smaller carbohydrates. They are aldehydes or ketones with two or more hydroxyl groups. The general chemical formula of an unmodified monosaccharide is (CH2O) n, literally a "carbon hydrate." Monosaccharides are important fuel molecules as well as building blocks for nucleic acids. The smallest monosaccharides, for which n = 3, are dihydroxyacetone and D- and L-glyceraldehydes.

Classification of monosaccharides

The and anomers of glucose. Note the position of the hydroxyl group (red or green) on the anomeric carbon relative to the CH2OH group bound to carbon 5: they are either on the opposite sides (), or the same side ().

Monosaccharides are classified according to three different characteristics: the placement of its carbonyl group, the number of carbon atoms it contains, and its chiral handedness. If the carbonyl group is an aldehyde, the monosaccharide is an aldose; if the carbonyl group is a ketone, the monosaccharide is a ketose. Monosaccharides with three carbon atoms are called trioses, those with four are called tetroses, five are called pentoses, six are hexoses, and so on.[6] These two systems of classification are often combined. For example, glucose is an aldohexose (a six-carbon aldehyde), ribose is an aldopentose (a five-carbon aldehyde), and fructose is a ketohexose (a six-carbon ketone).

D-glucose is an aldohexose with the formula (CH2O)6. The red atoms highlight the aldehyde group, and the blue atoms highlight the asymmetric center furthest from the aldehyde; because this -OH is on the right of the Fischer projection, this is a D sugar.

Carbohydrate Each carbon atom bearing a hydroxyl group (-OH), with the exception of the first and last carbons, are asymmetric, making them stereo centers with two possible configurations each (R or S). Because of this asymmetry, a number of isomers may exist for any given monosaccharide formula. The aldohexose D-glucose, for example, has the formula (CH2O) 6, of which all but two of its six carbons atoms are stereogenic, making D-glucose one of 24 = 16 possible stereoisomers. In the case of glyceraldehydes, an aldotriose, there is one pair of possible stereoisomers, which are enantiomers and epimers. 1, 3-dihydroxyacetone, the ketose corresponding to the aldose glyceraldehydes, is a symmetric molecule with no stereo centers). The assignment of D or L is made according to the orientation of the asymmetric carbon furthest from the carbonyl group: in a standard Fischer projection if the hydroxyl group is on the right the molecule is a D sugar, otherwise it is an L sugar. The "D-" and "L-" prefixes should not be confused with "d-" or "l-", which indicate the direction that the sugar rotates plane polarized light. This usage of "d-" and "l-" is no longer followed in carbohydrate chemistry.[7]

188

Ring-straight chain isomerism

The aldehyde or ketone group of a straight-chain monosaccharide will react reversibly with a hydroxyl group on a different carbon atom to form a hemiacetal or hemiketal, forming a heterocyclic ring with an oxygen bridge between two carbon atoms. Rings with five and six atoms are called furanose and pyranose forms, respectively, and exist in equilibrium with the straight-chain form.[8] During the conversion from straight-chain form to the cyclic form, the carbon atom containing the carbonyl oxygen, called the anomeric carbon, becomes a stereogenic center with two possible Glucose can exist in both a straight-chain and ring configurations: The oxygen atom may take a position either above form. or below the plane of the ring. The resulting possible pair of stereoisomers is called anomers. In the anomer, the -OH substituent on the anomeric carbon rests on the opposite side (trans) of the ring from the CH2OH side branch. The alternative form, in which the CH2OH substituent and the anomeric hydroxyl are on the same side (cis) of the plane of the ring, is called the anomer. You can remember that the anomer is cis by the mnemonic, "It's always better to e up". Because the ring and straight-chain forms readily interconvert, both anomers exist in equilibrium.[8] In a Fischer Projection, the anomer is represented with the anomeric hydroxyl group trans to the CH2OH and cis in the anomer.

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Use in living organisms

Monosaccharides are the major source of fuel for metabolism, being used both as an energy source (glucose being the most important in nature) and in biosynthesis. When monosaccharides are not immediately needed by many cells they are often converted to more space efficient forms, often polysaccharides. In many animals, including humans, this storage form is glycogen, especially in liver and muscle cells. In plants, starch is used for the same purpose.

Disaccharides
Two joined monosaccharides are called a disaccharide and these are the simplest polysaccharides. Examples include sucrose and lactose. They are composed of two monosaccharide units bound together by a covalent bond known as a glycosidic linkage formed via a dehydration reaction, resulting in the loss of a hydrogen atom from one monosaccharide and a hydroxyl group from the other. The formula of unmodified disaccharides is C12H22O11. Although there are numerous kinds of disaccharides, a handful of disaccharides are particularly notable.

Sucrose, also known as table sugar, is a common disaccharide. It is composed of two monosaccharides: D-glucose (left) and D-fructose (right).

Sucrose, pictured to the right, is the most abundant disaccharide, and the main form in which carbohydrates are transported in plants. It is composed of one D-glucose molecule and one D-fructose molecule. The systematic name for sucrose, O--D-glucopyranosyl-(12)-D-fructofuranoside, indicates four things: Its monosaccharides: glucose and fructose Their ring types: glucose is a pyranose, and fructose is a furanose How they are linked together: the oxygen on carbon number 1 (C1) of -D-glucose is linked to the C2 of D-fructose. The -oside suffix indicates that the anomeric carbon of both monosaccharides participates in the glycosidic bond. Lactose, a disaccharide composed of one D-galactose molecule and one D-glucose molecule, occurs naturally in mammalian milk. The systematic name for lactose is O--D-galactopyranosyl-(14)-D-glucopyranose. Other notable disaccharides include maltose (two D-glucoses linked -1,4) and cellulobiose (two D-glucoses linked -1,4). disaccharides can be classified into two types.They are reducing and non-reducing disaccahrides if the functional group is present in bonding with another sugar unit it is called a reducing disaccharide or biose.

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Oligosaccharides and polysaccharides

Oligosaccharides and polysaccharides are composed of longer chains of monosaccharide units bound together by glycosidic bonds. The distinction between the two is based upon the number of monosaccharide units present in the chain. Oligosaccharides typically contain between three and ten monosaccharide units, and polysaccharides contain greater than Amylose is a linear polymer of glucose mainly linked with (14) bonds. It can be made ten monosaccharide units. Definitions of several thousands of glucose units. It is one of the two components of starch, the other of how large a carbohydrate must be to being amylopectin. fall into each category vary according to personal opinion. Examples of oligosaccharides include the disaccharides mentioned above, the trisaccharide raffinose and the tetrasaccharide stachyose. Oligosaccharides are found as a common form of protein posttranslational modification. Such posttranslational modifications include the Lewis and ABO oligosaccharides responsible for blood group classifications and so of tissue incompatibilities, the alpha-Gal epitope responsible for hyperacute rejection in xenotransplantation, and O-GlcNAc modifications. Polysaccharides represent an important class of biological polymers. Their function in living organisms is usually either structure- or storage-related. Starch (a polymer of glucose) is used as a storage polysaccharide in plants, being found in the form of both amylose and the branched amylopectin. In animals, the structurally similar glucose polymer is the more densely branched glycogen, sometimes called 'animal starch'. Glycogen's properties allow it to be metabolized more quickly, which suits the active lives of moving animals. Cellulose and chitin are examples of structural polysaccharides. Cellulose is used in the cell walls of plants and other organisms, and is claimed to be the most abundant organic molecule on earth.[9] It has many uses such as a significant role in the paper and textile industries, and is used as a feedstock for the production of rayon (via the viscose process), cellulose acetate, celluloid, and nitrocellulose. Chitin has a similar structure, but has nitrogen-containing side branches, increasing its strength. It is found in arthropod exoskeletons and in the cell walls of some fungi. It also has multiple uses, including surgical threads. Other polysaccharides include callose or laminarin, chrysolaminarin, xylan, arabinoxylan, mannan, fucoidan and galactomannan.

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Nutrition
Foods high in carbohydrate include fruits, sweets, soft drinks, breads, pastas, beans, potatoes, bran, rice, and cereals. Carbohydrates are a common source of energy in living organisms; however, no carbohydrate is an essential nutrient in humans.[10] Carbohydrates are not necessary building blocks of other molecules, and the body can obtain all its energy from protein and fats.[11] [12] The brain fats contain 37.8 kilojoules (9 kilocalories) per gram. In the case of protein, this is somewhat misleading as only some amino acids are usable for fuel. Organisms typically cannot metabolize all types of carbohydrate to yield energy. Glucose is a nearly universal and accessible source of calories. Many organisms also have the ability to metabolize other monosaccharides and Disaccharides, though glucose is preferred. In Escherichia coli, for example, the lac operon will express enzymes for the digestion of lactose when it is present, but if both lactose and Grain products: rich sources of carbohydrates glucose are present the lac operon is repressed, resulting in the glucose being used first (see: Diauxie). Polysaccharides are also common sources of energy. Many organisms can easily break down starches into glucose, however, most organisms cannot metabolize cellulose or other polysaccharides like chitin and arabinoxylans. These carbohydrates types can be metabolized by some bacteria and protists. Ruminants and termites, for example, use microorganisms to process cellulose. Even though these complex carbohydrates are not very digestible, they may comprise important dietary elements for humans. Called dietary fiber, these carbohydrates enhance digestion among other benefits. Based on the effects on risk of heart disease and obesity, the Institute of Medicine recommends that American and Canadian adults get between 4565% of dietary energy from carbohydrates.[13] The Food and Agriculture Organization and World Health Organization jointly recommend that national dietary guidelines set a goal of 5575% of total energy from carbohydrates, but only 10% directly from sugars (their term for simple carbohydrates).[14]

Classification
Historically nutritionists have classified carbohydrates as either simple or complex, however, the exact delineation of these categories is ambiguous. Today, simple carbohydrate typically refers to monosaccharides and disaccharides and complex carbohydrate means polysaccharides (and oligosaccharides). However, the term complex carbohydrate was first used in slightly different context in the U.S. Senate Select Committee on Nutrition and Human Needs publication Dietary Goals for the United States (1977). In this work, complex carbohydrate were defined as "fruit, vegetables and whole-grains".[15] Some nutritionists use complex carbohydrate to refer to any sort of digestible saccharide present in a whole food, where fiber, vitamins and minerals are also found (as opposed to processed carbohydrates, which provide calories but few other nutrients). Some simple carbohydrates (e.g. fructose) are digested very slowly, while some complex carbohydrates (starches), especially if processed, raise blood sugar rapidly. The speed of digestion is determined by a variety of factors including which other nutrients are consumed with the carbohydrate, how the food is prepared, individual differences in metabolism, and the chemistry of the carbohydrate. The USDA's Dietary Guidelines for Americans 2005 dispensed with the simple/complex distinction, instead recommending fiber-rich foods and whole grains.[16] The glycemic index (GI) and glycemic load concepts have been developed to characterize food behavior during human digestion. They rank carbohydrate-rich foods based on the rapidity and magnitude of their effect on blood

Carbohydrate glucose levels. Glycemic index is a measure of how quickly food glucose is absorbed, while glycemic load is a measure of the total absorbable glucose in foods. The insulin index is a similar, more recent classification method that ranks foods based on their effects on blood insulin levels, which are caused by glucose (or starch) and some amino acids in food.

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Carbohydrate chemistry
Carbohydrate chemistry is a large and economically important branch of organic chemistry. Some of the main organic reactions that involve carbohydrates are: Carbohydrate acetalisation Cyanohydrin reaction Lobry-de Bruyn-van Ekenstein transformation Amadori rearrangement Nef reaction Wohl degradation KoenigsKnorr reaction

References
[1] Flitsch, SL & Ulijn, RV (2003). "Sugars tied to the spot." Nature 421: 219220 (http:/ / www. nature. com/ nature/ journal/ v421/ n6920/ pdf/ 421219a. pdf). [2] Maton, Anthea; Jean Hopkins, Charles William McLaughlin, Susan Johnson, Maryanna Quon Warner, David LaHart, Jill D. Wright (1993). Human Biology and Health. Englewood Cliffs, New Jersey, USA: Prentice Hall. pp.5259. ISBN0-13-981176-1. [3] John Merle Coulter, Charler Reid Barnes, Henry Chandler Cowles (1930), A Textbook of Botany for Colleges and Universities (http:/ / books. google. com. br/ books?id=WyZnVpCiTHIC& pg=PA375& dq=simplest+ carbohydrate)" [4] Carl A. Burtis, Edward R. Ashwood, Norbert W. Tietz (2000), Tietz fundamentals of clinical chemistry (http:/ / books. google. com/ books?id=l5hqAAAAMAAJ& q=simplest+ carbohydrate) [5] Matthews, C. E.; K. E. Van Holde; K. G. Ahern (1999) Biochemistry. 3rd edition. Benjamin Cummings. ISBN 0-8053-3066-6 [6] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [7] Pigman, Ward; Horton, D. (1972). "Chapter 1: Stereochemistry of the Monosaccharides". In Pigman and Horton. The Carbohydrates: Chemistry and Biochemistry Vol 1A (2nd ed.). San Diego: Academic Press. pp.167. [8] Pigman, Ward; Anet, E.F.L.J. (1972). "Chapter 4: Mutarotations and Actions of Acids and Bases". In Pigman and Horton. The Carbohydrates: Chemistry and Biochemistry Vol 1A (2nd ed.). San Diego: Academic Press. pp.165194. [9] N.A.Campbell (1996) Biology (4th edition). Benjamin Cummings NY. p.23 ISBN 0-8053-1957-3 [10] http:/ / www. ajcn. org/ content/ 75/ 5/ 951. 2. full [11] Is dietary carbohydrate essential for human nutrition? - Westman 75 (5): 951 - American Journal of Clinical Nutrition (http:/ / www. ajcn. org/ cgi/ content/ full/ 75/ 5/ 951-a) [12] A High-Protein, High-Fat, Carbohydrate-Free Diet Reduces Energy Intake, Hepatic Lipogenesis, and Adiposity in Rats - Pichon et al. 136 (5): 1256 - Journal of Nutrition (http:/ / jn. nutrition. org/ cgi/ reprint/ 136/ 5/ 1256?ijkey=ebf0450b5cf21e8d83dd43f62b5559254694f65f) [13] Food and Nutrition Board (2002/2005). Dietary Reference Intakes for Energy, Carbohydrate, Fiber, Fat, Fatty Acids, Cholesterol, Protein, and Amino Acids (http:/ / newton. nap. edu/ books/ 0309085373/ html). Washington, D.C.: The National Academies Press. Page 769 (http:/ / newton. nap. edu/ books/ 0309085373/ html/ 769. html). ISBN 0-309-08537-3. [14] Joint WHO/FAO expert consultation (2003). Diet, Nutrition and the Prevention of Chronic Diseases (http:/ / www. who. int/ hpr/ NPH/ docs/ who_fao_expert_report. pdf) (PDF). Geneva: World Health Organization. Pages 5556. ISBN 92-4-120916-X. [15] Joint WHO/FAO expert consultation (1998), Carbohydrates in human nutrition, chapter 1 (http:/ / www. fao. org/ docrep/ W8079E/ w8079e07. htm). ISBN 92-5-104114-8. [16] DHHS and USDA, Dietary Guidelines for Americans 2005, Chapter 7 Carbohydrates (http:/ / www. health. gov/ dietaryguidelines/ dga2005/ document/ html/ chapter7. htm)

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External links
Carbohydrates, including interactive models and animations (http://www2.ufp.pt/~pedros/bq/carb_en.htm) (Requires MDL Chime (http://www.mdl.com/products/framework/chime/)) IUPAC-IUBMB Joint Commission on Biochemical Nomenclature (JCBN): Carbohydrate Nomenclature (http:// www.chem.qmw.ac.uk/iupac/2carb/) Carbohydrates detailed (http://www.cem.msu.edu/~reusch/VirtualText/carbhyd.htm) Complex And Simple Carbohydrates (http://evilcyber.com/nutrition/complex-and-simple-carbohydrates/) Explanation of the differences Carbohydrates and Glycosylation - The Virtual Library of Biochemistry and Cell Biology (http://www. biochemweb.org/carbohydrates.shtml) Functional Glycomics Gateway (http://www.functionalglycomics.org/), a collaboration between the Consortium for Functional Glycomics and Nature Publishing Group Wine Carbohydrates (http://www.wineclubwizard.com/wine-carbohydrates.html)

Polysaccharide
Polysaccharides are long carbohydrate molecules, of repeated monomer units joined together by glycosidic bonds. They range in structure from linear to highly branched. Polysaccharides are often quite heterogeneous, containing slight modifications of the repeating unit. Depending on the structure, these macromolecules can have distinct properties from their monosaccharide building blocks. They may be amorphous or even insoluble in water.[1] [2]

3D structure of cellulose, a beta-glucan polysaccharide.

When all the monosaccharides in a polysaccharide are the same type, the polysaccharide is called a homopolysaccharide or homoglycan, but when more than one type of monosaccharide is present they are called heteropolysaccharides or heteroglycans. [3] [4] Examples include storage polysaccharides such as starch and glycogen, and structural polysaccharides such as cellulose and chitin. Polysaccharides have a general formula of Cx(H2O)y where x is usually a large number between 200 and 2500. Considering that the repeating units in the polymer backbone are often six-carbon monosaccharides, the general formula can also be represented as (C6H10O5)n where 40n3000.

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Structure
Natural saccharides are generally built of simple carbohydrates called monosaccharides with general formula (CH2O)n where n is three or more. A typical monosaccharide has the structure H-(CHOH)x(C=O)-(CHOH)y-H, that is, an aldehyde or ketone with many hydroxyl groups added, usually one on each carbon atom that is not part of the aldehyde or ketone functional group. Examples of monosaccharides are glucose, fructose, and glyceraldehyde[5] Polysaccharides are composed of long chains of monosaccharide units bound together by glycosidic bonds. Polysaccharides contain more than ten monosaccharide units. Definitions of how large a carbohydrate must be to fall into the categories polysaccharides or oligosaccharides vary according to personal opinion. Polysaccharides is an important class Amylose is a linear polymer of glucose mainly linked with (14) bonds. It can be made of biological polymers. Their function of several thousands of glucose units. It is one of the two components of starch, the other in living organisms is usually either being amylopectin. structure- or storage-related. Starch (a polymer of glucose) is used as a storage polysaccharide in plants, being found in the form of both amylose and the branched amylopectin. In animals, the structurally similar glucose polymer is the more densely branched glycogen, sometimes called 'animal starch'. Glycogen's properties allow it to be metabolized more quickly, which suits the active lives of moving animals. Cellulose and chitin are examples of structural polysaccharides. Cellulose is used in the cell walls of plants and other organisms, and is claimed to be the most abundant organic molecule on earth.[6] It has many uses such as a significant role in the paper and textile industries, and is used as a feedstock for the production of rayon (via the viscose process), cellulose acetate, celluloid, and nitrocellulose. Chitin has a similar structure, but has nitrogen-containing side branches, increasing its strength. It is found in arthropod exoskeletons and in the cell walls of some fungi. It also has multiple uses, including surgical threads. Polysaccharides also include callose or laminarin, chrysolaminarin, xylan, arabinoxylan, mannan, fucoidan and galactomannan.

Function
Nutrition
Polysaccharides are common sources of energy. Many organisms can easily break down starches into glucose, however, most organisms cannot metabolize cellulose or other polysaccharides like chitin and arabinoxylans. These carbohydrates types can be metabolized by some bacteria and protists. Ruminants and termites, for example, use microorganisms to process cellulose. Even though these complex carbohydrates are not very digestible, they may comprise important dietary elements for humans. Called dietary fiber, these carbohydrates enhance digestion among other benefits. The main action of dietary fiber is to change the nature of the contents of the gastrointestinal tract, and to change how other nutrients and chemicals are absorbed.[7] [8] Soluble fiber binds to bile acids in the small intestine, making them less likely to enter the body; this in turn lowers cholesterol levels in the blood.[9] Soluble fiber also attenuates the absorption of sugar, reduces sugar response after eating, normalizes blood lipid levels and, once fermented in the colon, produces short-chain fatty acids as byproducts with wide-ranging physiological activities (discussion below). Although

Polysaccharide insoluble fiber is associated with reduced diabetes risk, the mechanism by which this occurs is unknown.[10] Not yet formally proposed as an essential macronutrient, dietary fiber is nevertheless regarded as important for the diet, with regulatory authorities in many developed countries recommending increases in fiber intake.[7] [8] [11] [12]

195

Storage polysaccharides
Starches
Starches are glucose polymers in which glucopyranose units are bonded by alpha-linkages. It is made up of a mixture of Amylose (1520%) and Amylopectin (8085%). Amylose consists of a linear chain of several hundred glucose molecules and Amylopectin is a branched molecule made of several thousand glucose units (every chain of 2430 glucose units is one unit of Amylopectin). Starches are insoluble in water. They can be digested by hydrolysis, catalyzed by enzymes called amylases, which can break the alpha-linkages (glycosidic bonds). Humans and other animals have amylases, so they can digest starches. Potato, rice, wheat, and maize are major sources of starch in the human diet. The formations of starches are the ways that plants store glucose.

Glycogen
Glycogen serves as the secondary long-term energy storage in animal and fungal cells, with the primary energy stores being held in adipose tissue. Glycogen is made primarily by the liver and the muscles, but can also be made by glycogenesis within the brain and stomach.[14] Glycogen is the analogue of starch, a glucose polymer in plants, and is sometimes referred to as animal starch, having a similar structure to amylopectin but more extensively branched and compact than starch. Glycogen is a polymer of (14) glycosidic bonds linked, with (16)-linked branches. Glycogen is found in the form of granules in the cytosol/cytoplasm in many cell types, and plays an important role in the glucose cycle. Glycogen forms an energy reserve that can be quickly mobilized to meet a sudden need for glucose, but one that is less compact than the energy reserves of triglycerides (lipids).

Schematic 2-D cross-sectional view of glycogen. A core protein of glycogenin is surrounded by branches of glucose units. The entire globular granule may contain [13] approximately 30,000 glucose units.

In the liver hepatocytes, glycogen can compose up to eight percent of the fresh weight (100120g in an adult) soon after a meal.[15] Only the glycogen stored in the

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liver can be made accessible to other organs. In the muscles, glycogen is found in a low concentration (one to two percent of the muscle mass). However, the amount of glycogen stored in the bodyespecially within the muscles, liver, and red blood cells[16] [17] [18] mostly depends on physical training, basal metabolic rate, and eating habits such as intermittent fasting. Small amounts of glycogen are found in the kidneys, and even smaller amounts in certain glial cells in the brain and white blood cells. The uterus also stores glycogen during pregnancy to nourish the embryo.[19] Glycogen is composed of a branched chain of glucose residues. It is stored in liver and muscles. It is an energy reserve for animals. A view of the atomic structure of a single branched strand of glucose units in a It is the chief form of carbohydrate stored glycogen molecule. in animal body. It is insoluble in water. It turns red when mixed with iodine. It also yields glucose on hydrolysis.

Structural polysaccharides
Arabinoxylans
Arabinoxylans are found in both the primary and secondary cell walls of plants and are the copolymers of two pentose sugars: arabinose and xylose.

Cellulose
The structural component of plants are formed primarily from cellulose. Wood is largely cellulose and lignin, while paper and cotton are nearly pure cellulose. Cellulose is a polymer made with repeated glucose units bonded together by beta-linkages. Humans and many other animals lack an enzyme to break the beta-linkages, so they do not digest cellulose. Certain animals such as termites can digest cellulose, because bacteria possessing the enzyme are present in their gut. Cellulose is insoluble in water. It does not change color when mixed with iodine. On hydrolysis, it yields glucose. It is the most abundant carbohydrate in nature.

Chitin
Chitin is one of many naturally occurring polymers. It forms a structural component of many animals, such as exoskeletons. Over time it is bio-degradable in the natural environment. Its breakdown may be catalyzed by enzymes called chitinases, secreted by microorganisms such as bacteria and fungi, and produced by some plants. Some of these microorganisms have receptors to simple sugars from the decomposition of chitin. If chitin is detected, they then produce enzymes to digest it by cleaving the glycosidic bonds in order to convert it to simple sugars and ammonia.

Polysaccharide Chemically, chitin is closely related to chitosan (a more water-soluble derivative of chitin). It is also closely related to cellulose in that it is a long unbranched chain of glucose derivatives. Both materials contribute structure and strength, protecting the organism.

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Pectins
Pectins are a family of complex polysaccharides that contain 1,4-linked -D-galactosyluronic acid residues. They are present in most primary cell walls and in the non-woody parts of terrestrial plants.

Acidic polysaccharides
Acidic polysaccharides are polysaccharides that contain carboxyl groups, phosphate groups and/or sulfuric ester groups.

Bacterial polysaccharides
Bacterial polysaccharides represent a diverse range of macromolecules that include peptidoglycan, lipopolysaccharides, capsules and exopolysaccharides; compounds whose functions range from structural cell-wall components (e.g., peptidoglycan), and important virulence factors (e.g., Poly-N-acetylglucosamine in S. aureus), to permitting the bacterium to survive in harsh environments (e.g., Pseudomonas aeruginosa in the human lung).[20] Polysaccharide biosynthesis is a tightly regulated, energy-intensive process and understanding the subtle interplay between the regulation and energy conservation, polymer modification and synthesis, and the external ecological functions is a huge area of research. The potential benefits are enormous and should enable for example the development of novel antibacterial strategies (e.g., new antibiotics and vaccines) and the commercial exploitation to develop novel applications.[21] [22]

Bacterial capsular polysaccharides

Pathogenic bacteria commonly produce a thick, mucous-like, layer of polysaccharide. This "capsule" cloaks antigenic proteins on the bacterial surface that would otherwise provoke an immune response and thereby lead to the destruction of the bacteria. Capsular polysaccharides are water soluble, commonly acidic, and have molecular weights on the order of 100-1000 kDa. They are linear and consist of regularly repeating subunits of one to six monosaccharides. There is enormous structural diversity; nearly two hundred different polysaccharides are produced by E. coli alone. Mixtures of capsular polysaccharides, either conjugated or native are used as vaccines. Bacteria and many other microbes, including fungi and algae, often secrete polysaccharides as an evolutionary adaptation to help them adhere to surfaces and to prevent them from drying out. Humans have developed some of these polysaccharides into useful products, including xanthan gum, dextran, welan gum, gellan gum, diutan gum and pullulan. Most of these polysaccharides exhibit interesting and very useful visco-elastic properties when dissolved in water at very low levels.[23] This gives many foods and various liquid consumer products, like lotions, cleaners and paints, for example, a viscous appearance when stationary, but fluidity when the slightest shear is applied, such as when wiped, poured or brushed. This property is referred to as pseudoplasticity, or shear thinning.

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Viscosity of Welan gum [24]

Shear Rate (rpm) 0.3 0.5 1 2 4 5 10 20 50 100 Viscosity (cP) 23330 16000 11000 5500 3250 2900 1700 900 520 310

Aqueous solutions of the polysaccharide alone have a curious behavior when stirred. After stopping, the swirl continues due to momentum, then stops, and then reverses direction briefly. This recoil demonstrates the elastic effect of the polysaccharide chains previously streched in solution, returning to their relaxed state. Cell-surface polysaccharides play diverse roles in bacterial ecology and physiology. They serve as a barrier between the cell wall and the environment, mediate host-pathogen interactions, and form structural components of biofilms. These polysaccharides are synthesized from nucleotide-activated precursors (called nucleotide sugars) and, in most cases, all the enzymes necessary for biosynthesis, assembly and transport of the completed polymer are encoded by genes organized in dedicated clusters within the genome of the organism. Lipopolysaccharide is one of the most important cell-surface polysaccharides, as it plays a key structural role in outer membrane integrity, as well as being an important mediator of host-pathogen interactions. The enzymes that make the A-band (homopolymeric) and B-band (heteropolymeric) O-antigens have been identified and the metabolic pathways defined.[25] The exopolysaccharide alginate is a linear copolymer of -1,4-linked D-mannuronic acid and L-guluronic acid residues, and is responsible for the mucoid phenotype of late-stage cystic fibrosis disease. The pel and psl loci are two recently discovered gene clusters that also encode exopolysaccharides found to be important for biofilm formation. Rhamnolipid is a biosurfactant whose production is tightly regulated at the transcriptional level, but the precise role that it plays in disease is not well understood at present. Protein glycosylation, particularly of pilin and flagellin, is a recent focus of research by several groups and it has been shown to be important for adhesion and invasion during bacterial infection.[26]

References
[1] Varki A, Cummings R, Esko J, Freeze H, Stanley P, Bertozzi C, Hart G, Etzler M (2008). Essentials of glycobiology (http:/ / www. ncbi. nlm. nih. gov/ bookshelf/ br. fcgi?book=glyco2). Cold Spring Harbor Laboratory Press; 2nd edition. ISBN0-87969-770-9. . [2] Varki A, Cummings R, Esko J, Jessica Freeze, Hart G, Marth J (1999). Essentials of glycobiology (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=glyco. TOC& depth=2). Cold Spring Harbor Laboratory Press. ISBN0-87969-560-9. . [3] IUPAC, Compendium of Chemical Terminology, 2nd ed. (the "Gold Book") (1997). Online corrected version: (2006) " homopolysaccharide (homoglycan) (http:/ / goldbook. iupac. org/ H02856. html)". [4] IUPAC, Compendium of Chemical Terminology, 2nd ed. (the "Gold Book") (1997). Online corrected version: (2006) " heteropolysaccharide (heteroglycan) (http:/ / goldbook. iupac. org/ H02812. html)". [5] Matthews, C. E.; K. E. Van Holde; K. G. Ahern (1999) Biochemistry. 3rd edition. Benjamin Cummings. ISBN 0-8053-3066-6 [6] N.A.Campbell (1996) Biology (4th edition). Benjamin Cummings NY. p.23 ISBN 0-8053-1957-3 [7] "Dietary Reference Intakes for Energy, Carbohydrate, fiber, Fat, Fatty Acids, Cholesterol, Protein, and Amino Acids (Macronutrients) (2005), Chapter 7: Dietary, Functional and Total fiber." (http:/ / www. nal. usda. gov/ fnic/ DRI/ / DRI_Energy/ 339-421. pdf). US Department of Agriculture, National Agricultural Library and National Academy of Sciences, Institute of Medicine, Food and Nutrition Board. .

Polysaccharide
[8] Eastwood M, Kritchevsky D (2005). "Dietary fiber: how did we get where we are?". Annu Rev Nutr 25: 18. doi:10.1146/annurev.nutr.25.121304.131658. PMID16011456. [9] Anderson JW, Baird P, Davis RH et al. (2009). "Health benefits of dietary fiber". Nutr Rev 67 (4): 188205. doi:10.1111/j.1753-4887.2009.00189.x. PMID19335713. [10] Weickert MO, Pfeiffer AF (2008). "Metabolic effects of dietary fiber consumption and prevention of diabetes". J Nutr 138 (3): 43942. PMID18287346. [11] "Dietary reference values for carbohydrates and dietary fiber" (http:/ / www. efsa. europa. eu/ EFSA/ DocumentSet/ nda_op_drv_carbohydrates_draft_en_released for consultation,0. pdf?ssbinary=true). European Food Safety Authority. . [12] Jones PJ, Varady KA (2008). "Are functional foods redefining nutritional requirements?" (http:/ / article. pubs. nrc-cnrc. gc. ca/ ppv/ RPViewDoc?issn=1715-5312& volume=33& issue=1& startPage=118) (PDF). Appl Physiol Nutr Metab 33 (1): 11823. doi:10.1139/H07-134. PMID18347661. . [13] Page 12 in: (http:/ / books. google. dk/ books?id=SRptlOx7yj4C& printsec=frontcover& hl=en) Exercise physiology: energy, nutrition, and human performance By William D. McArdle, Frank I. Katch, Victor L. Katch Edition: 6, illustrated Published by Lippincott Williams & Wilkins, 2006 ISBN 0781749905, 9780781749909, 1068 pages [14] Anatomy and Physiology. Saladin, Kenneth S. McGraw-Hill, 2007. [15] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [16] Moses SW, Bashan N, Gutman A (December 1972). "Glycogen metabolism in the normal red blood cell" (http:/ / www. bloodjournal. org/ cgi/ pmidlookup?view=long& pmid=5083874). Blood 40 (6): 83643. PMID5083874. . [17] http:/ / jeb. biologists. org/ cgi/ reprint/ 129/ 1/ 141. pdf [18] Miwa I, Suzuki S (November 2002). "An improved quantitative assay of glycogen in erythrocytes". Annals of Clinical Biochemistry 39 (Pt 6): 6123. doi:10.1258/000456302760413432. PMID12564847. [19] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [20] Sutherland, I. W. (2002). Vandamme, E. J., Ed.. ed. Polysaccharides from Microorganisms, Plants and Animals, in: Biopolymers, Volume 5, Polysaccharides I: Polysaccharides from Prokaryotes. Weiheim Wiley VCH. pp.119. ISBN978-3-527-30226-0. [21] Ullrich M (editor) (2009). Bacterial Polysaccharides: Current Innovations and Future Trends. Caister Academic Press. ISBN978-1-904455-45-5. [22] Rehm BHA (editor). (2009). Microbial Production of Biopolymers and Polymer Precursors: Applications and Perspectives. Caister Academic Press. ISBN978-1-904455-36-3. [23] Viscosity of Welan Gum vs. Concentration in Water. http:/ / www. xydatasource. com/ xy-showdatasetpage. php?datasetcode=345115& dsid=80 [24] http:/ / www. xydatasource. com/ xy-showdatasetpage. php?datasetcode=45615& dsid=76& searchtext=polysaccharide [25] Guo H, Yi W, Song JK, Wang PG (2008). "Current understanding on biosynthesis of microbial polysaccharides". Curr Top Med Chem 8 (2): 14151. doi:10.2174/156802608783378873. PMID18289083. [26] Cornelis P (editor). (2008). Pseudomonas: Genomics and Molecular Biology (http:/ / www. horizonpress. com/ pseudo) (1st ed.). Caister Academic Press. ISBN978-1-904455-19-6. . .

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External links
Polysaccharide Structure (http://employees.csbsju.edu/hjakubowski/classes/ch331/cho/complexoligosacch. htm) Applications and commercial sources of polysaccharides (http://www.polysaccharidecenter.com) European Polysaccharide Network of Excellence (http://www.epnoe.eu)

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Intermediary metabolism

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Metabolism
Overview of metabolism
Metabolism (from Greek: "metabol", "change" or Greek: metabolismos, "outthrow") is the set of chemical reactions that happen in the cells of living organisms to sustain life. These processes allow organisms to grow and reproduce, maintain their structures, and respond to their environments. Metabolism is usually divided into two categories. Catabolism breaks down organic matter, for example to harvest energy in cellular respiration. Anabolism uses energy to construct components of cells such as proteins and nucleic acids.

The chemical reactions of metabolism are organized into metabolic pathways, in which one chemical is transformed through a series of steps into another chemical, by a sequence of enzymes. Enzymes are crucial to metabolism because they allow organisms to drive desirable reactions that require energy and will not occur by themselves, by coupling them to spontaneous reactions that release energy. As enzymes act as catalysts they allow these reactions to proceed quickly and efficiently. Enzymes also allow the regulation of metabolic pathways in response to changes in the cell's environment or signals from other cells. The metabolism of an organism determines which substances it will find nutritious and which it will find poisonous. For example, some prokaryotes use hydrogen sulfide as a nutrient, yet this gas is poisonous to animals.[1] The speed of metabolism, the metabolic rate, influences how much food an organism will require, and also affects how it is able to obtain that food. A striking feature of metabolism is the similarity of the basic metabolic pathways and components between even vastly different species.[2] For example, the set of carboxylic acids that are best known as the intermediates in the citric acid cycle are present in all known organisms, being found in species as diverse as the unicellular bacteria Escherichia coli and huge multicellular organisms like elephants.[3] These striking similarities in metabolic pathways are likely due to their early appearance in evolutionary history, and being retained because of their efficacy.[4] [5]

Structure of adenosine triphosphate, a central intermediate in energy metabolism

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Key biochemicals
Further information: Biomolecule, cell (biology) and biochemistry Most of the structures that make up animals, plants and microbes are made from three basic classes of molecule: amino acids, carbohydrates and lipids (often called fats). As these molecules are vital for life, metabolic reactions either focus on making these molecules during the construction of cells and tissues, or breaking them down and using them as a source of energy, in the digestion and use of food. Many important biochemicals can be joined together to make polymers such as DNA and proteins. These macromolecules are essential.

Structure of a triacylglycerol lipid

Type of molecule Amino acids Carbohydrates Nucleic acids

Name of monomer forms Amino acids Monosaccharides Nucleotides

Name of polymer forms

Examples of polymer forms

Proteins (also called polypeptides) Fibrous proteins and globular proteins Polysaccharides Polynucleotides Starch, glycogen and cellulose DNA and RNA

Amino acids and proteins

Proteins are made of amino acids arranged in a linear chain and joined together by peptide bonds. Many proteins are the enzymes that catalyze the chemical reactions in metabolism. Other proteins have structural or mechanical functions, such as the proteins that form the cytoskeleton, a system of scaffolding that maintains the cell shape.[6] Proteins are also important in cell signaling, immune responses, cell adhesion, active transport across membranes, and the cell cycle.[7]

Lipids
Lipids are the most diverse group of biochemicals. Their main structural uses are as part of biological membranes such as the cell membrane, or as a source of energy.[7] Lipids are usually defined as hydrophobic or amphipathic biological molecules that will dissolve in organic solvents such as benzene or chloroform.[8] The fats are a large group of compounds that contain fatty acids and glycerol; a glycerol molecule attached to three fatty acid esters is a triacylglyceride.[9] Several variations on this basic structure exist, including alternate backbones such as sphingosine in the sphingolipids, and hydrophilic groups such as phosphate in phospholipids. Steroids such as cholesterol are another major class of lipids that are made in cells.[10]

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Carbohydrates
Carbohydrates are aldehydes or ketones with many hydroxyl groups that can exist as straight chains or rings. Carbohydrates are the most abundant biological molecules, and fill numerous roles, such as the storage and transport of energy (starch, glycogen) and structural components (cellulose in plants, chitin in animals).[7] The basic carbohydrate units are called monosaccharides and include galactose, fructose, and most importantly glucose. Monosaccharides can be linked together to form polysaccharides in almost limitless ways.[11]

Nucleotides

Glucose can exist in both a straight-chain and ring form.

The two nucleic acids, DNA and RNA are polymers of nucleotides, each nucleotide comprising a phosphate group, a ribose sugar group, and a nitrogenous base. Nucleic acids are critical for the storage and use of genetic information, through the processes of transcription and protein biosynthesis.[7] This information is protected by DNA repair mechanisms and propagated through DNA replication. Many viruses have an RNA genome, for example HIV, which uses reverse transcription to create a DNA template from its viral RNA genome.[12] RNA in ribozymes such as spliceosomes and ribosomes is similar to enzymes as it can catalyze chemical reactions. Individual nucleosides are made by attaching a nucleobase to a ribose sugar. These bases are heterocyclic rings containing nitrogen, classified as purines or pyrimidines. Nucleotides also act as coenzymes in metabolic group transfer reactions.[13]

Coenzymes
Further information: Coenzyme Metabolism involves a vast array of chemical reactions, but most fall under a few basic types of reactions that involve the transfer of functional groups.[14] This common chemistry allows cells to use a small set of metabolic intermediates to carry Structure of the coenzyme acetyl-CoA.The transferable acetyl group is bonded to chemical groups between different [13] the sulfur atom at the extreme left. reactions. These group-transfer intermediates are called coenzymes. Each class of group-transfer reaction is carried out by a particular coenzyme, which is the substrate for a set of enzymes that produce it, and a set of enzymes that consume it. These coenzymes are therefore continuously being made, consumed and then recycled.[15] One central coenzyme is adenosine triphosphate (ATP), the universal energy currency of cells. This nucleotide is used to transfer chemical energy between different chemical reactions. There is only a small amount of ATP in cells, but as it is continuously regenerated, the human body can use about its own weight in ATP per day.[15] ATP acts as a bridge between catabolism and anabolism, with catabolic reactions generating ATP and anabolic reactions consuming it. It also serves as a carrier of phosphate groups in phosphorylation reactions.

Overview of metabolism A vitamin is an organic compound needed in small quantities that cannot be made in the cells. In human nutrition, most vitamins function as coenzymes after modification; for example, all water-soluble vitamins are phosphorylated or are coupled to nucleotides when they are used in cells.[16] Nicotinamide adenine dinucleotide (NADH), a derivative of vitamin B3 (niacin), is an important coenzyme that acts as a hydrogen acceptor. Hundreds of separate types of dehydrogenases remove electrons from their substrates and reduce NAD+ into NADH. This reduced form of the coenzyme is then a substrate for any of the reductases in the cell that need to reduce their substrates.[17] Nicotinamide adenine dinucleotide exists in two related forms in the cell, NADH and NADPH. The NAD+/NADH form is more important in catabolic reactions, while NADP+/NADPH is used in anabolic reactions.

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Minerals and cofactors

Further information: Metal Ions in Life Sciences, Metal metabolism, and bioinorganic chemistry Inorganic elements play critical roles in metabolism; some are abundant (e.g. sodium and potassium) while others function at minute concentrations. About 99% of a mammal's mass is made up of the elements carbon, nitrogen, calcium, sodium, chlorine, potassium, hydrogen, phosphorus, oxygen and sulfur.[18] Organic compounds (proteins, lipids and carbohydrates) contain the majority of the carbon and nitrogen; most of the oxygen and hydrogen is present as water.[18] The abundant inorganic elements act as ionic electrolytes. The most important ions Structure of hemoglobin. The protein subunits are in red and blue, and the [1] iron-containing heme groups in green. From PDB 1GZX . are sodium, potassium, calcium, magnesium, chloride, phosphate and the organic ion bicarbonate. The maintenance of precise gradients across cell membranes maintains osmotic pressure and pH.[19] Ions are also critical for nerve and muscle function, as action potentials in these tissues are produced by the exchange of electrolytes between the extracellular fluid and the cytosol.[20] Electrolytes enter and leave cells through proteins in the cell membrane called ion channels. For example, muscle contraction depends upon the movement of calcium, sodium and potassium through ion channels in the cell membrane and T-tubules.[21] Transition metals are usually present as trace elements in organisms, with zinc and iron being most abundant.[22] [23] These metals are used in some proteins as cofactors and are essential for the activity of enzymes such as catalase and oxygen-carrier proteins such as hemoglobin.[24] Metal cofactors are bound tightly to specific sites in proteins; although enzyme cofactors can be modified during catalysis, they always return to their original state by the end of the reaction catalyzed. Metal micronutrients are taken up into organisms by specific transporters and bind to storage proteins such as ferritin or metallothionein when not being used.[25] [26]

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Catabolism
Further information: Catabolism Catabolism is the set of metabolic processes that break down large molecules. These include breaking down and oxidizing food molecules. The purpose of the catabolic reactions is to provide the energy and components needed by anabolic reactions. The exact nature of these catabolic reactions differ from organism to organism and organisms can be classified based on their sources of energy and carbon (their primary nutritional groups), as shown in the table below. Organic molecules are used as a source of energy by organotrophs, while lithotrophs use inorganic substrates and phototrophs capture sunlight as chemical energy. However, all these different forms of metabolism depend on redox reactions that involve the transfer of electrons from reduced donor molecules such as organic molecules, water, ammonia, hydrogen sulfide or ferrous ions to acceptor molecules such as oxygen, nitrate or sulfate.[27] In animals these reactions involve complex organic molecules being broken down to simpler molecules, such as carbon dioxide and water. In photosynthetic organisms such as plants and cyanobacteria, these electron-transfer reactions do not release energy, but are used as a way of storing energy absorbed from sunlight.[7]

Classification of organisms based on their metabolism

Energy source sunlight photo-troph

Preformed molecules chemoElectron donor organic compound inorganic compound Carbon source organic compound inorganic compound organolithoheteroauto-

The most common set of catabolic reactions in animals can be separated into three main stages. In the first, large organic molecules such as proteins, polysaccharides or lipids are digested into their smaller components outside cells. Next, these smaller molecules are taken up by cells and converted to yet smaller molecules, usually acetyl coenzyme A (acetyl-CoA), which releases some energy. Finally, the acetyl group on the CoA is oxidised to water and carbon dioxide in the citric acid cycle and electron transport chain, releasing the energy that is stored by reducing the coenzyme nicotinamide adenine dinucleotide (NAD+) into NADH.

Digestion
Further information: Digestion and gastrointestinal tract Macromolecules such as starch, cellulose or proteins cannot be rapidly taken up by cells and need to be broken into their smaller units before they can be used in cell metabolism. Several common classes of enzymes digest these polymers. These digestive enzymes include proteases that digest proteins into amino acids, as well as glycoside hydrolases that digest polysaccharides into monosaccharides. Microbes simply secrete digestive enzymes into their surroundings,[28] [29] while animals only secrete these enzymes from specialized cells in their guts.[30] The amino acids or sugars released by these extracellular enzymes are then pumped into cells by specific active transport proteins.[31] [32]

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Energy from organic compounds

Further information: Cellular respiration, fermentation, carbohydrate catabolism, fat catabolism and protein catabolism Carbohydrate catabolism is the breakdown of carbohydrates into smaller units. Carbohydrates are usually taken into cells once they have been digested into monosaccharides.[33] Once inside, the major route of breakdown is glycolysis, where sugars such as glucose and fructose are converted into pyruvate and some ATP is generated.[34] Pyruvate is an intermediate in several metabolic pathways, but the majority is converted to acetyl-CoA and fed into the citric acid cycle. Although some more ATP is generated in the citric acid cycle, the most A simplified outline of the catabolism of proteins, carbohydrates and fats important product is NADH, which is made from NAD+ as the acetyl-CoA is oxidized. This oxidation releases carbon dioxide as a waste product. In anaerobic conditions, glycolysis produces lactate, through the enzyme lactate dehydrogenase re-oxidizing NADH to NAD+ for re-use in glycolysis. An alternative route for glucose breakdown is the pentose phosphate pathway, which reduces the coenzyme NADPH and produces pentose sugars such as ribose, the sugar component of nucleic acids. Fats are catabolised by hydrolysis to free fatty acids and glycerol. The glycerol enters glycolysis and the fatty acids are broken down by beta oxidation to release acetyl-CoA, which then is fed into the citric acid cycle. Fatty acids release more energy upon oxidation than carbohydrates because carbohydrates contain more oxygen in their structures. Amino acids are either used to synthesize proteins and other biomolecules, or oxidized to urea and carbon dioxide as a source of energy.[35] The oxidation pathway starts with the removal of the amino group by a transaminase. The amino group is fed into the urea cycle, leaving a deaminated carbon skeleton in the form of a keto acid. Several of these keto acids are intermediates in the citric acid cycle, for example the deamination of glutamate forms -ketoglutarate.[36] The glucogenic amino acids can also be converted into glucose, through gluconeogenesis (discussed below).[37]

Energy transformations
Oxidative phosphorylation
Further information: Oxidative phosphorylation, chemiosmosis and mitochondrion In oxidative phosphorylation, the electrons removed from organic molecules in areas such as the protagon acid cycle are transferred to oxygen and the energy released is used to make ATP. This is done in eukaryotes by a series of proteins in the membranes of mitochondria called the electron transport chain. In prokaryotes, these proteins are found in the cell's inner membrane.[38] These proteins use the energy released from passing electrons from reduced molecules like NADH onto oxygen to pump protons across a membrane.[39] Pumping protons out of the mitochondria creates a proton concentration difference across the membrane and generates an electrochemical gradient.[40] This force drives protons back into the mitochondrion through the base of

Overview of metabolism an enzyme called ATP synthase. The flow of protons makes the stalk subunit rotate, causing the active site of the synthase domain to change shape and phosphorylate adenosine diphosphate turning it into ATP.[15]

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Energy from inorganic compounds

Further information: Microbial metabolism and nitrogen cycle Chemolithotrophy is a type of metabolism found in prokaryotes where energy is obtained from the oxidation of inorganic compounds. These organisms can use hydrogen,[41] reduced sulfur compounds (such as sulfide, hydrogen sulfide and thiosulfate),[1] ferrous iron (FeII)[42] or ammonia[43] as sources of reducing power and they gain energy from the oxidation of these compounds with electron acceptors such as oxygen or nitrite.[44] These microbial processes are important in global biogeochemical cycles such as acetogenesis, nitrification and denitrification and are critical for soil fertility.[45] [46]

Energy from light

Further information: Phototroph, photophosphorylation, chloroplast The energy in sunlight is captured by plants, cyanobacteria, purple bacteria, green sulfur bacteria and some protists. This process is often coupled to the conversion of carbon dioxide into organic compounds, as part of photosynthesis, which is discussed below. The energy capture and carbon fixation systems can however operate separately in prokaryotes, as purple bacteria and green sulfur bacteria can use sunlight as a source of energy, while switching between carbon fixation and the fermentation of organic compounds.[47] [48] In many organisms the capture of solar energy is similar in principle to oxidative phosphorylation, as it involves energy being stored as a proton concentration gradient and this proton motive force then driving ATP synthesis.[15] The electrons needed to drive this electron transport chain come from light-gathering proteins called photosynthetic reaction centres or rhodopsins. Reaction centers are classed into two types depending on the type of photosynthetic pigment present, with most photosynthetic bacteria only having one type, while plants and cyanobacteria have two.[49] In plants, algae, and cyanobacteria, photosystem II uses light energy to remove electrons from water, releasing oxygen as a waste product. The electrons then flow to the cytochrome b6f complex, which uses their energy to pump protons across the thylakoid membrane in the chloroplast.[7] These protons move back through the membrane as they drive the ATP synthase, as before. The electrons then flow through photosystem I and can then either be used to reduce the coenzyme NADP+, for use in the Calvin cycle which is discussed below, or recycled for further ATP generation.[50]

Anabolism
Further information: Anabolism Anabolism is the set of constructive metabolic processes where the energy released by catabolism is used to synthesize complex molecules. In general, the complex molecules that make up cellular structures are constructed step-by-step from small and simple precursors. Anabolism involves three basic stages. Firstly, the production of precursors such as amino acids, monosaccharides, isoprenoids and nucleotides, secondly, their activation into reactive forms using energy from ATP, and thirdly, the assembly of these precursors into complex molecules such as proteins, polysaccharides, lipids and nucleic acids. Organisms differ in how many of the molecules in their cells they can construct for themselves. Autotrophs such as plants can construct the complex organic molecules in cells such as polysaccharides and proteins from simple molecules like carbon dioxide and water. Heterotrophs, on the other hand, require a source of more complex substances, such as monosaccharides and amino acids, to produce these complex molecules. Organisms can be further classified by ultimate source of their energy: photoautotrophs and photoheterotrophs obtain energy from

Overview of metabolism light, whereas chemoautotrophs and chemoheterotrophs obtain energy from inorganic oxidation reactions.

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Carbon fixation
Further information: Photosynthesis, carbon fixation and chemosynthesis Photosynthesis is the synthesis of carbohydrates from sunlight and carbon dioxide (CO2). In plants, cyanobacteria and algae, oxygenic photosynthesis splits water, with oxygen produced as a waste product. This process uses the ATP and NADPH produced by the photosynthetic reaction centres, as described above, to convert CO2 into glycerate 3-phosphate, which can then be converted into glucose. This carbon-fixation reaction is carried out by the enzyme RuBisCO as part of the Calvin Benson cycle.[51] Three types of photosynthesis occur in plants, C3 carbon fixation, C4 carbon fixation and CAM photosynthesis. These differ by the route that carbon dioxide takes to the Calvin cycle, with C3 plants fixing CO2 directly, while C4 and CAM photosynthesis incorporate the CO2 into other compounds first, as adaptations to deal with intense sunlight and dry conditions.[52]

Plant cells (bounded by purple walls) filled with chloroplasts (green), which are the site of photosynthesis

In photosynthetic prokaryotes the mechanisms of carbon fixation are more diverse. Here, carbon dioxide can be fixed by the Calvin Benson cycle, a reversed citric acid cycle,[53] or the carboxylation of acetyl-CoA.[54] [55] Prokaryotic chemoautotrophs also fix CO2 through the Calvin Benson cycle, but use energy from inorganic compounds to drive the reaction.[56]

Carbohydrates and glycans

Further information: Gluconeogenesis, glyoxylate cycle, glycogenesis and glycosylation In carbohydrate anabolism, simple organic acids can be converted into monosaccharides such as glucose and then used to assemble polysaccharides such as starch. The generation of glucose from compounds like pyruvate, lactate, glycerol, glycerate 3-phosphate and amino acids is called gluconeogenesis. Gluconeogenesis converts pyruvate to glucose-6-phosphate through a series of intermediates, many of which are shared with glycolysis.[34] However, this pathway is not simply glycolysis run in reverse, as several steps are catalyzed by non-glycolytic enzymes. This is important as it allows the formation and breakdown of glucose to be regulated separately and prevents both pathways from running simultaneously in a futile cycle.[57] [58] Although fat is a common way of storing energy, in vertebrates such as humans the fatty acids in these stores cannot be converted to glucose through gluconeogenesis as these organisms cannot convert acetyl-CoA into pyruvate; plants do, but animals do not, have the necessary enzymatic machinery.[59] As a result, after long-term starvation, vertebrates need to produce ketone bodies from fatty acids to replace glucose in tissues such as the brain that cannot metabolize fatty acids.[60] In other organisms such as plants and bacteria, this metabolic problem is solved using the glyoxylate cycle, which bypasses the decarboxylation step in the citric acid cycle and allows the transformation of acetyl-CoA to oxaloacetate, where it can be used for the production of glucose.[59] [61] Polysaccharides and glycans are made by the sequential addition of monosaccharides by glycosyltransferase from a reactive sugar-phosphate donor such as uridine diphosphate glucose (UDP-glucose) to an acceptor hydroxyl group on the growing polysaccharide. As any of the hydroxyl groups on the ring of the substrate can be acceptors, the polysaccharides produced can have straight or branched structures.[62] The polysaccharides produced can have structural or metabolic functions themselves, or be transferred to lipids and proteins by enzymes called oligosaccharyltransferases.[63] [64]

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Fatty acids, isoprenoids and steroids

Further information: Fatty acid synthesis, steroid metabolism Fatty acids are made by fatty acid synthases that polymerize and then reduce acetyl-CoA units. The acyl chains in the fatty acids are extended by a cycle of reactions that add the actyl group, reduce it to an alcohol, dehydrate it to an alkene group and then reduce it again to an alkane group. The enzymes of fatty acid biosynthesis are divided into two groups, in animals and fungi all these fatty acid synthase reactions are carried out by a single multifunctional type I protein,[65] while in plant plastids and bacteria separate type II enzymes perform each step in the pathway.[66] [67] Terpenes and isoprenoids are a large class of lipids that include the carotenoids and form the largest class of plant natural products.[68] These compounds are made by the assembly and modification of isoprene units Simplified version of the steroid synthesis pathway with the intermediates isopentenyl donated from the reactive precursors pyrophosphate (IPP), dimethylallyl pyrophosphate (DMAPP), geranyl pyrophosphate isopentenyl pyrophosphate and (GPP) and squalene shown. Some intermediates are omitted for clarity. dimethylallyl pyrophosphate.[69] These precursors can be made in different ways. In animals and archaea, the mevalonate pathway produces these compounds from acetyl-CoA,[70] while in plants and bacteria the non-mevalonate pathway uses pyruvate and glyceraldehyde 3-phosphate as substrates.[69] [71] One important reaction that uses these activated isoprene donors is steroid biosynthesis. Here, the isoprene units are joined together to make squalene and then folded up and formed into a set of rings to make lanosterol.[72] Lanosterol can then be converted into other steroids such as cholesterol and ergosterol.[72] [73]

Proteins
Further information: Protein biosynthesis, amino acid synthesis Organisms vary in their ability to synthesize the 20 common amino acids. Most bacteria and plants can synthesize all twenty, but mammals can synthesize only eleven nonessential amino acids.[7] Thus, nine essential amino acids must be obtained from food. All amino acids are synthesized from intermediates in glycolysis, the citric acid cycle, or the pentose phosphate pathway. Nitrogen is provided by glutamate and glutamine. Amino acid synthesis depends on the formation of the appropriate alpha-keto acid, which is then transaminated to form an amino acid.[74] Amino acids are made into proteins by being joined together in a chain by peptide bonds. Each different protein has a unique sequence of amino acid residues: this is its primary structure. Just as the letters of the alphabet can be combined to form an almost endless variety of words, amino acids can be linked in varying sequences to form a huge variety of proteins. Proteins are made from amino acids that have been activated by attachment to a transfer RNA

Overview of metabolism molecule through an ester bond. This aminoacyl-tRNA precursor is produced in an ATP-dependent reaction carried out by an aminoacyl tRNA synthetase.[75] This aminoacyl-tRNA is then a substrate for the ribosome, which joins the amino acid onto the elongating protein chain, using the sequence information in a messenger RNA.[76]

210

Nucleotide synthesis and salvage

Further information: Nucleotide salvage, pyrimidine biosynthesis, and purine metabolism Nucleotides are made from amino acids, carbon dioxide and formic acid in pathways that require large amounts of metabolic energy.[77] Consequently, most organisms have efficient systems to salvage preformed nucleotides.[77] [78] Purines are synthesized as nucleosides (bases attached to ribose). Both adenine and guanine are made from the precursor nucleoside inosine monophosphate, which is synthesized using atoms from the amino acids glycine, glutamine, and aspartic acid, as well as formate transferred from the coenzyme tetrahydrofolate. Pyrimidines, on the other hand, are synthesized from the base orotate, which is formed from glutamine and aspartate.[79]

Xenobiotics and redox metabolism

Further information: Xenobiotic metabolism, drug metabolism, Alcohol metabolism and antioxidants All organisms are constantly exposed to compounds that they cannot use as foods and would be harmful if they accumulated in cells, as they have no metabolic function. These potentially damaging compounds are called xenobiotics.[80] Xenobiotics such as synthetic drugs, natural poisons and antibiotics are detoxified by a set of xenobiotic-metabolizing enzymes. In humans, these include cytochrome P450 oxidases,[81] UDP-glucuronosyltransferases,[82] and glutathione S-transferases.[83] This system of enzymes acts in three stages to firstly oxidize the xenobiotic (phase I) and then conjugate water-soluble groups onto the molecule (phase II). The modified water-soluble xenobiotic can then be pumped out of cells and in multicellular organisms may be further metabolized before being excreted (phase III). In ecology, these reactions are particularly important in microbial biodegradation of pollutants and the bioremediation of contaminated land and oil spills.[84] Many of these microbial reactions are shared with multicellular organisms, but due to the incredible diversity of types of microbes these organisms are able to deal with a far wider range of xenobiotics than multicellular organisms, and can degrade even persistent organic pollutants such as organochloride compounds.[85] A related problem for aerobic organisms is oxidative stress.[86] Here, processes including oxidative phosphorylation and the formation of disulfide bonds during protein folding produce reactive oxygen species such as hydrogen peroxide.[87] These damaging oxidants are removed by antioxidant metabolites such as glutathione and enzymes such as catalases and peroxidases.[88] [89]

Thermodynamics of living organisms

Further information: Biological thermodynamics Living organisms must obey the laws of thermodynamics, which describe the transfer of heat and work. The second law of thermodynamics states that in any closed system, the amount of entropy (disorder) will tend to increase. Although living organisms' amazing complexity appears to contradict this law, life is possible as all organisms are open systems that exchange matter and energy with their surroundings. Thus living systems are not in equilibrium, but instead are dissipative systems that maintain their state of high complexity by causing a larger increase in the entropy of their environments.[90] The metabolism of a cell achieves this by coupling the spontaneous processes of catabolism to the non-spontaneous processes of anabolism. In thermodynamic terms, metabolism maintains order by creating disorder.[91]

Overview of metabolism

211

Regulation and control

Further information: Metabolic pathway, metabolic control analysis, hormone, regulatory enzymes, and cell signaling As the environments of most organisms are constantly changing, the reactions of metabolism must be finely regulated to maintain a constant set of conditions within cells, a condition called homeostasis.[92] [93] Metabolic regulation also allows organisms to respond to signals and interact actively with their environments.[94] Two closely linked concepts are important for understanding how metabolic pathways are controlled. Firstly, the regulation of an enzyme in a pathway is how its activity is increased and decreased in response to signals. Secondly, the control exerted by this enzyme is the effect that these changes in its activity have on the overall rate of the pathway (the flux through the pathway).[95] For example, an enzyme may show large changes in activity (i.e. it is highly regulated) but if these changes have little effect on the flux of a metabolic pathway, then this enzyme is not involved in the control of the pathway.[96] There are multiple levels of metabolic regulation. In intrinsic regulation, the metabolic pathway self-regulates to respond to changes in the levels of substrates or products; for example, a decrease in the amount of product can increase the flux through the pathway to compensate.[95] This type of regulation often involves allosteric regulation of the activities of multiple enzymes in the pathway.[97] Extrinsic control involves a cell in a multicellular Effect of insulin on glucose uptake and metabolism. Insulin binds to its receptor organism changing its metabolism in (1) which in turn starts many protein activation cascades (2). These include: response to signals from other cells. These translocation of Glut-4 transporter to the plasma membrane and influx of glucose (3), glycogen synthesis (4), glycolysis (5) and fatty acid synthesis (6). signals are usually in the form of soluble messengers such as hormones and growth factors and are detected by specific receptors on the cell surface.[98] These signals are then transmitted inside the cell by second messenger systems that often involved the phosphorylation of proteins.[99] A very well understood example of extrinsic control is the regulation of glucose metabolism by the hormone insulin.[100] Insulin is produced in response to rises in blood glucose levels. Binding of the hormone to insulin receptors on cells then activates a cascade of protein kinases that cause the cells to take up glucose and convert it into storage molecules such as fatty acids and glycogen.[101] The metabolism of glycogen is controlled by activity of phosphorylase, the enzyme that breaks down glycogen, and glycogen synthase, the enzyme that makes it. These enzymes are regulated in a reciprocal fashion, with phosphorylation inhibiting glycogen synthase, but activating phosphorylase. Insulin causes glycogen synthesis by activating protein phosphatases and producing a decrease in the phosphorylation of these enzymes.[102]

Overview of metabolism

212

Evolution
Further information: Molecular evolution and phylogenetics The central pathways of metabolism described above, such as glycolysis and the citric acid cycle, are present in all three domains of living things and were present in the last universal ancestor.[3] [103] This universal ancestral cell was prokaryotic and probably a methanogen that had extensive amino acid, nucleotide, carbohydrate and lipid [104] [105] metabolism. The retention of these ancient pathways during later evolution may be the result of these reactions being an optimal solution to their particular metabolic problems, Evolutionary tree showing the common ancestry of organisms from all three domains of life. Bacteria are colored blue, eukaryotes red, and archaea green. Relative positions of with pathways such as glycolysis and some of the phyla included are shown around the tree. the citric acid cycle producing their end products highly efficiently and in a [4] [5] minimal number of steps. Mutation changes that affect non-coding DNA segments may merely affect the metabolic efficiency of the individual for whom the mutation occurs.[106] The first pathways of enzyme-based metabolism may have been parts of purine nucleotide metabolism, with previous metabolic pathways being part of the ancient RNA world.[107] Many models have been proposed to describe the mechanisms by which novel metabolic pathways evolve. These include the sequential addition of novel enzymes to a short ancestral pathway, the duplication and then divergence of entire pathways as well as the recruitment of pre-existing enzymes and their assembly into a novel reaction pathway.[108] The relative importance of these mechanisms is unclear, but genomic studies have shown that enzymes in a pathway are likely to have a shared ancestry, suggesting that many pathways have evolved in a step-by-step fashion with novel functions being created from pre-existing steps in the pathway.[109] An alternative model comes from studies that trace the evolution of proteins' structures in metabolic networks, this has suggested that enzymes are pervasively recruited, borrowing enzymes to perform similar functions in different metabolic pathways (evident in the MANET database)[110] These recruitment processes result in an evolutionary enzymatic mosaic.[111] A third possibility is that some parts of metabolism might exist as "modules" that can be reused in different pathways and perform similar functions on different molecules.[112] As well as the evolution of new metabolic pathways, evolution can also cause the loss of metabolic functions. For example, in some parasites metabolic processes that are not essential for survival are lost and preformed amino acids, nucleotides and carbohydrates may instead be scavenged from the host.[113] Similar reduced metabolic capabilities are seen in endosymbiotic organisms.[114]

Overview of metabolism

213

Investigation and manipulation

Further information: Protein methods, proteomics, metabolomics and metabolic network modelling Classically, metabolism is studied by a reductionist approach that focuses on a single metabolic pathway. Particularly valuable is the use of radioactive tracers at the whole-organism, tissue and cellular levels, which define the paths from precursors to final products by identifying radioactively labelled intermediates and products.[115] The enzymes that catalyze these chemical reactions can then be purified and their kinetics and responses to inhibitors investigated. A parallel approach is to identify the small molecules in a cell or tissue; the complete set of these molecules is called the metabolome. Overall, these studies give a good view of the structure and function of simple metabolic pathways, but are inadequate when applied to more complex systems such as the metabolism of a complete cell.[116]

An idea of the complexity of the metabolic networks in cells that contain thousands of different enzymes is given by the figure showing the interactions between just 43 proteins and 40 metabolites to the right: the sequences of genomes provide lists containing anything up to 45,000 genes.[117] However, it is now possible to use this genomic data to reconstruct complete networks of biochemical reactions and produce more holistic mathematical models that may explain and predict their behavior.[118] These models are especially powerful when used to integrate the pathway and metabolite data obtained through classical methods with data on gene expression from proteomic and DNA microarray studies.[119] Using these techniques, a model of human metabolism has now been produced, which will guide future drug discovery and biochemical research.[120] These models are now being used in network analysis, to classify human diseases into groups that share common proteins or metabolites.[121] [122] Bacterial metabolic networks seem to be a striking example of bow-tie[123] [124] [125] organization, an architecture able to input a wide range of nutrients and produce a large variety of products and complex macromolecules using a relatively few intermediate common currencies. A major technological application of this information is metabolic engineering. Here, organisms such as yeast, plants or bacteria are genetically modified to make them more useful in biotechnology and aid the production of drugs such as antibiotics or industrial chemicals such as 1,3-propanediol and shikimic acid.[126] These genetic modifications usually aim to reduce the amount of energy used to produce the product, increase yields and reduce the production of wastes.[127]

Metabolic network of the Arabidopsis thaliana citric acid cycle. Enzymes and metabolites are shown as red squares and the interactions between them as black lines.

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214

History
Further information: History of biochemistry and history of molecular biology The term metabolism is derived from the Greek "Metabolismos" for "change", or "overthrow".[128] The history of the scientific study of metabolism spans several centuries and has moved from examining whole animals in early studies, to examining individual metabolic reactions in modern biochemistry. The first controlled experiments in human metabolism were published by Santorio Santorio in 1614 in his book Ars de statica medicina.[129] He described how he weighed himself before and after eating, sleep, working, sex, fasting, drinking, and excreting. He found that most of the food he took in was lost through what he called "insensible perspiration". In these early studies, the mechanisms of these metabolic processes had not been identified and a vital force was thought to animate living tissue.[130] In the 19th century, when studying the fermentation of sugar to alcohol by yeast, Louis Pasteur concluded that fermentation was catalyzed by substances Santorio Santorio in his steelyard within the yeast cells he called "ferments". He wrote that "alcoholic balance, from Ars de statica medicina, fermentation is an act correlated with the life and organization of the yeast first published 1614 cells, not with the death or putrefaction of the cells."[131] This discovery, along with the publication by Friedrich Whler in 1828 of the chemical synthesis of urea,[132] notable for being the first organic compound prepared from wholly inorganic precursors, proved that the organic compounds and chemical reactions found in cells were no different in principle than any other part of chemistry. It was the discovery of enzymes at the beginning of the 20th century by Eduard Buchner that separated the study of the chemical reactions of metabolism from the biological study of cells, and marked the beginnings of biochemistry.[133] The mass of biochemical knowledge grew rapidly throughout the early 20th century. One of the most prolific of these modern biochemists was Hans Krebs who made huge contributions to the study of metabolism.[134] He discovered the urea cycle and later, working with Hans Kornberg, the citric acid cycle and the glyoxylate cycle.[135] [61] Modern biochemical research has been greatly aided by the development of new techniques such as chromatography, X-ray diffraction, NMR spectroscopy, radioisotopic labelling, electron microscopy and molecular dynamics simulations. These techniques have allowed the discovery and detailed analysis of the many molecules and metabolic pathways in cells.

References
[1] Friedrich C (1998). "Physiology and genetics of sulfur-oxidizing bacteria". Adv Microb Physiol 39: 23589. doi:10.1016/S0065-2911(08)60018-1. PMID9328649. [2] Pace NR (January 2001). "The universal nature of biochemistry". Proc. Natl. Acad. Sci. U.S.A. 98 (3): 8058. Bibcode2001PNAS...98..805P. doi:10.1073/pnas.98.3.805. PMC33372. PMID11158550. [3] Smith E, Morowitz H (2004). "Universality in intermediary metabolism" (http:/ / www. pnas. org/ cgi/ pmidlookup?view=long& pmid=15340153). Proc Natl Acad Sci USA 101 (36): 1316873. Bibcode2004PNAS..10113168S. doi:10.1073/pnas.0404922101. PMC516543. PMID15340153. . [4] Ebenhh O, Heinrich R (2001). "Evolutionary optimization of metabolic pathways. Theoretical reconstruction of the stoichiometry of ATP and NADH producing systems". Bull Math Biol 63 (1): 2155. doi:10.1006/bulm.2000.0197. PMID11146883. [5] Melndez-Hevia E, Waddell T, Cascante M (1996). "The puzzle of the Krebs citric acid cycle: assembling the pieces of chemically feasible reactions, and opportunism in the design of metabolic pathways during evolution". J Mol Evol 43 (3): 293303. doi:10.1007/BF02338838. PMID8703096. [6] Michie K, Lwe J (2006). "Dynamic filaments of the bacterial cytoskeleton". Annu Rev Biochem 75: 46792. doi:10.1146/annurev.biochem.75.103004.142452. PMID16756499.

Overview of metabolism
[7] Nelson, David L.; Michael M. Cox (2005). Lehninger Principles of Biochemistry. New York: W. H. Freeman and company. pp.841. ISBN0-7167-4339-6. [8] Fahy E, Subramaniam S, Brown H, Glass C, Merrill A, Murphy R, Raetz C, Russell D, Seyama Y, Shaw W, Shimizu T, Spener F, van Meer G, VanNieuwenhze M, White S, Witztum J, Dennis E (2005). "A comprehensive classification system for lipids" (http:/ / www. jlr. org/ cgi/ content/ full/ 46/ 5/ 839). J Lipid Res 46 (5): 83961. doi:10.1194/jlr.E400004-JLR200. PMID15722563. . [9] "Nomenclature of Lipids" (http:/ / www. chem. qmul. ac. uk/ iupac/ lipid/ ). IUPAC-IUB Commission on Biochemical Nomenclature (CBN). . Retrieved 2007-03-08. [10] Hegardt F (1999). "Mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase: a control enzyme in ketogenesis". Biochem J 338 (Pt 3): 56982. doi:10.1042/0264-6021:3380569. PMC1220089. PMID10051425. [11] Raman R, Raguram S, Venkataraman G, Paulson J, Sasisekharan R (2005). "Glycomics: an integrated systems approach to structure-function relationships of glycans". Nat Methods 2 (11): 81724. doi:10.1038/nmeth807. PMID16278650. [12] Sierra S, Kupfer B, Kaiser R (2005). "Basics of the virology of HIV-1 and its replication". J Clin Virol 34 (4): 23344. doi:10.1016/j.jcv.2005.09.004. PMID16198625. [13] Wimmer M, Rose I (1978). "Mechanisms of enzyme-catalyzed group transfer reactions". Annu Rev Biochem 47: 103178. doi:10.1146/annurev.bi.47.070178.005123. PMID354490. [14] Mitchell P (1979). "The Ninth Sir Hans Krebs Lecture. Compartmentation and communication in living systems. Ligand conduction: a general catalytic principle in chemical, osmotic and chemiosmotic reaction systems". Eur J Biochem 95 (1): 120. doi:10.1111/j.1432-1033.1979.tb12934.x. PMID378655. [15] Dimroth P, von Ballmoos C, Meier T (March 2006). "Catalytic and mechanical cycles in F-ATP synthases. Fourth in the Cycles Review Series". EMBO Rep 7 (3): 27682. doi:10.1038/sj.embor.7400646. PMC1456893. PMID16607397. [16] Coulston, Ann; Kerner, John; Hattner, JoAnn; Srivastava, Ashini (2006). "Nutrition Principles and Clinical Nutrition". Stanford School of Medicine Nutrition Courses. SUMMIT. [17] Pollak N, Dlle C, Ziegler M (2007). "The power to reduce: pyridine nucleotidessmall molecules with a multitude of functions". Biochem J 402 (2): 20518. doi:10.1042/BJ20061638. PMC1798440. PMID17295611. [18] Heymsfield S, Waki M, Kehayias J, Lichtman S, Dilmanian F, Kamen Y, Wang J, Pierson R (1991). "Chemical and elemental analysis of humans in vivo using improved body composition models". Am J Physiol 261 (2 Pt 1): E1908. PMID1872381. [19] Sychrov H (2004). "Yeast as a model organism to study transport and homeostasis of alkali metal cations" (http:/ / www. biomed. cas. cz/ physiolres/ pdf/ 53 Suppl 1/ 53_S91. pdf) (PDF). Physiol Res 53 Suppl 1: S918. PMID15119939. . [20] Levitan I (1988). "Modulation of ion channels in neurons and other cells". Annu Rev Neurosci 11: 11936. doi:10.1146/annurev.ne.11.030188.001003. PMID2452594. [21] Dulhunty A (2006). "Excitation-contraction coupling from the 1950s into the new millennium". Clin Exp Pharmacol Physiol 33 (9): 76372. doi:10.1111/j.1440-1681.2006.04441.x. PMID16922804. [22] Mahan D, Shields R (1998). "Macro- and micromineral composition of pigs from birth to 145 kilograms of body weight" (http:/ / jas. fass. org/ cgi/ reprint/ 76/ 2/ 506). J Anim Sci 76 (2): 50612. PMID9498359. . [23] Husted S, Mikkelsen B, Jensen J, Nielsen N (2004). "Elemental fingerprint analysis of barley (Hordeum vulgare) using inductively coupled plasma mass spectrometry, isotope-ratio mass spectrometry, and multivariate statistics". Anal Bioanal Chem 378 (1): 17182. doi:10.1007/s00216-003-2219-0. PMID14551660. [24] Finney L, O'Halloran T (2003). "Transition metal speciation in the cell: insights from the chemistry of metal ion receptors". Science 300 (5621): 9316. Bibcode2003Sci...300..931F. doi:10.1126/science.1085049. PMID12738850. [25] Cousins R, Liuzzi J, Lichten L (2006). "Mammalian zinc transport, trafficking, and signals" (http:/ / www. jbc. org/ cgi/ content/ full/ 281/ 34/ 24085). J Biol Chem 281 (34): 240859. doi:10.1074/jbc.R600011200. PMID16793761. . [26] Dunn L, Rahmanto Y, Richardson D (2007). "Iron uptake and metabolism in the new millennium". Trends Cell Biol 17 (2): 93100. doi:10.1016/j.tcb.2006.12.003. PMID17194590. [27] Nealson K, Conrad P (1999). "Life: past, present and future" (http:/ / rstb. royalsocietypublishing. org/ cgi/ pmidlookup?view=long& pmid=10670014). Philos Trans R Soc Lond B Biol Sci 354 (1392): 192339. doi:10.1098/rstb.1999.0532. PMC1692713. PMID10670014. . [28] Hse C, Finkelstein R (December 1993). "Bacterial extracellular zinc-containing metalloproteases" (http:/ / mmbr. asm. org/ cgi/ pmidlookup?view=long& pmid=8302217). Microbiol Rev 57 (4): 82337. PMC372940. PMID8302217. . [29] Gupta R, Gupta N, Rathi P (2004). "Bacterial lipases: an overview of production, purification and biochemical properties". Appl Microbiol Biotechnol 64 (6): 76381. doi:10.1007/s00253-004-1568-8. PMID14966663. [30] Hoyle T (1997). "The digestive system: linking theory and practice". Br J Nurs 6 (22): 128591. PMID9470654. [31] Souba W, Pacitti A (1992). "How amino acids get into cells: mechanisms, models, menus, and mediators". JPEN J Parenter Enteral Nutr 16 (6): 56978. doi:10.1177/0148607192016006569. PMID1494216. [32] Barrett M, Walmsley A, Gould G (1999). "Structure and function of facilitative sugar transporters". Curr Opin Cell Biol 11 (4): 496502. doi:10.1016/S0955-0674(99)80072-6. PMID10449337. [33] Bell G, Burant C, Takeda J, Gould G (1993). "Structure and function of mammalian facilitative sugar transporters". J Biol Chem 268 (26): 191614. PMID8366068. [34] Bouch C, Serdy S, Kahn C, Goldfine A (2004). "The cellular fate of glucose and its relevance in type 2 diabetes" (http:/ / edrv. endojournals. org/ cgi/ content/ full/ 25/ 5/ 807). Endocr Rev 25 (5): 80730. doi:10.1210/er.2003-0026. PMID15466941. .

215

Overview of metabolism
[35] Sakami W, Harrington H (1963). "Amino acid metabolism". Annu Rev Biochem 32: 35598. doi:10.1146/annurev.bi.32.070163.002035. PMID14144484. [36] Brosnan J (2000). "Glutamate, at the interface between amino acid and carbohydrate metabolism" (http:/ / jn. nutrition. org/ cgi/ content/ full/ 130/ 4/ 988S). J Nutr 130 (4S Suppl): 988S90S. PMID10736367. . [37] Young V, Ajami A (2001). "Glutamine: the emperor or his clothes?" (http:/ / jn. nutrition. org/ cgi/ content/ full/ 131/ 9/ 2449S). J Nutr 131 (9 Suppl): 2449S59S; discussion 2486S7S. PMID11533293. . [38] Hosler J, Ferguson-Miller S, Mills D (2006). "Energy transduction: proton transfer through the respiratory complexes". Annu Rev Biochem 75: 16587. doi:10.1146/annurev.biochem.75.062003.101730. PMC2659341. PMID16756489. [39] Schultz B, Chan S (2001). "Structures and proton-pumping strategies of mitochondrial respiratory enzymes". Annu Rev Biophys Biomol Struct 30: 2365. doi:10.1146/annurev.biophys.30.1.23. PMID11340051. [40] Capaldi R, Aggeler R (2002). "Mechanism of the F(1)F(0)-type ATP synthase, a biological rotary motor". Trends Biochem Sci 27 (3): 15460. doi:10.1016/S0968-0004(01)02051-5. PMID11893513. [41] Friedrich B, Schwartz E (1993). "Molecular biology of hydrogen utilization in aerobic chemolithotrophs". Annu Rev Microbiol 47: 35183. doi:10.1146/annurev.mi.47.100193.002031. PMID8257102. [42] Weber K, Achenbach L, Coates J (2006). "Microorganisms pumping iron: anaerobic microbial iron oxidation and reduction". Nat Rev Microbiol 4 (10): 75264. doi:10.1038/nrmicro1490. PMID16980937. [43] Jetten M, Strous M, van de Pas-Schoonen K, Schalk J, van Dongen U, van de Graaf A, Logemann S, Muyzer G, van Loosdrecht M, Kuenen J (1998). "The anaerobic oxidation of ammonium". FEMS Microbiol Rev 22 (5): 42137. doi:10.1111/j.1574-6976.1998.tb00379.x. PMID9990725. [44] Simon J (2002). "Enzymology and bioenergetics of respiratory nitrite ammonification". FEMS Microbiol Rev 26 (3): 285309. doi:10.1111/j.1574-6976.2002.tb00616.x. PMID12165429. [45] Conrad R (1996). "Soil microorganisms as controllers of atmospheric trace gases (H2, CO, CH4, OCS, N2O, and NO)" (http:/ / mmbr. asm. org/ cgi/ pmidlookup?view=long& pmid=8987358). Microbiol Rev 60 (4): 60940. PMC239458. PMID8987358. . [46] Barea J, Pozo M, Azcn R, Azcn-Aguilar C (2005). "Microbial co-operation in the rhizosphere" (http:/ / jxb. oxfordjournals. org/ cgi/ content/ full/ 56/ 417/ 1761). J Exp Bot 56 (417): 176178. doi:10.1093/jxb/eri197. PMID15911555. . [47] van der Meer M, Schouten S, Bateson M, Nbel U, Wieland A, Khl M, de Leeuw J, Sinninghe Damst J, Ward D (July 2005). "Diel variations in carbon metabolism by green nonsulfur-like bacteria in alkaline siliceous hot spring microbial mats from Yellowstone National Park" (http:/ / aem. asm. org/ cgi/ pmidlookup?view=long& pmid=16000812). Appl Environ Microbiol 71 (7): 397886. doi:10.1128/AEM.71.7.3978-3986.2005. PMC1168979. PMID16000812. . [48] Tichi M, Tabita F (2001). "Interactive control of Rhodobacter capsulatus redox-balancing systems during phototrophic metabolism" (http:/ / jb. asm. org/ cgi/ pmidlookup?view=long& pmid=11591679). J Bacteriol 183 (21): 634454. doi:10.1128/JB.183.21.6344-6354.2001. PMC100130. PMID11591679. . [49] Allen J, Williams J (1998). "Photosynthetic reaction centers". FEBS Lett 438 (12): 59. doi:10.1016/S0014-5793(98)01245-9. PMID9821949. [50] Munekage Y, Hashimoto M, Miyake C, Tomizawa K, Endo T, Tasaka M, Shikanai T (2004). "Cyclic electron flow around photosystem I is essential for photosynthesis". Nature 429 (6991): 57982. Bibcode2004Natur.429..579M. doi:10.1038/nature02598. PMID15175756. [51] Miziorko H, Lorimer G (1983). "Ribulose-1,5-bisphosphate carboxylase-oxygenase". Annu Rev Biochem 52: 50735. doi:10.1146/annurev.bi.52.070183.002451. PMID6351728. [52] Dodd A, Borland A, Haslam R, Griffiths H, Maxwell K (2002). "Crassulacean acid metabolism: plastic, fantastic" (http:/ / jxb. oxfordjournals. org/ cgi/ content/ full/ 53/ 369/ 569). J Exp Bot 53 (369): 56980. doi:10.1093/jexbot/53.369.569. PMID11886877. . [53] Hgler M, Wirsen C, Fuchs G, Taylor C, Sievert S (May 2005). "Evidence for autotrophic CO2 fixation via the reductive tricarboxylic acid cycle by members of the epsilon subdivision of proteobacteria" (http:/ / jb. asm. org/ cgi/ pmidlookup?view=long& pmid=15838028). J Bacteriol 187 (9): 30207. doi:10.1128/JB.187.9.3020-3027.2005. PMC1082812. PMID15838028. . [54] Strauss G, Fuchs G (1993). "Enzymes of a novel autotrophic CO2 fixation pathway in the phototrophic bacterium Chloroflexus aurantiacus, the 3-hydroxypropionate cycle". Eur J Biochem 215 (3): 63343. doi:10.1111/j.1432-1033.1993.tb18074.x. PMID8354269. [55] Wood H (1991). "Life with CO or CO2 and H2 as a source of carbon and energy" (http:/ / www. fasebj. org/ cgi/ reprint/ 5/ 2/ 156). FASEB J 5 (2): 15663. PMID1900793. . [56] Shively J, van Keulen G, Meijer W (1998). "Something from almost nothing: carbon dioxide fixation in chemoautotrophs". Annu Rev Microbiol 52: 191230. doi:10.1146/annurev.micro.52.1.191. PMID9891798. [57] Boiteux A, Hess B (1981). "Design of glycolysis". Philos Trans R Soc Lond B Biol Sci 293 (1063): 522. Bibcode1981RSPTB.293....5B. doi:10.1098/rstb.1981.0056. PMID6115423. [58] Pilkis S, el-Maghrabi M, Claus T (1990). "Fructose-2,6-bisphosphate in control of hepatic gluconeogenesis. From metabolites to molecular genetics". Diabetes Care 13 (6): 58299. doi:10.2337/diacare.13.6.582. PMID2162755. [59] Ensign S (2006). "Revisiting the glyoxylate cycle: alternate pathways for microbial acetate assimilation". Mol Microbiol 61 (2): 2746. doi:10.1111/j.1365-2958.2006.05247.x. PMID16856935. [60] Finn P, Dice J (2006). "Proteolytic and lipolytic responses to starvation". Nutrition 22 (78): 83044. doi:10.1016/j.nut.2006.04.008. PMID16815497. [61] Kornberg H, Krebs H (1957). "Synthesis of cell constituents from C2-units by a modified tricarboxylic acid cycle". Nature 179 (4568): 98891. Bibcode1957Natur.179..988K. doi:10.1038/179988a0. PMID13430766.

216

Overview of metabolism
[62] Rademacher T, Parekh R, Dwek R (1988). "Glycobiology". Annu Rev Biochem 57: 785838. doi:10.1146/annurev.bi.57.070188.004033. PMID3052290. [63] Opdenakker G, Rudd P, Ponting C, Dwek R (1993). "Concepts and principles of glycobiology" (http:/ / www. fasebj. org/ cgi/ reprint/ 7/ 14/ 1330). FASEB J 7 (14): 13307. PMID8224606. . [64] McConville M, Menon A (2000). "Recent developments in the cell biology and biochemistry of glycosylphosphatidylinositol lipids (review)". Mol Membr Biol 17 (1): 116. doi:10.1080/096876800294443. PMID10824734. [65] Chirala S, Wakil S (2004). "Structure and function of animal fatty acid synthase". Lipids 39 (11): 104553. doi:10.1007/s11745-004-1329-9. PMID15726818. [66] White S, Zheng J, Zhang Y (2005). "The structural biology of type II fatty acid biosynthesis". Annu Rev Biochem 74: 791831. doi:10.1146/annurev.biochem.74.082803.133524. PMID15952903. [67] Ohlrogge J, Jaworski J (1997). "Regulation of fatty acid synthesis". Annu Rev Plant Physiol Plant Mol Biol 48: 109136. doi:10.1146/annurev.arplant.48.1.109. PMID15012259. [68] Dubey V, Bhalla R, Luthra R (2003). "An overview of the non-mevalonate pathway for terpenoid biosynthesis in plants" (http:/ / www. ias. ac. in/ jbiosci/ sep2003/ 637. pdf) (PDF). J Biosci 28 (5): 63746. doi:10.1007/BF02703339. PMID14517367. . [69] Kuzuyama T, Seto H (2003). "Diversity of the biosynthesis of the isoprene units". Nat Prod Rep 20 (2): 17183. doi:10.1039/b109860h. PMID12735695. [70] Grochowski L, Xu H, White R (May 2006). "Methanocaldococcus jannaschii uses a modified mevalonate pathway for biosynthesis of isopentenyl diphosphate" (http:/ / jb. asm. org/ cgi/ pmidlookup?view=long& pmid=16621811). J Bacteriol 188 (9): 31928. doi:10.1128/JB.188.9.3192-3198.2006. PMC1447442. PMID16621811. . [71] Lichtenthaler H (1999). "The 1-Ddeoxy-D-xylulose-5-phosphate pathway of isoprenoid biosynthesis in plants". Annu Rev Plant Physiol Plant Mol Biol 50: 4765. doi:10.1146/annurev.arplant.50.1.47. PMID15012203. [72] Schroepfer G (1981). "Sterol biosynthesis". Annu Rev Biochem 50: 585621. doi:10.1146/annurev.bi.50.070181.003101. PMID7023367. [73] Lees N, Skaggs B, Kirsch D, Bard M (1995). "Cloning of the late genes in the ergosterol biosynthetic pathway of Saccharomyces cerevisiaea review". Lipids 30 (3): 2216. doi:10.1007/BF02537824. PMID7791529. [74] Guyton, Arthur C.; John E. Hall (2006). Textbook of Medical Physiology. Philadelphia: Elsevier. pp.8556. ISBN0-7216-0240-1. [75] Ibba M, Sll D (2001). "The renaissance of aminoacyl-tRNA synthesis" (http:/ / www. molcells. org/ home/ journal/ include/ downloadPdf. asp?articleuid={A158E3B4-2423-4806-9A30-4B93CDA76DA0}). EMBO Rep 2 (5): 3827. doi:10.1093/embo-reports/kve095. PMC1083889. PMID11375928. . [76] Lengyel P, Sll D (1969). "Mechanism of protein biosynthesis". Bacteriol Rev 33 (2): 264301. PMC378322. PMID4896351. [77] Rudolph F (1994). "The biochemistry and physiology of nucleotides". J Nutr 124 (1 Suppl): 124S127S. PMID8283301. Zrenner R, Stitt M, Sonnewald U, Boldt R (2006). "Pyrimidine and purine biosynthesis and degradation in plants". Annu Rev Plant Biol 57: 80536. doi:10.1146/annurev.arplant.57.032905.105421. PMID16669783. [78] Stasolla C, Katahira R, Thorpe T, Ashihara H (2003). "Purine and pyrimidine nucleotide metabolism in higher plants". J Plant Physiol 160 (11): 127195. doi:10.1078/0176-1617-01169. PMID14658380. [79] Smith J (1995). "Enzymes of nucleotide synthesis". Curr Opin Struct Biol 5 (6): 7527. doi:10.1016/0959-440X(95)80007-7. PMID8749362. [80] Testa B, Krmer S (2006). "The biochemistry of drug metabolisman introduction: part 1. Principles and overview". Chem Biodivers 3 (10): 1053101. doi:10.1002/cbdv.200690111. PMID17193224. [81] Danielson P (2002). "The cytochrome P450 superfamily: biochemistry, evolution and drug metabolism in humans". Curr Drug Metab 3 (6): 56197. doi:10.2174/1389200023337054. PMID12369887. [82] King C, Rios G, Green M, Tephly T (2000). "UDP-glucuronosyltransferases". Curr Drug Metab 1 (2): 14361. doi:10.2174/1389200003339171. PMID11465080. [83] Sheehan D, Meade G, Foley V, Dowd C (November 2001). "Structure, function and evolution of glutathione transferases: implications for classification of non-mammalian members of an ancient enzyme superfamily" (http:/ / www. biochemj. org/ bj/ 360/ 0001/ bj3600001. htm). Biochem J 360 (Pt 1): 116. doi:10.1042/0264-6021:3600001. PMC1222196. PMID11695986. . [84] Galvo T, Mohn W, de Lorenzo V (2005). "Exploring the microbial biodegradation and biotransformation gene pool". Trends Biotechnol 23 (10): 497506. doi:10.1016/j.tibtech.2005.08.002. PMID16125262. [85] Janssen D, Dinkla I, Poelarends G, Terpstra P (2005). "Bacterial degradation of xenobiotic compounds: evolution and distribution of novel enzyme activities". Environ Microbiol 7 (12): 186882. doi:10.1111/j.1462-2920.2005.00966.x. PMID16309386. [86] Davies K (1995). "Oxidative stress: the paradox of aerobic life". Biochem Soc Symp 61: 131. PMID8660387. [87] Tu B, Weissman J (2004). "Oxidative protein folding in eukaryotes: mechanisms and consequences" (http:/ / www. jcb. org/ cgi/ content/ full/ 164/ 3/ 341). J Cell Biol 164 (3): 3416. doi:10.1083/jcb.200311055. PMC2172237. PMID14757749. . [88] Sies H (1997). "Oxidative stress: oxidants and antioxidants" (http:/ / ep. physoc. org/ cgi/ reprint/ 82/ 2/ 291. pdf) (PDF). Exp Physiol 82 (2): 2915. PMID9129943. . [89] Vertuani S, Angusti A, Manfredini S (2004). "The antioxidants and pro-antioxidants network: an overview". Curr Pharm Des 10 (14): 167794. doi:10.2174/1381612043384655. PMID15134565. [90] von Stockar U, Liu J (1999). "Does microbial life always feed on negative entropy? Thermodynamic analysis of microbial growth". Biochim Biophys Acta 1412 (3): 191211. doi:10.1016/S0005-2728(99)00065-1. PMID10482783.

217

Overview of metabolism
[91] Demirel Y, Sandler S (2002). "Thermodynamics and bioenergetics". Biophys Chem 97 (23): 87111. doi:10.1016/S0301-4622(02)00069-8. PMID12050002. [92] Albert R (2005). "Scale-free networks in cell biology" (http:/ / jcs. biologists. org/ cgi/ content/ full/ 118/ 21/ 4947). J Cell Sci 118 (Pt 21): 494757. doi:10.1242/jcs.02714. PMID16254242. . [93] Brand M (1997). "Regulation analysis of energy metabolism" (http:/ / jeb. biologists. org/ cgi/ reprint/ 200/ 2/ 193). J Exp Biol 200 (Pt 2): 193202. PMID9050227. . [94] Soyer O, Salath M, Bonhoeffer S (2006). "Signal transduction networks: topology, response and biochemical processes". J Theor Biol 238 (2): 41625. doi:10.1016/j.jtbi.2005.05.030. PMID16045939. [95] Salter M, Knowles R, Pogson C (1994). "Metabolic control". Essays Biochem 28: 112. PMID7925313. [96] Westerhoff H, Groen A, Wanders R (1984). "Modern theories of metabolic control and their applications (review)". Biosci Rep 4 (1): 122. doi:10.1007/BF01120819. PMID6365197. [97] Fell D, Thomas S (1995). "Physiological control of metabolic flux: the requirement for multisite modulation". Biochem J 311 (Pt 1): 359. PMC1136115. PMID7575476. [98] Hendrickson W (2005). "Transduction of biochemical signals across cell membranes". Q Rev Biophys 38 (4): 32130. doi:10.1017/S0033583506004136. PMID16600054. [99] Cohen P (2000). "The regulation of protein function by multisite phosphorylationa 25 year update". Trends Biochem Sci 25 (12): 596601. doi:10.1016/S0968-0004(00)01712-6. PMID11116185. [100] Lienhard G, Slot J, James D, Mueckler M (1992). "How cells absorb glucose". Sci Am 266 (1): 8691. doi:10.1038/scientificamerican0192-86. PMID1734513. [101] Roach P (2002). "Glycogen and its metabolism". Curr Mol Med 2 (2): 10120. doi:10.2174/1566524024605761. PMID11949930. [102] Newgard C, Brady M, O'Doherty R, Saltiel A (2000). "Organizing glucose disposal: emerging roles of the glycogen targeting subunits of protein phosphatase-1" (http:/ / diabetes. diabetesjournals. org/ cgi/ reprint/ 49/ 12/ 1967. pdf) (PDF). Diabetes 49 (12): 196777. doi:10.2337/diabetes.49.12.1967. PMID11117996. . [103] Romano A, Conway T (1996). "Evolution of carbohydrate metabolic pathways". Res Microbiol 147 (67): 44855. doi:10.1016/0923-2508(96)83998-2. PMID9084754. [104] Koch A (1998). "How did bacteria come to be?". Adv Microb Physiol 40: 35399. doi:10.1016/S0065-2911(08)60135-6. PMID9889982. [105] Ouzounis C, Kyrpides N (1996). "The emergence of major cellular processes in evolution". FEBS Lett 390 (2): 11923. doi:10.1016/0014-5793(96)00631-X. PMID8706840. [106] C.Michael Hogan. 2010. Mutation. ed. E.Monosson and C.J.Cleveland. Encyclopedia of Earth. National Council for Science and the Environment. Washington DC (http:/ / www. eoearth. org/ article/ Mutation?topic=49496) [107] Caetano-Anolles G, Kim HS, Mittenthal JE (2007). "The origin of modern metabolic networks inferred from phylogenomic analysis of protein architecture". Proc Natl Acad Sci USA 104 (22): 935863. Bibcode2007PNAS..104.9358C. doi:10.1073/pnas.0701214104. PMC1890499. PMID17517598. [108] Schmidt S, Sunyaev S, Bork P, Dandekar T (2003). "Metabolites: a helping hand for pathway evolution?". Trends Biochem Sci 28 (6): 33641. doi:10.1016/S0968-0004(03)00114-2. PMID12826406. [109] Light S, Kraulis P (2004). "Network analysis of metabolic enzyme evolution in Escherichia coli". BMC Bioinformatics 5: 15. doi:10.1186/1471-2105-5-15. PMC394313. PMID15113413. Alves R, Chaleil R, Sternberg M (2002). "Evolution of enzymes in metabolism: a network perspective". J Mol Biol 320 (4): 75170. doi:10.1016/S0022-2836(02)00546-6. PMID12095253. [110] Kim HS, Mittenthal JE, Caetano-Anolles G (2006). "MANET: tracing evolution of protein architecture in metabolic networks". BMC Bioinformatics 19 (7): 351. doi:10.1186/1471-2105-7-351. PMC1559654. PMID16854231. [111] Teichmann SA, Rison SC, Thornton JM, Riley M, Gough J, Chothia C (2001). "Small-molecule metabolsim: an enzyme mosaic". Trends Biotechnol 19 (12): 4826. doi:10.1016/S0167-7799(01)01813-3. PMID11711174. [112] Spirin V, Gelfand M, Mironov A, Mirny L (June 2006). "A metabolic network in the evolutionary context: multiscale structure and modularity" (http:/ / www. pnas. org/ cgi/ pmidlookup?view=long& pmid=16731630). Proc Natl Acad Sci USA 103 (23): 87749. Bibcode2006PNAS..103.8774S. doi:10.1073/pnas.0510258103. PMC1482654. PMID16731630. . [113] Lawrence J (2005). "Common themes in the genome strategies of pathogens". Curr Opin Genet Dev 15 (6): 5848. doi:10.1016/j.gde.2005.09.007. PMID16188434. Wernegreen J (2005). "For better or worse: genomic consequences of intracellular mutualism and parasitism". Curr Opin Genet Dev 15 (6): 57283. doi:10.1016/j.gde.2005.09.013. PMID16230003. [114] Pl C, Papp B, Lercher M, Csermely P, Oliver S, Hurst L (2006). "Chance and necessity in the evolution of minimal metabolic networks". Nature 440 (7084): 66770. Bibcode2006Natur.440..667P. doi:10.1038/nature04568. PMID16572170. [115] Rennie M (1999). "An introduction to the use of tracers in nutrition and metabolism". Proc Nutr Soc 58 (4): 93544. doi:10.1017/S002966519900124X. PMID10817161. [116] Phair R (1997). "Development of kinetic models in the nonlinear world of molecular cell biology". Metabolism 46 (12): 148995. doi:10.1016/S0026-0495(97)90154-2. PMID9439549. [117] Sterck L, Rombauts S, Vandepoele K, Rouz P, Van de Peer Y (2007). "How many genes are there in plants (... and why are they there)?". Curr Opin Plant Biol 10 (2): 199203. doi:10.1016/j.pbi.2007.01.004. PMID17289424. [118] Borodina I, Nielsen J (2005). "From genomes to in silico cells via metabolic networks". Curr Opin Biotechnol 16 (3): 3505. doi:10.1016/j.copbio.2005.04.008. PMID15961036.

218

Overview of metabolism
[119] Gianchandani E, Brautigan D, Papin J (2006). "Systems analyses characterize integrated functions of biochemical networks". Trends Biochem Sci 31 (5): 28491. doi:10.1016/j.tibs.2006.03.007. PMID16616498. [120] Duarte NC, Becker SA, Jamshidi N, et al. (February 2007). "Global reconstruction of the human metabolic network based on genomic and bibliomic data" (http:/ / www. pnas. org/ cgi/ pmidlookup?view=long& pmid=17267599). Proc. Natl. Acad. Sci. U.S.A. 104 (6): 177782. Bibcode2007PNAS..104.1777D. doi:10.1073/pnas.0610772104. PMC1794290. PMID17267599. . [121] Goh KI, Cusick ME, Valle D, Childs B, Vidal M, Barabsi AL (May 2007). "The human disease network". Proc. Natl. Acad. Sci. U.S.A. 104 (21): 868590. Bibcode2007PNAS..104.8685G. doi:10.1073/pnas.0701361104. PMC1885563. PMID17502601. [122] Lee DS, Park J, Kay KA, Christakis NA, Oltvai ZN, Barabsi AL (July 2008). "The implications of human metabolic network topology for disease comorbidity" (http:/ / www. pnas. org/ lookup/ pmid?view=long& pmid=18599447). Proc. Natl. Acad. Sci. U.S.A. 105 (29): 98809885. Bibcode2008PNAS..105.9880L. doi:10.1073/pnas.0802208105. PMC2481357. PMID18599447. . [123] Csete M, Doyle J (2004). "Bow ties, metabolism and disease". Trends Biotechnol. 22 (9): 44650. doi:10.1016/j.tibtech.2004.07.007. PMID15331224. [124] Ma HW, Zeng AP (2003). "The connectivity structure, giant strong component and centrality of metabolic networks". Bioinformatics 19 (11): 142330. doi:10.1093/bioinformatics/btg177. PMID12874056. [125] Zhao J, Yu H, Luo JH, Cao ZW, Li YX (2006). "Hierarchical modularity of nested bow-ties in metabolic networks". BMC Bioinformatics 7: 386. doi:10.1186/1471-2105-7-386. PMC1560398. PMID16916470. [126] Thykaer J, Nielsen J (2003). "Metabolic engineering of beta-lactam production". Metab Eng 5 (1): 5669. doi:10.1016/S1096-7176(03)00003-X. PMID12749845. Gonzlez-Pajuelo M, Meynial-Salles I, Mendes F, Andrade J, Vasconcelos I, Soucaille P (2005). "Metabolic engineering of Clostridium acetobutylicum for the industrial production of 1,3-propanediol from glycerol". Metab Eng 7 (56): 32936. doi:10.1016/j.ymben.2005.06.001. PMID16095939. Krmer M, Bongaerts J, Bovenberg R, Kremer S, Mller U, Orf S, Wubbolts M, Raeven L (2003). "Metabolic engineering for microbial production of shikimic acid". Metab Eng 5 (4): 27783. doi:10.1016/j.ymben.2003.09.001. PMID14642355. [127] Koffas M, Roberge C, Lee K, Stephanopoulos G (1999). "Metabolic engineering". Annu Rev Biomed Eng 1: 53557. doi:10.1146/annurev.bioeng.1.1.535. PMID11701499. [128] "Metabolism" (http:/ / www. etymonline. com/ index. php?term=metabolism). The Online Etymology Dictionary. . Retrieved 2007-02-20. [129] Eknoyan G (1999). "Santorio Sanctorius (15611636) founding father of metabolic balance studies". Am J Nephrol 19 (2): 22633. doi:10.1159/000013455. PMID10213823. [130] Williams, H. S. (1904) A History of Science: in Five Volumes. Volume IV: Modern Development of the Chemical and Biological Sciences (http:/ / etext. lib. virginia. edu/ toc/ modeng/ public/ Wil4Sci. html) Harper and Brothers (New York) Retrieved on 2007-03-26 [131] Dubos J. (1951). "Louis Pasteur: Free Lance of Science, Gollancz. Quoted in Manchester K. L. (1995) Louis Pasteur (18221895)chance and the prepared mind". Trends Biotechnol 13 (12): 511515. doi:10.1016/S0167-7799(00)89014-9. PMID8595136. [132] Kinne-Saffran E, Kinne R (1999). "Vitalism and synthesis of urea. From Friedrich Whler to Hans A. Krebs". Am J Nephrol 19 (2): 2904. doi:10.1159/000013463. PMID10213830. [133] Eduard Buchner's 1907 Nobel lecture (http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1907/ buchner-lecture. html) at http:/ / nobelprize. org Accessed 2007-03-20 [134] Kornberg H (2000). "Krebs and his trinity of cycles". Nat Rev Mol Cell Biol 1 (3): 2258. doi:10.1038/35043073. PMID11252898. [135] Krebs HA, Henseleit K (1932). "Untersuchungen ber die Harnstoffbildung im tierkorper". Z. Physiol. Chem. 210: 3366. Krebs H, Johnson W (April 1937). "Metabolism of ketonic acids in animal tissues". Biochem J 31 (4): 64560. PMC1266984. PMID16746382.

219

Further reading
Introductory Rose, S. and Mileusnic, R., The Chemistry of Life. (Penguin Press Science, 1999), ISBN 0-14-027273-9 Schneider, E. D. and Sagan, D., Into the Cool: Energy Flow, Thermodynamics, and Life. (University Of Chicago Press, 2005), ISBN 0-226-73936-8 Lane, N., Oxygen: The Molecule that Made the World. (Oxford University Press, USA, 2004), ISBN 0-19-860783-0 Advanced Price, N. and Stevens, L., Fundamentals of Enzymology: Cell and Molecular Biology of Catalytic Proteins. (Oxford University Press, 1999), ISBN 0-19-850229-X Berg, J. Tymoczko, J. and Stryer, L., Biochemistry. (W. H. Freeman and Company, 2002), ISBN 0-7167-4955-6 Cox, M. and Nelson, D. L., Lehninger Principles of Biochemistry. (Palgrave Macmillan, 2004), ISBN 0-7167-4339-6

Overview of metabolism Brock, T. D. Madigan, M. T. Martinko, J. and Parker J., Brock's Biology of Microorganisms. (Benjamin Cummings, 2002), ISBN 0-13-066271-2 Da Silva, J.J.R.F. and Williams, R. J. P., The Biological Chemistry of the Elements: The Inorganic Chemistry of Life. (Clarendon Press, 1991), ISBN 0-19-855598-9 Nicholls, D. G. and Ferguson, S. J., Bioenergetics. (Academic Press Inc., 2002), ISBN 0-12-518121-3

220

External links
biochemical families: prot nucl carb (glpr, alco, glys) lipd (fata/i, phld, strd, gllp, eico) amac/i ncbs/i ttpy/i

221

Carbohydrate metabolism
Glycolysis
Glycolysis (from glycose, an older term[1] for glucose + -lysis degradation) is the metabolic pathway that converts glucose C6H12O6, into pyruvate, CH3COCOO + H+. The free energy released in this process is used to form the high-energy compounds ATP (adenosine triphosphate) and NADH (reduced nicotinamide adenine dinucleotide).
Glycolysis overview

Glycolysis is a definite sequence of ten reactions involving ten intermediate compounds (one of the steps involves two intermediates). The intermediates provide entry points to glycolysis. For example, most monosaccharides, such as fructose, glucose, and galactose, can be converted to one of these intermediates. The intermediates may also be directly useful. For example, the intermediate dihydroxyacetone phosphate (DHAP) is a source of the glycerol that combines with fatty acids to form fat. It occurs, with variations, in nearly all organisms, both aerobic and anaerobic. The wide occurrence of glycolysis indicates that it is one of the most ancient known metabolic pathways.[2] It occur in the cytosol of the cell. The most common type of glycolysis is the Embden-Meyerhof-Parnas pathway (EMP pathway), which was first discovered by Gustav Embden, Otto Meyerhof and Jakub Karol Parnas. Glycolysis also refers to other pathways, such as the EntnerDoudoroff pathway and various heterofermentative and homofermentative pathways. However, the discussion here will be limited to the Embden-Meyerhof pathway. The entire glycolysis pathway can be separated into two phases[3] : 1. The Preparatory Phase - in which ATP is consumed and is hence also known as the investment phase 2. The Pay Off Phase - in which ATP is produced.

Overview
The overall reaction of glycolysis is:
D-[Glucose] + 2 [NAD]+ + 2 [ADP] + 2 [P]i 2 [Pyruvate] + 2 [NADH] + 2 H+ + 2 [ATP] + 2 H2O

The use of symbols in this equation makes it appear unbalanced with respect to oxygen atoms, hydrogen atoms, and charges. Atom balance is maintained by the two phosphate (Pi) groups:[4] each exists in the form of a hydrogen phosphate anion (HPO42-), dissociating to contribute 2 H+ overall

Glycolysis each liberates an oxygen atom when it binds to an ADP (adenosine diphosphate) molecule, contributing 2 O overall Charges are balanced by the difference between ADP and ATP. In the cellular environment, all three hydroxy groups of ADP dissociate into -O- and H+, giving ADP3-, and this ion tends to exist in an ionic bond with Mg2+, giving ADPMg-. ATP behaves identically except that it has four hydroxy groups, giving ATPMg2-. When these differences along with the true charges on the two phosphate groups are considered together, the net charges of -4 on each side are balanced. For simple anaerobic fermentations, the metabolism of one molecule of glucose to two molecules of pyruvate has a net yield of two molecules of ATP. Most cells will then carry out further reactions to 'repay' the used NAD+ and produce a final product of ethanol or lactic acid. Many bacteria use inorganic compounds as hydrogen acceptors to regenerate the NAD+. Cells performing aerobic respiration synthesize much more ATP, but not as part of glycolysis. These further aerobic reactions use pyruvate and NADH + H+ from glycolysis. Eukaryotic aerobic respiration produces approximately 34 additional molecules of ATP for each glucose molecule, however most of these are produced by a vastly different mechanism to the substrate-level phosphorylation in glycolysis.
Glycolysis

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The lower-energy production, per glucose, of anaerobic respiration relative to aerobic respiration, results in greater flux through the pathway under hypoxic (low-oxygen) conditions, unless alternative sources of anaerobically-oxidizable substrates, such as fatty acids, are found.

Elucidation of the pathway

In 1860, Louis Pasteur discovered that microorganisms are responsible for fermentation. In 1897, Eduard Buchner found that extracts of certain cells can cause fermentation. In 1905, Arthur Harden and William Youngalong with Nick Sheppard determined that a heat-sensitive high-molecular-weight subcellular fraction (the enzymes) and a heat-insensitive low-molecular-weight cytoplasm fraction (ADP, ATP and NAD+ and other cofactors) are required together for fermentation to proceed. The details of the pathway were eventually determined by 1940, with a major input from Otto Meyerhof and some years later by Luis Leloir. The biggest difficulties in determining the intricacies of the pathway were due to the very short lifetime and low steady-state concentrations of the intermediates of the fast glycolytic reactions.

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Sequence of reactions
Preparatory phase
The first five steps are regarded as the preparatory (or investment) phase, since they consume energy to convert the glucose into two three-carbon sugar phosphates[3] (G3P).
The first step in glycolysis is phosphorylation of glucose by a family of enzymes called hexokinases to form glucose 6-phosphate (G6P). This reaction consumes ATP, but it acts to keep the glucose concentration low, promoting continuous transport of glucose into the cell through the plasma membrane transporters. In addition, it blocks the glucose from leaking out the cell lacks transporters for G6P, and free diffusion out of the cell is prevented due to the charged nature of G6P. Glucose may alternatively be from the phosphorolysis or hydrolysis of intracellular starch or glycogen. In animals, an isozyme of hexokinase called glucokinase is also used in the liver, which has a much lower affinity for glucose (Km in the vicinity of normal glycemia), and differs in regulatory properties. The different substrate affinity and alternate regulation of this enzyme are a reflection of the role of the liver in maintaining blood sugar levels. Cofactors: Mg2+

D-Glucose (Glc)

Hexokinase (HK) a transferase

-D-Glucose-6-phosphate (G6P)

ATP

H+ + ADP

G6P is then rearranged into fructose 6-phosphate (F6P) by glucose phosphate isomerase. Fructose can also enter the glycolytic pathway by phosphorylation at this point. The change in structure is an isomerization, in which the G6P has been converted to F6P. The reaction requires an enzyme, phosphohexose isomerase, to proceed. This reaction is freely reversible under normal cell conditions. However, it is often driven forward because of a low concentration of F6P, which is constantly consumed during the next step of glycolysis. Under conditions of high F6P concentration, this reaction readily runs in reverse. This phenomenon can be explained through Le Chatelier's Principle. Isomerization to a keto sugar is necessary for carbanion stabilization in the fourth reaction step (below).

-D-Glucose 6-phosphate (G6P)

Phosphoglucose isomerase an isomerase

-D-Fructose 6-phosphate (F6P)

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224

The energy expenditure of another ATP in this step is justified in 2 ways: The glycolytic process (up to this step) is now irreversible, and the energy supplied destabilizes the molecule. Because the reaction catalyzed by Phosphofructokinase 1 (PFK-1) is coupled to the hydrolysis of ATP, an energetically favorable step, it is, in essence, irreversible, and a different pathway must be used to do the reverse conversion during gluconeogenesis. This makes the reaction a key regulatory point (see below). This is also the rate-limiting step. Furthermore, the second phosphorylation event is necessary to allow the formation of two charged groups (rather than only one) in the subsequent step of glycolysis, ensuring the prevention of free diffusion of substrates out of the cell. The same reaction can also be catalyzed by pyrophosphate-dependent phosphofructokinase (PFP or PPi-PFK), which is found in most plants, some bacteria, archea, and protists, but not in animals. This enzyme uses pyrophosphate (PPi) as a phosphate donor instead of ATP. It is a reversible reaction, [5] increasing the flexibility of glycolytic metabolism. A rarer ADP-dependent PFK enzyme variant has [6] been identified in archaean species. Cofactors: Mg2+

-D-Fructose 6-phosphate (F6P)

phosphofructokinase (PFK-1) a transferase ATP H+ + ADP

-D-Fructose 1,6-bisphosphate (F1,6BP)

Destabilizing the molecule in the previous reaction allows the hexose ring to be split by aldolase into two triose sugars, dihydroxyacetone phosphate, a ketone, and glyceraldehyde 3-phosphate, an aldehyde. There are two classes of aldolases: class I aldolases, present in animals and plants, and class II aldolases, present in fungi and bacteria; the two classes use different mechanisms in cleaving the ketose ring. Electrons delocalized in the carbon-carbon bond cleavage associate with the alcohol group. The resulting carbanion is stabilized by the structure of the carbanion itself via resonance charge distribution and by the presence of a charged ion prosthetic group.

-D-Fructose 1,6-bisphosphate (F1,6BP)

fructose bisphosphate aldolase (ALDO) a lyase

D-glyceraldehyde 3-phosphate (GADP)

Dihydroxyacetone phosphate (DHAP)

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225

Triosephosphate isomerase rapidly interconverts dihydroxyacetone phosphate with glyceraldehyde 3-phosphate (GADP) that proceeds further into glycolysis. This is advantageous, as it directs dihydroxyacetone phosphate down the same pathway as glyceraldehyde 3-phosphate, simplifying regulation.

Dihydroxyacetone phosphate (DHAP)

triosephosphate isomerase (TPI) an isomerase

D-glyceraldehyde 3-phosphate (GADP)

Pay-off phase
The second half of glycolysis is known as the pay-off phase, characterised by a net gain of the energy-rich molecules ATP and NADH[3] . Since glucose leads to two triose sugars in the preparatory phase, each reaction in the pay-off phase occurs twice per glucose molecule. This yields 2 NADH molecules and 4 ATP molecules, leading to a net gain of 2 NADH molecules and 2 ATP molecules from the glycolytic pathway per glucose.
The triose sugars are dehydrogenated and inorganic phosphate is added to them, forming 1,3-bisphosphoglycerate. The hydrogen is used to reduce two molecules of NAD+, a hydrogen carrier, to give NADH + H+ for each triose. Hydrogen atom balance and charge balance are both maintained because the phosphate (Pi) group actually exists in the form of a [4] hydrogen phosphate anion (HPO42-), which dissociates to contribute the extra H+ ion and gives a net charge of -3 on both sides.

glyceraldehyde 3-phosphate (GADP)

glyceraldehyde phosphate dehydrogenase (GAPDH) an oxidoreductase

D-1,3-bisphosphoglycerate (1,3BPG)

NAD+ + Pi

NADH + H+

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226

This step is the enzymatic transfer of a phosphate group from 1,3-bisphosphoglycerate to ADP by phosphoglycerate kinase, forming ATP and 3-phosphoglycerate. At this step, glycolysis has reached the break-even point: 2 molecules of ATP were consumed, and 2 new molecules have now been synthesized. This step, one of the two substrate-level phosphorylation steps, requires ADP; thus, when the cell has plenty of ATP (and little ADP), this reaction does not occur. Because ATP decays relatively quickly when it is not metabolized, this is an important regulatory point in the glycolytic pathway. ADP actually exists as ADPMg-, and ATP as ATPMg2-, balancing the charges at -5 both sides. Cofactors: Mg2+

1,3-bisphosphoglycerate (1,3-BPG)

phosphoglycerate kinase (PGK) a transferase

3-phosphoglycerate (3-P-G)

ADP

ATP

phosphoglycerate kinase (PGK)

Phosphoglycerate mutase now forms 2-phosphoglycerate.

3-phosphoglycerate (3PG)

phosphoglycerate mutase (PGM) a mutase

2-phosphoglycerate (2PG)

Enolase next forms phosphoenolpyruvate from 2-phosphoglycerate. Cofactors: 2 Mg2+: one "conformational" ion to coordinate with the carboxylate group of the substrate, and one "catalytic" ion that participates in the dehydration.

2-phosphoglycerate (2PG)

enolase (ENO) a lyase H2O

phosphoenolpyruvate (PEP)

enolase (ENO)

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227

A final substrate-level phosphorylation now forms a molecule of pyruvate and a molecule of ATP by means of the enzyme pyruvate kinase. This serves as an additional regulatory step, similar to the phosphoglycerate kinase step. Cofactors: Mg2+

phosphoenolpyruvate (PEP)

pyruvate kinase (PK) a transferase ADP + H+ ATP

pyruvate (Pyr)

Regulation
Glycolysis is regulated by slowing down or speeding up certain steps in the glycolysis pathway. This is accomplished by inhibiting or activating the enzymes that are involved. The steps that are regulated may be determined by calculating the change in free energy, G, for each step. If a step's products and reactants are in equilibrium, then the step is assumed to not be regulated. Since the change in free energy is zero for a system at equilibrium, any step with a free energy change near zero is not being regulated. If a step is being regulated, then that step's enzyme is not converting reactants into products as fast as it could, resulting in a build-up of reactants, which would be converted to products if the enzyme were operating faster. Since the reaction is thermodynamically favorable, the change in free energy for the step will be negative. A step with a large negative change in free energy is assumed to be regulated.

Free energy changes Concentrations of metabolites in erythrocytes[7]

Compound glucose glucose-6-phosphate fructose-6-phosphate fructose-1,6-bisphosphate dihydroxyacetone phosphate glyceraldehyde-3-phosphate 1,3-bisphosphoglycerate 2,3-bisphosphoglycerate 3-phosphoglycerate 2-phosphoglycerate phosphoenolpyruvate pyruvate ATP ADP Pi 5.0 0.083 0.014 0.031 0.14 0.019 0.001 4.0 0.12 0.03 0.023 0.051 1.85 0.14 1.0 Concentration / mM

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228

The change in free energy for each step of glycolysis estimated from the concentration of metabolites in an erythrocyte.

The change in free energy, G, for each step in the glycolysis pathway can be calculated using G = G' + RTln Q, where Q is the reaction quotient. This requires knowing the concentrations of the metabolites. All of these values are available for erythrocytes, with the exception of the concentrations of NAD+ and NADH. The ratio of NAD+ to NADH in the cytoplasm is approximately 1000 in the step 6, something that makes the oxidation of glyceraldehyde-3-phosphate more favourable. Using the measured concentrations of each step, and the standard free energy changes, the actual free energy change can be calculated. (Neglecting this is very common - the delta G of ATP hydrolysis in cells is not the standard free energy change of ATP hydrolysis quoted in textbooks).

Change in free energy for each step of glycolysis[8]

Step 1 2 3 4 5 6 7 8 9 10 Reaction glucose + ATP4- glucose-6-phosphate2- + ADP3- + H+ glucose-6-phosphate2- fructose-6-phosphate2fructose-6-phosphate2- + ATP4- fructose-1,6-bisphosphate4- + ADP3- + H+ G' / (kJ/mol) G / (kJ/mol) -16.7 1.67 -14.2 -34 -2.9 -19 -0.23 2.4 -1.29 0.09 0.83 1.1 -23.0

fructose-1,6-bisphosphate4- dihydroxyacetone phosphate2- + glyceraldehyde-3-phosphate2- 23.9 dihydroxyacetone phosphate2- glyceraldehyde-3-phosphate2glyceraldehyde-3-phosphate2- + Pi2- + NAD+ 1,3-bisphosphoglycerate4- + NADH + H+ 1,3-bisphosphoglycerate4- + ADP3- 3-phosphoglycerate3- + ATP43-phosphoglycerate3- 2-phosphoglycerate32-phosphoglycerate3- phosphoenolpyruvate3- + H2O phosphoenolpyruvate3- + ADP3- + H+ pyruvate- + ATP47.56 6.30 -18.9 4.4 1.8 -31.7

From measuring the physiological concentrations of metabolites in an erythrocyte it seems that about seven of the steps in glycolysis are in equilibrium for that cell type. Three of the steps the ones with large negative free energy changes are not in equilibrium and are referred to as irreversible; such steps are often subject to regulation. Step 5 in the figure is shown behind the other steps, because that step is a side-reaction that can decrease or increase the concentration of the intermediate glyceraldehyde-3-phosphate. That compound is converted to dihydroxyacetone

Glycolysis phosphate by the enzyme triose phosphate isomerase, which is a catalytically perfect enzyme; its rate is so fast that the reaction can be assumed to be in equilibrium. The fact that G is not zero indicates that the actual concentrations in the erythrocyte are not accurately known.

229

Biochemical logic
The existence of more than one point of regulation indicates that intermediates between those points enter and leave the glycolysis pathway by other processes. For example, in the first regulated step, hexokinase converts glucose into glucose-6-phosphate. Instead of continuing through the glycolysis pathway, this intermediate can be converted into glucose storage molecules, such as glycogen or starch. The reverse reaction, breaking down, e.g., glycogen, produces mainly glucose-6-phosphate; very little free glucose is formed in the reaction. The glucose-6-phosphate so produced can enter glycolysis after the first control point. In the second regulated step (the third step of glycolysis), phosphofructokinase converts fructose-6-phosphate into fructose-1,6-bisphosphate, which then is converted into glyceraldehyde-3-phosphate and dihydroxyacetone phosphate. The dihydroxyacetone phosphate can be removed from glycolysis by conversion into glycerol-3-phosphate, which can be used to form triglycerides.[9] On the converse, triglycerides can be broken down into fatty acids and glycerol; the latter, in turn, can be converted into dihydroxyacetone phosphate, which can enter glycolysis after the second control point.

Regulation
The three regulated enzymes are hexokinase, phosphofructokinase, and pyruvate kinase. The flux through the glycolytic pathway is adjusted in response to conditions both inside and outside the cell. The rate in liver is regulated to meet major cellular needs: (1) the production of ATP, (2) the provision of building blocks for biosynthetic reactions, and (3) to lower blood glucose, one of the major functions of the liver. When blood sugar falls, glycolysis is halted in the liver to allow the reverse process, gluconeogenesis. In glycolysis, the reactions catalyzed by hexokinase, phosphofructokinase, and pyruvate kinase are effectively irreversible in most organisms. In metabolic pathways, such enzymes are potential sites of control, and all three enzymes serve this purpose in glycolysis. Hexokinase In animals, regulation of blood glucose levels by the pancreas in conjunction with the liver is a vital part of homeostasis. In liver cells, extra G6P (glucose-6-phosphate) may be converted to G1P for conversion to glycogen, or it is alternatively converted by glycolysis to acetyl-CoA and then citrate. Excess citrate is exported to the cytosol, where ATP citrate lyase will regenerate acetyl-CoA and OAA. The acetyl-CoA is then used for fatty acid synthesis and cholesterol synthesis, two important ways of utilizing excess glucose when its [10] Yeast hexokinase B. PDB 1IG8 . concentration is high in blood. Liver contains both hexokinase and glucokinase; the latter catalyses the phosphorylation of glucose to G6P and is not inhibited by G6P. Thus, it allows glucose to be converted into glycogen, fatty acids, and cholesterol even when hexokinase activity is low.[11] This is important when blood glucose levels are high. During hypoglycemia, the glycogen can be converted back to G6P and then converted to glucose by the liver-specific enzyme glucose 6-phosphatase. This reverse reaction is an important role of liver cells to maintain blood sugars levels during fasting. This is critical for brain function, since the brain utilizes glucose as an energy source under most conditions.

Glycolysis Phosphofructokinase Phosphofructokinase is an important control point in the glycolytic pathway, since it is one of the irreversible steps and has key allosteric effectors, AMP and fructose 2,6-bisphosphate (F2,6BP). Fructose 2,6-bisphosphate (F2,6BP) is a very potent activator of phosphofructokinase (PFK-1), which is synthesised when F6P is phosphorylated by a second phosphofructokinase (PFK2). In liver, when blood sugar is low and glucagon elevates cAMP, PFK2 is phosphorylated by protein kinase A. The phosphorylation inactivates PFK2, and another domain on this protein becomes active as fructose 2,6-bisphosphatase, which converts F2,6BP back to F6P. Both Bacillus stearothermophilus phosphofructokinase. [12] glucagon and epinephrine cause high levels of cAMP in the liver. The PDB 6PFK . result of lower levels of liver fructose-2,6-bisphosphate is a decrease in activity of phosphofructokinase and an increase in activity of fructose 1,6-bisphosphatase, so that gluconeogenesis (in essence, "glycolysis in reverse") is favored. This is consistent with the role of the liver in such situations, since the response of the liver to these hormones is to release glucose to the blood. ATP competes with AMP for the allosteric effector site on the PFK enzyme. ATP concentrations in cells are much higher than those of AMP, typically 100-fold higher,[13] but the concentration of ATP does not change more than about 10% under physiological conditions, whereas a 10% drop in ATP results in a 6-fold increase in AMP.[14] Thus, the relevance of ATP as an allosteric effector is questionable. An increase in AMP is a consequence of a decrease in energy charge in the cell. Citrate inhibits phosphofructokinase when tested in vitro by enhancing the inhibitory effect of ATP. However, it is doubtful that this is a meaningful effect in vivo, because citrate in the cytosol is utilized mainly for conversion to acetyl-CoA for fatty acid and cholesterol synthesis. Pyruvate kinase This enzyme catalyzes the last step of glycolysis, in which pyruvate and ATP are formed. Regulation of this enzyme is discussed in the main topic, pyruvate kinase.

230

Post-glycolysis processes
The overall process of glycolysis is: glucose + 2 NAD+ + 2 ADP + 2 Pi 2 pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O If glycolysis were to continue indefinitely, all of the NAD+ would be used up, and glycolysis would stop. To allow glycolysis to continue, organisms must be able to oxidize NADH back to NAD+.
Yeast pyruvate kinase. PDB 1A3W [15] .

Fermentation
One method of doing this is to simply have the pyruvate do the oxidation; in this process, the pyruvate is converted to lactate (the conjugate base of lactic acid) in a process called lactic acid fermentation: pyruvate + NADH + H+ lactate + NAD+

Glycolysis This process occurs in the bacteria involved in making yogurt (the lactic acid causes the milk to curdle). This process also occurs in animals under hypoxic (or partially-anaerobic) conditions, found, for example, in overworked muscles that are starved of oxygen, or in infarcted heart muscle cells. In many tissues, this is a cellular last resort for energy; most animal tissue cannot maintain anaerobic respiration for an extended length of time. Some organisms, such as yeast, convert NADH back to NAD+ in a process called ethanol fermentation. In this process, the pyruvate is converted first to acetaldehyde and carbon dioxide, then to ethanol. Lactic acid fermentation and ethanol fermentation can occur in the absence of oxygen. This anaerobic fermentation allows many single-cell organisms to use glycolysis as their only energy source.

231

Anaerobic respiration
In the above two examples of fermentation, NADH is oxidized by transferring two electrons to pyruvate. However, anaerobic bacteria use a wide variety of compounds as the terminal electron acceptors in cellular respiration: nitrogenous compounds, such as nitrates and nitrites; sulfur compounds, such as sulfates, sulfites, sulfur dioxide, and elemental sulfur; carbon dioxide; iron compounds; manganese compounds; cobalt compounds; and uranium compounds.

Aerobic respiration
In aerobic organisms, a complex mechanism has been developed to use the oxygen in air as the final electron acceptor of respiration. First, pyruvate is converted to acetyl-CoA and CO2 within the mitochondria in a process called pyruvate decarboxylation. Second, the acetyl-CoA enters the citric acid cycle, also known as Krebs Cycle, where it is fully oxidized to carbon dioxide and water, producing yet more NADH. Third, the NADH is oxidized to NAD+ by the electron transport chain, using oxygen as the final electron acceptor. This process creates a "hydrogen ion gradient" across the inner membrane of the mitochondria. Fourth, the proton gradient is used to produce a large amount of ATP in a process called oxidative phosphorylation.

Intermediates for other pathways

This article concentrates on the catabolic role of glycolysis with regard to converting potential chemical energy to usable chemical energy during the oxidation of glucose to pyruvate.many of the metabolites in the glycolytic pathway are also used by anabolic pathways, and, as a consequence, flux through the pathway is critical to maintain a supply of carbon skeletons for biosynthesis. In addition, not all carbon entering the pathway leaves as pyruvate and may be extracted at earlier stages to provide carbon compounds for other pathways. These metabolic pathways are all strongly reliant on glycolysis as a source of metabolites: Gluconeogenesis Lipid metabolism Pentose phosphate pathway Citric acid cycle, which in turn leads to: Amino acid synthesis Nucleotide synthesis Tetrapyrrole synthesis From an anabolic metabolism perspective, the NADH has a role to drive synthetic reactions, doing so by directly or indirectly reducing the pool of NADP+ in the cell to NADPH, which is another important reducing agent for

Glycolysis biosynthetic pathways in a cell.

232

Glycolysis in disease
Genetic diseases
Glycolytic mutations are generally rare due to importance of the metabolic pathway, this means that the majority of occurring mutations result in an inability for the cell to respire, and therefore cause the death of the cell at an early stage. However, some mutations are seen with one notable example being Pyruvate kinase deficiency, leading to chronic hemolytic anemia.

Cancer
Malignant rapidly-growing tumor cells typically have glycolytic rates that are up to 200 times higher than those of their normal tissues of origin. This phenomenon was first described in 1930 by Otto Warburg and is referred to as the Warburg effect. The Warburg hypothesis claims that cancer is primarily caused by dysfunctionality in mitochondrial metabolism, rather than because of uncontrolled growth of cells. A number of theories have been advanced to explain the Warburg effect. This high glycolysis rate has important medical applications, as high aerobic glycolysis by malignant tumors is utilized clinically to diagnose and monitor treatment responses of cancers by imaging uptake of 2-18F-2-deoxyglucose (FDG) (a radioactive modified hexokinase substrate) with positron emission tomography (PET).[16] [17] There is ongoing research to affect mitochondrial metabolism and treat cancer by reducing glycolysis and thus starving cancerous cells in various new ways, including a ketogenic diet.

Alzheimer's disease
Disfunctioning glycolysis or glucose metabolism in fronto-temporo-parietal and cingulate cortices has been associated with Alzheimer's disease,[18] probably due to the decreased amyloid (1-42) (A42) and increased tau, phosphorylated tau in cerebrospinal fluid (CSF) [19]

Alternative nomenclature
Some of the metabolites in glycolysis have alternative names and nomenclature. In part, this is because some of them are common to other pathways, such as the Calvin cycle.
This article 1 3 4 5 6 7 glucose fructose 6-phosphate fructose 1,6-bisphosphate Glc F6P F1,6BP fructose 1,6-diphosphate glycerone phosphate 3-phosphoglyceraldehyde PGAL, G3P, GALP,GAP,TP FBP, FDP, F1,6DP Alternative names dextrose Alternative nomenclature

dihydroxyacetone phosphate DHAP glyceraldehyde 3-phosphate GADP 1,3-bisphosphoglycerate

1,3BPG glycerate PGAP, BPG, DPG 1,3-bisphosphate, glycerate 1,3-diphosphate, 1,3-diphosphoglycerate 3PG 2PG PEP glycerate 3-phosphate glycerate 2-phosphate PGA, GP

8 9

3-phosphoglycerate 2-phosphoglycerate

10 phosphoenolpyruvate

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233
11 pyruvate Pyr pyruvic acid

References
[1] [2] [3] [4] Webster's New International Dictionary of the English Language, 2nd ed. (1937) Merriam Company, Springfield, Mass. Romano AH, Conway T. (1996) Evolution of carbohydrate metabolic pathways. Res Microbiol. 147(6-7):448-55 PMID 9084754 Glycolysis - Animation and Notes (http:/ / pharmaxchange. info/ press/ 2011/ 09/ glycolysis-animation-and-notes/ ) Lane, A. N.; Fan, T. W. -M.; Higashi, R. M. (2009). "Metabolic acidosis and the importance of balanced equations". Metabolomics 5 (2): 163165. doi:10.1007/s11306-008-0142-2. [5] Reeves, R. E.; South D. J., Blytt H. J. and Warren L. G. (1974). "Pyrophosphate: D-fructose 6-phosphate 1-phosphotransferase. A new enzyme with the glycolytic function 6-phosphate 1-phosphotransferase". J Biol Chem 249 (24): 77377741. PMID4372217. [6] Selig, M.; Xavier K. B., Santos H. and Schnheit P. (1997). "Comparative analysis of Embden-Meyerhof and Entner-Doudoroff glycolytic pathways in hyperthermophilic archaea and the bacterium Thermotoga". Arch Microbiol 167 (4): 217232. PMID9075622. [7] Garrett, R.; Grisham, C. M. (2005). Biochemistry (3rd ed.). Belmont, CA: Thomson Brooks/Cole. p.584. ISBN0-534-49011-6. [8] Garrett, R.; Grisham, C. M. (2005). Biochemistry (3rd ed.). Belmont, CA: Thomson Brooks/Cole. pp.582583. ISBN0-534-49011-6. [9] Berg, J. M.; Tymoczko, J. L.; Stryer, L. (2007). Biochemistry (6th ed.). New York: Freeman. p.622. ISBN0-534-49011-6. [10] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1IG8 [11] Voet D., and Voet J. G. (2004). Biochemistry 3rd Edition (New York, John Wiley & Sons, Inc.) [12] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=6PFK [13] Beis I., and Newsholme E. A. (1975). The contents of adenine nucleotides, phosphagens and some glycolytic intermediates in resting muscles from vertebrates and invertebrates. Biochem J 152, 23-32. [14] Voet D., and Voet J. G. (2004). Biochemistry 3rd Edition (New York, John Wiley & Sons, Inc.). [15] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=1A3W [16] "PET Scan: PET Scan Info Reveals ..." (http:/ / www. petscaninfo. com/ ). . Retrieved December 5, 2005. [17] "4320139 549..559" (http:/ / biogenomica. com/ PDFs/ PauwelsPETandHexokinase. pdf). . Retrieved December 5, 2005. [18] Hunt, A . et al.; Schonknecht, P; Henze, M; Seidl, U; Haberkorn, U; Schroder, J (2007). "Reduced cerebral glucose metabolism in patients at risk for Alzheimer's disease". Psychiatry Research: Neuroimaging 155 (2): 147154. doi:10.1016/j.pscychresns.2006.12.003. PMID17524628. [19] Hunt, A . et al.; Van Der Flier, WM; Blankenstein, MA; Bouwman, FH; Van Kamp, GJ; Barkhof, F; Scheltens, P (2008). "CSF and MRI markers independently contribute to the diagnosis of Alzheimer's disease". Neurobiology of Aging 29 (5): 669675. doi:10.1016/j.neurobiolaging.2006.11.018. PMID17208336.

External links
A Detailed Glycolysis Animation provided by [[IUBMB (http://www.iubmb-nicholson.org/swf/glycolysis. swf)]] ( Adobe Flash (http://get.adobe.com/flashplayer/) Required) The Glycolytic enzymes in Glycolysis (http://nist.rcsb.org/pdb/molecules/pdb50_1.html) at Protein Data Bank Glycolytic cycle with animations (http://www.wdv.com/CellWorld/Biochemistry/Glycolytic) at wdv.com Metabolism, Cellular Respiration and Photosynthesis - The Virtual Library of Biochemistry and Cell Biology (http://www.biochemweb.org/metabolism.shtml) at biochemweb.org notes on glycolysis (http://www.rahulgladwin.com/blog/2007/01/notes-on-glycolysis.html) at rahulgladwin.com The chemical logic behind glycolysis (http://www2.ufp.pt/~pedros/bq/glycolysis.htm) at ufp.pt Expasy biochemical pathways poster (http://www.expasy.org/tools/pathways/boehringer_legends.html) at ExPASy Mnemonic at medicalmnemonics.com 317 5468 (http://www.medicalmnemonics.com/cgi-bin/lookup. cfm?id1=317&id2=5468&id3=&id4=)

Gluconeogenesis

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Gluconeogenesis
Gluconeogenesis (abbreviated GNG) is a metabolic pathway that results in the generation of glucose from non-carbohydrate carbon substrates such as lactate, glycerol, and glucogenic amino acids. It is one of the two main mechanisms humans and many other animals use to keep blood glucose levels from dropping too low (hypoglycemia). The other means of maintaining blood glucose levels is through the degradation of glycogen (glycogenolysis).[1] Gluconeogenesis is a ubiquitous process, present in plants, animals, fungi, bacteria, and other microorganisms.[2] In animals, gluconeogenesis takes place mainly in the liver and, to a lesser extent, in the cortex of kidneys. This process occurs during periods of fasting, starvation, low-carbohydrate diets, or intense exercise and is highly endergonic. For example, the pathway leading from pyruvate to glucose-6-phosphate requires 4 molecules of ATP and 2 molecules of GTP. Gluconeogenesis is often associated with ketosis. Gluconeogenesis is also a target of therapy for type II diabetes, such as metformin, which inhibits glucose formation and stimulates glucose uptake by cells.[3]

Entering the pathway

Lactate is transported back to the liver where it is converted into pyruvate by the Cori cycle using the enzyme lactate dehydrogenase. Pyruvate, the first designated substrate of the gluconeogenic pathway, can then be used to generate glucose.[4]

Gluconeogenesis pathway with key molecules and enzymes. Many steps are the opposite of those found in the glycolysis.

All citric acid cycle intermediates, through conversion to oxaloacetate, amino acids other than lysine or leucine, and glycerol can also function as substrates for gluconeogenesis.[4] Transamination or deamination of amino acids facilitates entering of their carbon skeleton into the cycle directly (as pyruvate or oxaloacetate), or indirectly via the citric acid cycle. Whether fatty acids can be converted into glucose in animals has been a longstanding question in biochemistry.[6] It is known that odd-chain fatty acids can be oxidized to yield propionyl CoA, a precursor for succinyl CoA, which can be converted to

Catabolism of proteinogenic amino acids. Amino acids are classified according the [5] abilities of their products to enter gluconeogenesis: Glucogenic amino acids have this abilityKetogenic amino acids do not. These products may still be used for ketogenesis or lipid synthesis.Some amino acids are catabolized into both glucogenic and ketogenic products.

Gluconeogenesis pyruvate and enter into gluconeogenesis. In plants, specifically seedlings, the glyoxylate cycle can be used to convert fatty acids (acetate) into the primary carbon source of the organism. The glyoxylate cycle produces four-carbon dicarboxylic acids that can enter gluconeogenesis.[4] In 1995, researchers identified the glyoxylate cycle in nematodes.[7] In addition, the glyoxylate enzymes malate synthase and isocitrate lyase have been found in animal tissues.[8] Genes coding for malate synthase gene have been identified in other [metazoans] including arthropods, echinoderms, and even some vertebrates. Mammals found to possess these genes include monotremes (platypus) and marsupials (opossum) but not placental mammals. Genes for isocitrate lyase are found only in nematodes, in which, it is apparent, they originated in horizontal gene transfer from bacteria. The existence of glyoxylate cycles in humans has not been established, and it is widely held that fatty acids cannot be converted to glucose in humans directly. However, carbon-14 has been shown to end up in glucose when it is supplied in fatty acids.[9] Despite these findings, it is considered unlikely that the 2-carbon acetyl-CoA derived from the oxidation of fatty acids would produce a net yield of glucose via the citric acid cycle.[6] Glycerol, which is a part of the triacylglycerol molecule, can be used in gluconeogenesis.

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Location
In humans, gluconeogenesis is restricted to the liver and to a lesser extent the kidney.[10] In all species, the formation of oxaloacetate from pyruvate and TCA cycle intermediates is restricted to the mitochondrion, and the enzymes that convert PEP to glucose are found in the cytosol.[11] The location of the enzyme that links these two parts of gluconeogenesis by converting oxaloacetate to PEP, PEP carboxykinase, is variable by species: it can be found entirely within the mitochondria, entirely within the cytosol, or dispersed evenly between the two, as it is in humans.[11] Transport of PEP across the mitochondrial membrane is accomplished by dedicated transport proteins; however no such proteins exist for oxaloacetate.[11] Therefore species that lack intra-mitochondrial PEP, oxaloacetate must be converted into malate or asparate, exported from the mitochondrion, and converted back into oxaloacetate in order to allow gluconeogenesis to continue.[11]

Pathway
Gluconeogenesis is a pathway consisting of eleven enzyme-catalyzed reactions. The pathway can begin in the mitochondria or cytoplasm, depending on the substrate being used. Many of the reactions are the reversible steps found in glycolysis. Gluconeogenesis begins in the mitochondria with the formation of oxaloacetate through carboxylation of pyruvate. This reaction also requires one molecule of ATP, and is catalyzed by pyruvate carboxylase. This enzyme is stimulated by high levels of acetyl-CoA (produced in -oxidation in the liver) and inhibited by high levels of ADP. Oxaloacetate is reduced to malate using NADH, a step required for transport out of the mitochondria. Malate is oxidized to oxaloacetate using NAD+ in the cytoplasm, where the remaining steps of gluconeogenesis occur. Oxaloacetate is decarboxylated and phosphorylated to produce phosphoenolpyruvate by phosphoenolpyruvate carboxykinase. One molecule of GTP is hydrolyzed to GDP during this reaction. The next steps in the reaction are the same as reversed glycolysis. However, fructose-1,6-bisphosphatase converts fructose-1,6-bisphosphate to fructose 6-phosphate, requiring one water molecule and releasing one phosphate. This is also the rate-limiting step of gluconeogenesis. Glucose-6-phosphate is formed from fructose 6-phosphate by phosphoglucoisomerase. Glucose-6-phosphate can be used in other metabolic pathways or dephosphorylated to free glucose. Whereas free glucose can easily diffuse in and out of the cell, the phosphorylated form (glucose-6-phosphate) is locked in the cell, a mechanism by which

Gluconeogenesis intracellular glucose levels are controlled by cells. The final reaction of gluconeogenesis, the formation of glucose, occurs in the lumen of the endoplasmic reticulum, where glucose-6-phosphate is hydrolyzed by glucose-6-phosphatase to produce glucose. Glucose is shuttled into the cytosol by glucose transporters located in the membrane of the endoplasmic reticulum.

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Regulation
While most steps in gluconeogenesis are the reverse of those found in glycolysis, three regulated and strongly exergonic reactions are replaced with more kinetically favorable reactions. Hexokinase/glucokinase, phosphofructokinase, and pyruvate kinase enzymes of glycolysis are replaced with glucose-6-phosphatase, fructose-1,6-bisphosphatase, and PEP carboxykinase. This system of reciprocal control allow glycolysis and gluconeogenesis to inhibit each other and prevent the formation of a futile cycle. The majority of the enzymes responsible for gluconeogenesis are found in the cytoplasm; the exceptions are mitochondrial pyruvate carboxylase and, in animals, phosphoenolpyruvate carboxykinase. The latter exists as an isozyme located in both the mitochondrion and the cytosol.[12] The rate of gluconeogenesis is ultimately controlled by the action of a key enzyme, fructose-1,6-bisphosphatase, which is also regulated through signal transduction by cAMP and its phosphorylation. Most factors that regulate the activity of the gluconeogenesis pathway do so by inhibiting the activity or expression of key enzymes. However, both acetyl CoA and citrate activate gluconeogenesis enzymes (pyruvate carboxylase and fructose-1,6-bisphosphatase, respectively). Due to the reciprocal control of the cycle, acetyl-CoA and citrate also have inhibitory roles in the activity of pyruvate kinase. Global control of gluconeogenesis is mediated by glucagon (released when blood glucose is low); it triggers phosphorylation of enzymes and regulatory proteins by Protein Kinase A (a cyclic AMP regulated kinase) resulting in inhibition of glycolysis and stimulation of gluconeogenesis, thus bringing blood glucose levels up.[13]

References
[1] Silva, Pedro. "The Chemical Logic Behind Gluconeogenesis" (http:/ / www2. ufp. pt/ ~pedros/ bq/ gng. htm). . Retrieved September 8, 2009. [2] David L Nelson and Michael M Cox (2000). Lehninger Principles of Biochemistry. USA: Worth Publishers. pp.724. ISBN1-57259-153-6. [3] Hundal R, Krssak M, Dufour S, Laurent D, Lebon V, Chandramouli V, Inzucchi S, Schumann W, Petersen K, Landau B, Shulman G (2000). "Mechanism by Which Metformin Reduces Glucose Production in Type 2 Diabetes". Diabetes 49 (12): 20639. doi:10.2337/diabetes.49.12.2063. PMC2995498. PMID11118008. Free full text (http:/ / diabetes. diabetesjournals. org/ cgi/ reprint/ 49/ 12/ 2063)PDF(82KiB) [4] Garrett, Reginald H.; Charles M. Grisham (2002). Principles of Biochemistry with a Human Focus. USA: Brooks/Cole, Thomson Learning. pp.578, 585. ISBN0-03-097369-4. [5] Chapter 20 (Amino Acid Degradation and Synthesis) in: Denise R., PhD. Ferrier. Lippincott's Illustrated Reviews: Biochemistry (Lippincott's Illustrated Reviews). Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-2265-9. [6] Figueiredo, Luis F., Stefan Schuster, Christoph Kaleta, David A. Fell (2009). "Can sugars be produced from fatty acids? A test case for pathway analysis tools". Bioinformatics 25 (1): 152158. doi:10.1093/bioinformatics/btn621. PMID19117076. [7] Liu, F., et al. (1995). "Bifunctional glyoxylate cycle protein of Caenorhabditis elegans: a developmentally regulated protein of intestine and muscle". Developmental Biology 169 (2): 399414. doi:10.1006/dbio.1995.1156. PMID7781887. [8] Fyodor A Kondrashov, Eugene V Koonin, Igor G Morgunov, Tatiana V Finogenova, Marie N Kondrashova (2006). "Evolution of glyoxylate cycle enzymes in Metazoa: evidence of multiple horizontal transfer events and pseudogene formation". Biology Direct 1: 31. doi:10.1186/1745-6150-1-31. PMC1630690. PMID17059607. [9] Weinman, E.O., et al. (1957). "Conversion of fatty acids to carbohydrate: application of isotopes to this problem and role of the Krebs cycle as a synthetic pathway". Physiol. Rev. 37 (2): 25272. PMID13441426. [10] Widmaier, Eric (2006). Vander's Human Physiology. McGraw Hill. pp.96. ISBN0-07-282741-6. [11] Voet, Donald; Judith Voet, Charlotte Pratt (2008). Fundamentals of Biochemistry. John Wiley & Sons Inc. p.556. ISBN978-0470-12930-2. [12] Chakravarty, K., Cassuto, H., Resef, L., & Hanson, R.W. (2005) Factors that control the tissue-specific transcription of the gene for phosphoenolpyruvate carboxykinase-C. Critical Reviews of Biochemistry and Molecular Biology, 40(3), 129-154. [13] http:/ / rpi. edu/ dept/ bcbp/ molbiochem/ MBWeb/ mb1/ part2/ gluconeo. htm - Gluconeogenesis, by Diwan. Cites no sources.

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External links
Overview at indstate.edu (http://themedicalbiochemistrypage.org/gluconeogenesis.html) Interactive diagram at uakron.edu (http://ull.chemistry.uakron.edu/Pathways/gluconeogenesis/index.html#) The chemical logic behind gluconeogenesis (http://homepage.ufp.pt/pedros/bq/gng.htm)

Glycogen
Glycogen is a molecule that serves as the secondary long-term energy storage in animal and fungal cells, with the primary energy stores being held in adipose tissue. Glycogen is made primarily by the liver and the muscles, but can also be made by glycogenesis within the brain and stomach.[2] Glycogen is the analogue of starch, a glucose polymer in plants, and is sometimes referred to as animal starch, having a similar structure to amylopectin but more extensively branched and compact than starch. Glycogen is a polymer of (14) glycosidic bonds linked, with (16)-linked branches. Glycogen is found in the form of granules in the cytosol/cytoplasm in many cell types, and plays an important role in the glucose cycle. Glycogen forms an energy reserve that can be quickly mobilized to meet a sudden need for glucose, but one that is less compact than the energy reserves of triglycerides (lipids).

Schematic 2-D cross-sectional view of glycogen. A core protein of glycogenin is surrounded by branches of glucose units. The entire globular granule may contain [1] approximately 30,000 glucose units.

In the liver hepatocytes, glycogen can compose up to eight percent of the fresh weight (100120g in an adult) soon after a meal.[3] Only the glycogen stored in the liver can be made accessible to other organs. In the muscles, glycogen is found in a low concentration (one to two percent of the muscle mass). However, the amount of glycogen stored in the bodyespecially within the muscles, liver, and red blood cells[4] [5] [6] mostly depends on physical training, basal metabolic rate, and eating

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habits such as intermittent fasting. Small amounts of glycogen are found in the kidneys, and even smaller amounts in certain glial cells in the brain and white blood cells. The uterus also stores glycogen during pregnancy to nourish the embryo.[7]

Function and regulation of liver glycogen

As a meal containing carbohydrates is eaten and digested, blood glucose levels rise, and the pancreas secretes insulin. Glucose from the portal vein enters liver cells (hepatocytes). Insulin acts on the hepatocytes to stimulate the action of several enzymes, including glycogen synthase. Glucose molecules are added to the chains of glycogen as long as both insulin and glucose remain plentiful. In this postprandial or "fed" state, the liver takes in more glucose from the blood than it releases.

A view of the atomic structure of a single branched strand of glucose units in a glycogen molecule.

After a meal has been digested and glucose levels begin to fall, insulin secretion is reduced, and glycogen synthesis stops. When it is needed for energy, glycogen is broken down and converted again to glucose. Glycogen phosphorylase is the primary enzyme of glycogen breakdown. For the next 812 hours, glucose derived from liver glycogen will be the primary source of blood glucose to be used by the rest of the body for fuel. Glucagon is another hormone produced by the pancreas, which in many respects serves as a counter-signal to insulin. In response to insulin level below normal (when blood levels of glucose begin to fall below the normal range), glucagon is secreted in increasing amounts to stimulate glycogenolysis and gluconeogenesis pathways.

In muscle and other cells

Muscle cell glycogen appears to function as an immediate reserve source of available glucose for muscle cells. Other cells that contain small amounts use it locally as well. Muscle cells lack the enzyme glucose-6-phosphatase, which is required to pass glucose into the blood, so the glycogen they store is destined for internal use and is not shared with other cells. (This is in contrast to liver cells, which, on demand, readily do break down their stored glycogen into glucose and send it through the blood stream as fuel for the brain or muscles). Glycogen is also a suitable storage substance due to its insolubility in water, which means it does not affect the osmotistic levels and pressure of a cell.

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Glycogen depletion and endurance exercise

Long-distance athletes such as marathon runners, cross-country skiers, and cyclists often experience glycogen depletion, where almost all of the athlete's glycogen stores are depleted after long periods of exertion without enough energy consumption. This phenomenon is referred to as "hitting the wall". In marathon runners, it normally happens around the 20-mile (32km) point of a marathon, depending on the size of the runner and the race course. Glycogen depletion can be forestalled in four possible ways. First, during exercise carbohydrates with the highest possible rate of conversion to blood glucose per time (high Glycemic Index) are ingested continuously. The best possible outcome of this strategy replaces about 35% of glucose consumed at heart rates above about 80% of maximum. Second, through training, the body can be conditioned to burn fat earlier, faster, and more efficiently, sparing carbohydrate use from all sources. Third, by consuming foods low on the glycemic index for 1218 hours before the event, the liver and muscles will store the resulting slow but steady stream of glucose as glycogen, instead of fat. This process is known as carbohydrate loading. . When experiencing glycogen debt, athletes often experience extreme fatigue to the point that it is difficult to move. As a reference, the very best professional cyclists in the world will usually finish a 4-5hr stage race right at the limit of glycogen depletion using the first 3 strategies. A study published in the Journal of Applied Physiology (online May 8, 2008) suggests that, when athletes ingest both carbohydrate and caffeine following exhaustive exercise, their glycogen is replenished more rapidly.[8] [9]

Disorders of glycogen metabolism

The most common disease in which glycogen metabolism becomes abnormal is diabetes, in which, because of abnormal amounts of insulin, liver glycogen can be abnormally accumulated or depleted. Restoration of normal glucose metabolism usually normalizes glycogen metabolism as well. In hypoglycemia caused by excessive insulin, liver glycogen levels are high, but the high insulin level prevents the glycogenolysis necessary to maintain normal blood sugar levels. Glucagon is a common treatment for this type of hypoglycemia. Various inborn errors of metabolism are caused by deficiencies of enzymes necessary for glycogen synthesis or breakdown. These are collectively referred to as glycogen storage diseases.

Synthesis
Glycogen synthesis is, unlike its breakdown, endergonic. This means that glycogen synthesis requires the input of energy. Energy for glycogen synthesis comes from UTP, which reacts with glucose-1-phosphate, forming UDP-glucose, in a reaction catalysed by UDP-glucose pyrophosphorylase. Glycogen is synthesized from monomers of Glycogen Structure Segment UDP-glucose by the enzyme glycogen synthase, which progressively lengthens the glycogen chain with (14) bonded glucose. As glycogen synthase can lengthen only an existing chain, the protein glycogenin is needed to initiate the synthesis of glycogen. The glycogen-branching enzyme, amylo (14) to (16) transglycosylase, catalyzes the transfer of a terminal

Glycogen fragment of 6-7 glucose residues from a nonreducing end to the C-6 hydroxyl group of a glucose residue deeper into the interior of the glycogen molecule. The branching enzyme can act upon only a branch having at least 11 residues, and the enzyme may transfer to the same glucose chain or adjacent glucose chains.

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Breakdown
Glycogen is cleaved from the nonreducing ends of the chain by the enzyme glycogen phosphorylase to produce monomers of glucose-1-phosphate, which is then converted to glucose 6-phosphate by phosphoglucomutase. A special debranching enzyme is needed to remove the alpha(1-6) branches in branched glycogen and reshape the chain into linear polymer. The G6P monomers produced have three possible fates:

G6P can continue on the glycolysis pathway and be used as fuel. G6P can enter the pentose phosphate pathway via the enzyme Glucose-6-phosphate dehydrogenase to produce NADPH and 5-carbon sugars. In the liver and kidney, G6P can be dephosphorylated back to Glucose by the enzyme Glucose 6-phosphatase. This is the final step in the gluconeogenesis pathway.

References
[1] Page 12 in: (http:/ / books. google. dk/ books?id=SRptlOx7yj4C& printsec=frontcover& hl=en) Exercise physiology: energy, nutrition, and human performance By William D. McArdle, Frank I. Katch, Victor L. Katch Edition: 6, illustrated Published by Lippincott Williams & Wilkins, 2006 ISBN 0781749905, 9780781749909, 1068 pages [2] Anatomy and Physiology. Saladin, Kenneth S. McGraw-Hill, 2007. [3] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [4] Moses SW, Bashan N, Gutman A (December 1972). "Glycogen metabolism in the normal red blood cell" (http:/ / www. bloodjournal. org/ cgi/ pmidlookup?view=long& pmid=5083874). Blood 40 (6): 83643. PMID5083874. . [5] http:/ / jeb. biologists. org/ cgi/ reprint/ 129/ 1/ 141. pdf [6] Miwa I, Suzuki S (November 2002). "An improved quantitative assay of glycogen in erythrocytes". Annals of Clinical Biochemistry 39 (Pt 6): 6123. doi:10.1258/000456302760413432. PMID12564847. [7] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [8] Pedersen DJ, Lessard SJ, Coffey VG, et al. (July 2008). "High rates of muscle glycogen resynthesis after exhaustive exercise when carbohydrate is coingested with caffeine". Journal of Applied Physiology 105 (1): 713. doi:10.1152/japplphysiol.01121.2007. PMID18467543. [9] Post-exercise Caffeine Helps Muscles Refuel (http:/ / newswise. com/ articles/ view/ 542216/ ) Newswise, Retrieved on July 6, 2008.

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External links
Glycogen detection using Periodic Acid Schiff Staining (http://www.histochem.net/protocol periodic acid schiff.htm) Glycogen storage disease - McArdle's Disease Website (http://mcardlesdisease.org) MeSH Glycogen (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=&term=Glycogen)

Pentose phosphate pathway

The pentose phosphate pathway (also called the phosphogluconate pathway and the hexose monophosphate shunt) is a process that generates NADPH and pentoses (5-carbon sugars). There are two distinct phases in the pathway. The first is the oxidative phase, in which NADPH is generated, and the second is the non-oxidative synthesis of 5-carbon sugars. This pathway is an alternative to glycolysis. While it does involve oxidation of glucose, its primary role is anabolic rather than catabolic. For most organisms, it takes place in the cytosol; in plants, most steps take place in plastids.[1]

Outcome
The primary results of the Pathway are: The generation of reducing equivalents, in the form of NADPH, used in reductive biosynthesis reactions within cells. (e.g. fatty acid synthesis) Production of ribose-5-phosphate (R5P), used in the synthesis of nucleotides and nucleic acids. Production of erythrose-4-phosphate (E4P), used in the synthesis of aromatic amino acids. Aromatic amino acids, in turn, are precursors for many biosynthetic pathways, notably including the lignin in wood.
The Pentose Phosphate Pathway

Dietary pentose sugars derived from the digestion of nucleic acids may be metabolized through the pentose phosphate pathway, and the carbon skeletons of dietary carbohydrates may be converted into glycolytic/gluconeogenic intermediates. In mammals, the PPP occurs exclusively in the cytoplasm, and is found to be most active in the liver, mammary gland and adrenal cortex in the human. However, the pathway is absent in skeletal muscle tissue. The PPP is one of the three main ways the body creates molecules with reducing power, accounting for approximately 60% of NADPH production in humans. One of the uses of NADPH in the cell is to prevent oxidative stress. It reduces glutathione via glutathione reductase, which converts reactive H2O2 into H2O by glutathione peroxidase. If absent, the H2O2 would be converted to hydroxyl free radicals by Fenton chemistry, which can attack the cell. In a significant step, erythrocytes generate, through the pentose phosphate pathway, a large amount of NADPH used in the reduction of glutathione. Hydrogen peroxide is also generated for phagocytes in a process often referred to as a respiratory burst.[2]

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Phases
Oxidative phase
In this phase, two molecules of NADP+ are reduced to NADPH, utilizing the energy from the conversion of glucose-6-phosphate into ribulose 5-phosphate.

Oxidative phase of pentose phosphate pathway. glucose-6-phosphate (1), 6-phosphoglucono--lactone (2), 6-phosphogluconate (3), ribulose 5-phosphate (4).

The entire set of reactions can be summarized as follows:

Reactants Glucose 6-phosphate + NADP+ Products 6-phosphoglucono--lactone + NADPH Enzyme glucose 6-phosphate dehydrogenase Description Dehydrogenation. The hemiacetal hydroxyl group located on carbon 1 of glucose 6-phosphate is converted into a carbonyl group, generating a lactone, and, in the process, NADPH is generated. Hydrolysis

6-phosphoglucono--lactone + H2O 6-phosphogluconate + NADP+

6-phosphogluconate + H+ ribulose 5-phosphate + NADPH + CO2

6-phosphogluconolactonase

6-phosphogluconate dehydrogenase

Oxidative decarboxylation. NADP+ is the electron acceptor, generating another molecule of NADPH, a CO2, and ribulose 5-phosphate.

The overall reaction for this process is: Glucose 6-phosphate + 2 NADP+ + H2O ribulose 5-phosphate + 2 NADPH + 2 H+ + CO2

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Non-oxidative phase

The pentose phosphate pathway's Nonoxidative phase

Reactants ribulose 5-phosphate ribulose 5-phosphate

Products ribose 5-phosphate xylulose 5-phosphate

Enzymes Ribulose 5-Phosphate Isomerase Ribulose 5-Phosphate 3-Epimerase transketolase

xylulose 5-phosphate + ribose 5-phosphate

glyceraldehyde 3-phosphate + sedoheptulose 7-phosphate erythrose 4-phosphate + fructose 6-phosphate

sedoheptulose 7-phosphate + glyceraldehyde 3-phosphate xylulose 5-phosphate + erythrose 4-phosphate

transaldolase

glyceraldehyde 3-phosphate + fructose 6-phosphate

transketolase

Net reaction: 3 ribulose-5-phosphate 1 ribose-5-phosphate + 2 xylulose-5-phosphate 2 fructose-6-phosphate + glyceraldehyde-3-phosphate

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Regulation
Glucose-6-phosphate dehydrogenase is the rate-controlling enzyme of this pathway. It is allosterically stimulated by NADP+. The ratio of NADPH:NADP+ is normally about 100:1 in liver cytosol. This makes the cytosol a highly-reducing environment. An NADPH-utilizing pathway forms NADP+, which stimulates Glucose-6-phosphate dehydrogenase to produce more NADPH.

Erythrocytes and the pentose phosphate pathway

Carbohydrates are metabolized in red blood cells mainly by glycolysis, the pentose phosphate pathway (PPP), and 2,3-bisphosphoglycerate (2,3-BPG) metabolism (refer to discussion of hemoglobin for the role of 2,3-BPG). Glycolysis provides ATP for membrane ion pumps and NADH for reduction of methemoglobin. The PPP supplies the red blood cell with NADPH, which in turn maintains the reduced state of glutathione. The inability to maintain reduced glutathione in red blood cells leads to increased accumulation of peroxides, predominantly H2O2, that in turn results in a weakening of the cell membrane and concomitant hemolysis. Accumulation of H2O2 also leads to increased rates of oxidation of hemoglobin to methemoglobin that also weakens the cell wall. Glutathione removes peroxides via the action of glutathione peroxidase. The PPP in erythrocytes is, in essence, the only pathway for these cells to produce NADPH. Any defect in the production of NADPH could, therefore, have profound effects on erythrocyte survival. Several deficiencies in the level of activity (not function) of glucose-6-phosphate dehydrogenase have been observed to be associated with resistance to the malarial parasite Plasmodium falciparum among individuals of Mediterranean and African descent. The basis for this resistance may be a weakening of the red cell membrane (the erythrocyte is the host cell for the parasite) such that it cannot sustain the parasitic life cycle long enough for productive growth.[3]

References
[1] Kruger NJ, von Schaewen A (June 2003). "The oxidative pentose phosphate pathway: structure and organisation" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S1369526603000396). Curr. Opin. Plant Biol. 6 (3): 23646. doi:10.1016/S1369-5266(03)00039-6. PMID12753973. . [2] Immunology at MCG 1/cytotox (http:/ / www. lib. mcg. edu/ edu/ esimmuno/ ch1/ cytotox. htm) [3] Cappadoro M, Giribaldi G, O'Brien E, et al. (October 1998). "Early phagocytosis of glucose-6-phosphate dehydrogenase (G6PD)-deficient erythrocytes parasitized by Plasmodium falciparum may explain malaria protection in G6PD deficiency" (http:/ / bloodjournal. hematologylibrary. org/ cgi/ content/ full/ 92/ 7/ 2527). Blood 92 (7): 252734. PMID9746794. .

External links
The chemical logic behind the pentose phosphate pathway (http://www2.ufp.pt/~pedros/bq/ppp.htm) MeSH Pentose+Phosphate+Pathway (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=& term=Pentose+Phosphate+Pathway) Pentose phosphate pathway Map - Homo sapiens (http://www.genome.jp/dbget-bin/ www_bget?path:hsa00030)

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Citric acid cycle

Citric acid cycle
The citric acid cycle also known as the tricarboxylic acid cycle (TCA cycle), the Krebs cycle, or the Szent-Gyrgyi-Krebs cycle[1] [2] is a series of chemical reactions which is used by all aerobic living organisms to generate energy through the oxidization of acetate derived from carbohydrates, fats and proteins into carbon dioxide and water. In addition, the cycle provides precursors for the biosynthesis of compounds including certain amino acids as well as the reducing agent NADH that is used in numerous biochemical reactions. Its central importance to many Overview of the citric acid cycle biochemical pathways suggests that it was one of the earliest established components of cellular metabolism and may have originated abiogenically.[3] The name of this metabolic pathway is derived from citric acid (a type of tricarboxylic acid) which is first consumed and then regenerated by this sequence of reactions to complete the cycle. In addition, the cycle consumes acetate in the form of acetyl-CoA, reduces NAD+ to NADH, and produces carbon dioxide. The NADH generated by the TCA cycle is fed into the oxidative phosphorylation pathway. The net result of these two closely linked pathways is the oxidation of nutrients to produce energy in the form of ATP. In eukaryotic cells, the citric acid cycle occurs in the matrix of the mitochondrion. Bacteria also use the TCA cycle to generate energy, but since they lack mitochondria, the reaction sequence is performed in the cytosol. The components and reactions of the citric acid cycle were established in the 1930s by seminal work from the Nobel laureates Albert Szent-Gyrgyi[4] and Hans Adolf Krebs.[5]

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Evolution
Components of the TCA cycle were derived from anaerobic bacteria and the TCA cycle itself may have evolved more than once.[6] Theoretically there are several alternatives to the TCA cycle, however the TCA cycle appears to be the most efficient.[7] If several alternatives independently evolved, they all undoubtedly rapidly converged to the TCA cycle.

Overview
The citric acid cycle is a key component of the metabolic pathway by which all aerobic organisms generate energy. Through catabolism of sugars, fats, and proteins, a two carbon organic product acetate in the form of acetyl-CoA is produced. Acetyl-CoA along with two equivalents of water (H2O) are consumed by the citric acid cycle producing two equivalents of carbon dioxide (CO2) and one equivalent of HS-CoA. In addition, one complete turn of the cycle converts three equivalents of nicotinamide adenine dinucleotide (NAD+) into three equivalents of reduced NAD+ (NADH), one equivalent of ubiquinone (Q) into one equivalent of reduced ubiquinone (QH2), and one equivalent each of guanosine diphosphate (GDP) and inorganic phosphate (Pi) into one equivalent of guanosine triphosphate (GTP). The NADH and QH2 that is generated by the citric acid cycle is in turn used by the oxidative phosphorylation pathway to generate energy rich adenosine triphosphate (ATP). One of the primary sources of acetyl-CoA are sugars that are broken down by glycolysis to produce pyruvate that in turn is decarboxylated by the enzyme pyruvate dehydrogenase generating acetyl-CoA according to the following reaction scheme: CH3C(=O)C(=O)O (pyruvate) + HSCoA + NAD+ CH3C(=O)SCoA (acetyl-CoA) + NADH + H+ + CO2 The product of this reaction, acetyl-CoA, is the starting point for the citric acid cycle. Below is a schematic outline of the cycle: The citric acid cycle begins with the transfer of a two-carbon acetyl group from acetyl-CoA to the four-carbon acceptor compound (oxaloacetate) to form a six-carbon compound (citrate). The citrate then goes through a series of chemical transformations, losing two carboxyl groups as CO2. The carbons lost as CO2 originate from what was oxaloacetate, not directly from acetyl-CoA. The carbons donated by acetyl-CoA become part of the oxaloacetate carbon backbone after the first turn of the citric acid cycle. Loss of the acetyl-CoA-donated carbons as CO2 requires several turns of the citric acid cycle. However, because of the role of the citric acid cycle in anabolism, they may not be lost, since many TCA cycle intermediates are also used as precursors for the biosynthesis of other molecules.[8] Most of the energy made available by the oxidative steps of the cycle is transferred as energy-rich electrons to NAD+, forming NADH. For each acetyl group that enters the citric acid cycle, three molecules of NADH are produced. Electrons are also transferred to the electron acceptor Q, forming QH2. At the end of each cycle, the four-carbon oxaloacetate has been regenerated, and the cycle continues.

Steps
Two carbon atoms are oxidized to CO2, the energy from these reactions being transferred to other metabolic processes by GTP (or ATP), and as electrons in NADH and QH2. The NADH generated in the TCA cycle may later donate its electrons in oxidative phosphorylation to drive ATP synthesis; FADH2 is covalently attached to succinate dehydrogenase, an enzyme functioning both in the TCA cycle and the mitochondrial electron transport chain in oxidative phosphorylation. FADH2, therefore, facilitates transfer of electrons to coenzyme Q, which is the final electron acceptor of the reaction catalyzed by the Succinate:ubiquinone oxidoreductase complex, also acting as an intermediate in the electron transport chain.[9]

Citric acid cycle The citric acid cycle is continuously supplied with new carbon in the form of acetyl-CoA, entering at step 1 below.[10]
Substrates 1 Oxaloacetate + Acetyl CoA + H2O Citrate Products Citrate + CoA-SH Enzyme Citrate synthase Reaction type Aldol condensation Comment irreversible, extends the 4C oxaloacetate to a 6C molecule

247

cis-Aconitate + H2O Isocitrate

Aconitase

Dehydration

reversible isomerisation

cis-Aconitate + H2O Isocitrate + NAD+

Hydration

Oxalosuccinate + NADH + H + -Ketoglutarate + CO2

Isocitrate dehydrogenase

Oxidation

generates NADH (equivalent of 2.5 ATP)

Oxalosuccinate

Decarboxylation

rate-limiting, irreversible stage, generates a 5C molecule

-Ketoglutarate + NAD+ + CoA-SH

Succinyl-CoA + -Ketoglutarate dehydrogenase NADH + H+ + CO2 Succinyl-CoA synthetase

Oxidative decarboxylation

irreversible stage, generates NADH (equivalent of 2.5 ATP), regenerates the 4C chain (CoA excluded)

Succinyl-CoA + Succinate + GDP + Pi CoA-SH + GTP Succinate + ubiquinone (Q)

substrate-level phosphorylation

or ADPATP instead of GDPGTP, generates 1 ATP or equivalent

[9]

Fumarate + Succinate ubiquinol (QH2) dehydrogenase

Oxidation

uses FAD as a prosthetic group (FADFADH2 in the [9] first step of the reaction) in the enzyme, generates the equivalent of 1.5 ATP

Fumarate + H2O

L-Malate

Fumarase

Hydration

10 L-Malate + NAD+

Oxaloacetate + NADH + H+

Malate dehydrogenase

Oxidation

reversible (in fact, equilibrium favors malate), generates NADH (equivalent of 2.5 ATP)

Mitochondria in animals, including humans, possess two succinyl-CoA synthetases: one that produces GTP from GDP, and another that produces ATP from ADP.[11] Plants have the type that produces ATP (ADP-forming succinyl-CoA synthetase).[10] Several of the enzymes in the cycle may be loosely-associated in a multienzyme protein complex within the mitochondrial matrix.[12] The GTP that is formed by GDP-forming succinyl-CoA synthetase may be utilized by nucleoside-diphosphate kinase to form ATP (the catalyzed reaction is GTP + ADP GDP + ATP).[9]

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Products
Products of the first turn of the cycle are: one GTP (or ATP), three NADH, one QH2, two CO2. Because two acetyl-CoA molecules are produced from each glucose molecule, two cycles are required per glucose molecule. Therefore, at the end of two cycles, the products are: two GTP, six NADH, two QH2, and four CO2
Description The sum of all reactions in the citric acid cycle is: Reactants Acetyl-CoA + 3 NAD+ + Q + GDP + Pi + 2 H2O Pyruvate ion + 4 NAD+ + Q + GDP + Pi + 2 H2O Glucose + 10 NAD+ + 2 Q + 2 ADP + 2 GDP + 4 Pi + 2 H2O Products CoA-SH + 3 NADH + 3 H+ + QH2 + GTP + 2 CO2 4 NADH + 4 H+ + QH2 + GTP + 3 CO2 10 NADH + 10 H+ + 2 QH2 + 2 ATP + 2 GTP + 6 CO2

Combining the reactions occurring during the pyruvate oxidation with those occurring during the citric acid cycle, the following overall pyruvate oxidation reaction is obtained: Combining the above reaction with the ones occurring in the course of glycolysis, the following overall glucose oxidation reaction (excluding reactions in the respiratory chain) is obtained:

The above reactions are balanced if Pi represents the H2PO4- ion, ADP and GDP the ADP2- and GDP2- ions, respectively, and ATP and GTP the ATP3- and GTP3- ions, respectively. The total number of ATP obtained after complete oxidation of one glucose in glycolysis, citric acid cycle, and oxidative phosphorylation is estimated to be between 30 and 38. A recent assessment of the total ATP yield with the updated proton-to-ATP ratios provides an estimate of 29.85 ATP per glucose molecule.[13]

Regulation
Although pyruvate dehydrogenase is not technically a part of the citric acid cycle, its regulation is included here. The regulation of the TCA cycle is largely determined by substrate availability and product inhibition. NADH, a product of all dehydrogenases in the TCA cycle with the exception of succinate dehydrogenase, inhibits pyruvate dehydrogenase, isocitrate dehydrogenase, -ketoglutarate dehydrogenase, and also citrate synthase. Acetyl-coA inhibits pyruvate dehydrogenase, while succinyl-CoA inhibits succinyl-CoA synthetase and citrate synthase. When tested in vitro with TCA enzymes, ATP inhibits citrate synthase and -ketoglutarate dehydrogenase; however, ATP levels do not change more than 10% in vivo between rest and vigorous exercise. There is no known allosteric mechanism that can account for large changes in reaction rate from an allosteric effector whose concentration changes less than 10%.[14] Calcium is used as a regulator. It activates pyruvate dehydrogenase, isocitrate dehydrogenase and -ketoglutarate dehydrogenase.[15] This increases the reaction rate of many of the steps in the cycle, and therefore increases flux throughout the pathway. Citrate is used for feedback inhibition, as it inhibits phosphofructokinase, an enzyme involved in glycolysis that catalyses formation of fructose 1,6-bisphosphate,a precursor of pyruvate. This prevents a constant high rate of flux when there is an accumulation of citrate and a decrease in substrate for the enzyme. Recent work has demonstrated an important link between intermediates of the citric acid cycle and the regulation of hypoxia-inducible factors (HIF). HIF plays a role in the regulation of oxygen homeostasis, and is a transcription factor that targets angiogenesis, vascular remodeling, glucose utilization, iron transport and apoptosis. HIF is synthesized consititutively, and hydroxylation of at least one of two critical proline residues mediates their interaction with the von Hippel Lindau E3 ubiquitin ligase complex, which targets them for rapid degradation. This reaction is catalysed by prolyl 4-hydroxylases. Fumarate and succinate have been identified as potent inhibitors of prolyl hydroxylases, thus leading to the stabilisation of HIF.[16]

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Major metabolic pathways converging on the TCA cycle

Several catabolic pathways converge on the TCA cycle. Reactions that form intermediates of the TCA cycle in order to replenish them (especially during the scarcity of the intermediates) are called anaplerotic reactions. The citric acid cycle is the third step in carbohydrate catabolism (the breakdown of sugars). Glycolysis breaks glucose (a six-carbon-molecule) down into pyruvate (a three-carbon molecule). In eukaryotes, pyruvate moves into the mitochondria. It is converted into acetyl-CoA by decarboxylation and enters the citric acid cycle. In protein catabolism, proteins are broken down by proteases into their constituent amino acids. The carbon backbone of these amino acids can become a source of energy by being converted to acetyl-CoA and entering into the citric acid cycle. In fat catabolism, triglycerides are hydrolyzed to break them into fatty acids and glycerol. In the liver the glycerol can be converted into glucose via dihydroxyacetone phosphate and glyceraldehyde-3-phosphate by way of gluconeogenesis. In many tissues, especially heart tissue, fatty acids are broken down through a process known as beta oxidation, which results in acetyl-CoA, which can be used in the citric acid cycle. Beta oxidation of fatty acids with an odd number of methylene groups produces propionyl CoA, which is then converted into succinyl-CoA and fed into the citric acid cycle.[17] The total energy gained from the complete breakdown of one molecule of glucose by glycolysis, the citric acid cycle, and oxidative phosphorylation equals about 30 ATP molecules, in eukaryotes. The citric acid cycle is called an amphibolic pathway because it participates in both catabolism and anabolism.

Interactive pathway map

Click on genes, proteins and metabolites below to link to respective articles.[18] [[File: <div style="display:block; width:60px; height:0px; overflow:hidden; position:relative; left:627.0px; top:127.333333333333px; background:transparent; border-top:3px blue solid"></div> <div style="display:block; width:104.333333333333px; height:0px; overflow:hidden; position:relative; left:178.0px; top:420px; background:transparent; border-top:3px blue solid"></div> <div style="display:block; width:60px; height:0px; overflow:hidden; position:relative; left:219.66666666666669px; top:535.333333333333px; background:transparent; border-top:3px blue solid"></div> <div style="display:block; width:71.0666666666666px; height:0px; overflow:hidden; position:relative; left:697.9333333333334px; top:367.166666666667px; background:transparent; border-top:3px blue solid"></div> <div style="display:block; width:60px; height:0px; overflow:hidden; position:relative; left:778.3332722981771px; top:208.666666666667px; background:transparent; border-top:3px red solid"></div> <div style="display:block; width:69.5333333333333px; height:0px; overflow:hidden; position:relative; left:633.4666666666667px; top:633.166666666667px; background:transparent; border-top:3px blue solid"></div> <div style="display:block; width:60px; height:0px; overflow:hidden; position:relative; top:150.333333333333px; background:transparent; border-top:3px blue solid"></div> left:512.0px;

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References
[1] Lowenstein JM (1969). Methods in Enzymology, Volume 13: Citric Acid Cycle. Boston: Academic Press. ISBN0-12-181870-5. [2] Krebs HA, Weitzman PDJ (1987). Krebs' citric acid cycle: half a century and still turning. London: Biochemical Society. ISBN0-904498-22-0. [3] Lane, Nick (2009). Life Ascending: The Ten Great Inventions of Evolution. New York: W.W. Norton & Co. ISBN0-393-06596-0. [4] "The Nobel Prize in Physiology or Medicine 1937" (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1937/ ). The Nobel Foundation. . Retrieved 2011-10-26. [5] "The Nobel Prize in Physiology or Medicine 1953" (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1953/ ). The Nobel Foundation. . Retrieved 2011-10-26. [6] Gest H (1987). "Evolutionary roots of the citric acid cycle in prokaryotes". Biochem. Soc. Symp. 54: 316. PMID3332996. [7] Melndez-Hevia E, Waddell TG, Cascante M (September 1996). "The puzzle of the Krebs citric acid cycle: assembling the pieces of chemically feasible reactions, and opportunism in the design of metabolic pathways during evolution". J. Mol. Evol. 43 (3): 293303. doi:10.1007/BF02338838. PMID8703096. [8] Wolfe RR, Jahoor F (February 1990). "Recovery of labeled CO2 during the infusion of C-1- vs C-2-labeled acetate: implications for tracer studies of substrate oxidation". Am. J. Clin. Nutr. 51 (2): 24852. PMID2106256. [9] Stryer L, Berg J, Tymoczko JL (2002). Biochemistry. San Francisco: W.H. Freeman. ISBN0-7167-4684-0. [10] Jones RC, Buchanan BB, Gruissem W (2000). Biochemistry & molecular biology of plants (1st ed.). Rockville, Md: American Society of Plant Physiologists. ISBN0-943088-39-9. [11] Johnson JD, Mehus JG, Tews K, Milavetz BI, Lambeth DO (October 1998). "Genetic evidence for the expression of ATP- and GTP-specific succinyl-CoA synthetases in multicellular eucaryotes". J. Biol. Chem. 273 (42): 275806. doi:10.1074/jbc.273.42.27580. PMID9765291. [12] Barnes SJ, Weitzman PD (June 1986). "Organization of citric acid cycle enzymes into a multienzyme cluster". FEBS Lett. 201 (2): 26770. doi:10.1016/0014-5793(86)80621-4. PMID3086126. [13] Rich PR (December 2003). "The molecular machinery of Keilin's respiratory chain". Biochem. Soc. Trans. 31 (Pt 6): 1095105. doi:10.1042/BST0311095. PMID14641005. [14] Voet D, Voet JG (2004). Biochemistry (3rd ed.). New York: John Wiley & Sons, Inc.. p.615. [15] Denton RM, Randle PJ, Bridges BJ, Cooper RH, Kerbey AL, Pask HT, Severson DL, Stansbie D, Whitehouse S (October 1975). "Regulation of mammalian pyruvate dehydrogenase". Mol. Cell. Biochem. 9 (1): 2753. doi:10.1007/BF01731731. PMID171557. [16] Koivunen P, Hirsil M, Remes AM, Hassinen IE, Kivirikko KI, Myllyharju J (February 2007). "Inhibition of hypoxia-inducible factor (HIF) hydroxylases by citric acid cycle intermediates: possible links between cell metabolism and stabilization of HIF". J. Biol. Chem. 282 (7): 452432. doi:10.1074/jbc.M610415200. PMID17182618. [17] Halarnkar PP, Blomquist GJ (1989). "Comparative aspects of propionate metabolism". Comp. Biochem. Physiol., B 92 (2): 22731. doi:10.1016/0305-0491(89)90270-8. PMID2647392. [18] The interactive pathway map can be edited at WikiPathways: "TCACycle_WP78" (http:/ / www. wikipathways. org/ index. php/ Pathway:WP78). . [19] http:/ / www. wikipathways. org/ index. php/ Pathway:WP78

External links
Citric acid cycle Animation (http://www.1lec.com/Biochemistry/How the Krebs Cycle Works/index. html)(flash required) An animation of the citric acid cycle (http://www.science.smith.edu/departments/Biology/Bio231/krebs. html) at Smith College Notes on citric acid cycle (http://www.rahulgladwin.com/blog/2007/01/notes-on-citric-acid-cycle-glyoxylate. html) at rahulgladwin.com Citric acid cycle variants (http://biocyc.org/META/NEW-IMAGE?object=TCA-VARIANTS) at MetaCyc Pathways connected to the citric acid cycle (http://www.genome.ad.jp/kegg/pathway/map/map00020.html) at Kyoto Encyclopedia of Genes and Genomes A more detailed tutorial animation (http://www.johnkyrk.com/krebs.html) at johnkyrk.com A citric-acid cycle self quiz flash applet (http://www.pitt.edu/AFShome/j/b/jbrodsky/public/html/1820/tca. htm) at University of Pittsburgh The chemical logic behind the citric acid cycle (http://www2.ufp.pt/~pedros/bq/tca.htm) The Krebs cycle song (http://www.science-groove.org/Now/) by Lynda Jones.

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Oxidative phosphorylation
Oxidative phosphorylation
Oxidative phosphorylation (or OXPHOS in short) is a metabolic pathway that uses energy released by the oxidation of nutrients to produce adenosine triphosphate (ATP). Although the many forms of life on earth use a range of different nutrients, almost all aerobic organisms carry out oxidative phosphorylation to produce ATP, the molecule that supplies energy to metabolism. This pathway is probably so pervasive because it is a highly efficient way of releasing energy, compared to alternative fermentation processes such as anaerobic glycolysis. During oxidative phosphorylation, electrons are transferred from electron The electron transport chain in the mitochondrion is the site of oxidative phosphorylation donors to electron acceptors such as in eukaryotes. The NADH and succinate generated in the citric acid cycle are oxidized, oxygen, in redox reactions. These releasing energy to power the ATP synthase. redox reactions release energy, which is used to form ATP. In eukaryotes, these redox reactions are carried out by a series of protein complexes within mitochondria, whereas, in prokaryotes, these proteins are located in the cells' inner membranes. These linked sets of proteins are called electron transport chains. In eukaryotes, five main protein complexes are involved, whereas in prokaryotes many different enzymes are present, using a variety of electron donors and acceptors. The energy released by electrons flowing through this electron transport chain is used to transport protons across the inner mitochondrial membrane, in a process called chemiosmosis. This generates potential energy in the form of a pH gradient and an electrical potential across this membrane. This store of energy is tapped by allowing protons to flow back across the membrane and down this gradient, through a large enzyme called ATP synthase. This enzyme uses this energy to generate ATP from adenosine diphosphate (ADP), in a phosphorylation reaction. This reaction is driven by the proton flow, which forces the rotation of a part of the enzyme; the ATP synthase is a rotary mechanical motor. Although oxidative phosphorylation is a vital part of metabolism, it produces reactive oxygen species such as superoxide and hydrogen peroxide, which lead to propagation of free radicals, damaging cells and contributing to disease and, possibly, aging (senescence). The enzymes carrying out this metabolic pathway are also the target of many drugs and poisons that inhibit their activities.

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Overview of energy transfer by chemiosmosis

Further information: Chemiosmosis and Bioenergetics Oxidative phosphorylation works by using energy-releasing chemical reactions to drive energy-requiring reactions: The two sets of reactions are said to be coupled. This means one cannot occur without the other. The flow of electrons through the electron transport chain, from electron donors such as NADH to electron acceptors such as oxygen, is an exergonic process it releases energy, whereas the synthesis of ATP is an endergonic process, which requires an input of energy. Both the electron transport chain and the ATP synthase are embedded in a membrane, and energy is transferred from electron transport chain to the ATP synthase by movements of protons across this membrane, in a process called chemiosmosis.[1] In practice, this is like a simple electric circuit, with a current of protons being driven from the negative N-side of the membrane to the positive P-side by the proton-pumping enzymes of the electron transport chain. These enzymes are like a battery, as they perform work to drive current through the circuit. The movement of protons creates an electrochemical gradient across the membrane, which is often called the proton-motive force. This gradient has two components: a difference in proton concentration (a H+ gradient) and a difference in electric potential, with the N-side having a negative charge. The energy is stored largely as the difference of electric potentials in mitochondria, but also as a pH gradient in chloroplasts.[2] ATP synthase releases this stored energy by completing the circuit and allowing protons to flow down the electrochemical gradient, back to the N-side of the membrane.[3] This enzyme is like an electric motor as it uses the proton-motive force to drive the rotation of part of its structure and couples this motion to the synthesis of ATP. The amount of energy released by oxidative phosphorylation is high, compared with the amount produced by anaerobic fermentation. Glycolysis produces only 2 ATP molecules, but somewhere between 30 and 36 ATPs are produced by the oxidative phosphorylation of the 10 NADH and 2 succinate molecules made by converting one molecule of glucose to carbon dioxide and water,[4] while each cycle of beta oxidation of a fatty acid yields about 14 ATPs. These ATP yields are theoretical maximum values; in practice, some protons leak across the membrane, lowering the yield of ATP.[5]

Electron and proton transfer molecules

Further information: Coenzyme and Cofactor The electron transport chain carries both protons and electrons, passing electrons from donors to acceptors, and transporting protons across a membrane. These processes use both soluble and protein-bound transfer molecules. In mitochondria, electrons are transferred within the intermembrane space by the water-soluble electron transfer protein cytochrome c.[6] This carries only electrons, and these are transferred by the reduction and oxidation of an iron atom that the protein holds within a heme group in its structure. Cytochrome c is also found in some bacteria, where it is located within the periplasmic space.[7]

Within the inner mitochondrial membrane, the lipid-soluble electron carrier coenzyme Q10 (Q) carries [8] both electrons and protons by a redox cycle. This small benzoquinone molecule is very hydrophobic, so it diffuses freely within the membrane. When Q accepts two electrons and two protons, it becomes reduced to the ubiquinol form (QH2); when QH2 releases two electrons and two protons, it becomes oxidized back to the ubiquinone (Q) form. As a result, if two enzymes are arranged so that Q is reduced on one side of the membrane and QH2 oxidized

Reduction of coenzyme Q from its ubiquinone form (Q) to the reduced ubiquinol form (QH2).

Oxidative phosphorylation on the other, ubiquinone will couple these reactions and shuttle protons across the membrane.[9] Some bacterial electron transport chains use different quinones, such as menaquinone, in addition to ubiquinone.[10] Within proteins, electrons are transferred between flavin cofactors,[3] [11] ironsulfur clusters, and cytochromes. There are several types of ironsulfur cluster. The simplest kind found in the electron transfer chain consists of two iron atoms joined by two atoms of inorganic sulfur; these are called [2Fe2S] clusters. The second kind, called [4Fe4S], contains a cube of four iron atoms and four sulfur atoms. Each iron atom in these clusters is coordinated by an additional amino acid, usually by the sulfur atom of cysteine. Metal ion cofactors undergo redox reactions without binding or releasing protons, so in the electron transport chain they serve solely to transport electrons through proteins. Electrons move quite long distances through proteins by hopping along chains of these cofactors.[12] This occurs by quantum tunnelling, which is rapid over distances of less than 1.4109 m.[13]

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Eukaryotic electron transport chains

Further information: Electron transport chain and Chemiosmosis Many catabolic biochemical processes, such as glycolysis, the citric acid cycle, and beta oxidation, produce the reduced coenzyme NADH. This coenzyme contains electrons that have a high transfer potential; in other words, they will release a large amount of energy upon oxidation. However, the cell does not release this energy all at once, as this would be an uncontrollable reaction. Instead, the electrons are removed from NADH and passed to oxygen through a series of enzymes that each release a small amount of the energy. This set of enzymes, consisting of complexes I through IV, is called the electron transport chain and is found in the inner membrane of the mitochondrion. Succinate is also oxidized by the electron transport chain, but feeds into the pathway at a different point. In eukaryotes, the enzymes in this electron transport system use the energy released from the oxidation of NADH to pump protons across the inner membrane of the mitochondrion. This causes protons to build up in the intermembrane space, and generates an electrochemical gradient across the membrane. The energy stored in this potential is then used by ATP synthase to produce ATP. Oxidative phosphorylation in the eukaryotic mitochondrion is the best-understood example of this process. The mitochondrion is present in almost all eukaryotes, with the exception of anaerobic protozoa such as Trichomonas vaginalis that instead reduce protons to hydrogen in a remnant mitochondrion called a hydrogenosome.[14]

Typical respiratory enzymes and substrates in eukaryotes.

Respiratory enzyme NADH dehydrogenase Succinate dehydrogenase Redox pair NAD+ / NADH FMN or FAD / FMNH2 or FADH2 Midpoint potential (Volts) 0.32 0.20 +0.06 +0.12 +0.22 +0.29 +0.82 [15] [15]

Cytochrome bc1 complex Coenzyme Q10ox / Coenzyme Q10red Cytochrome bc1 complex Complex IV Complex IV Complex IV Conditions: pH = 7 [15] Cytochrome box / Cytochrome bred Cytochrome cox / Cytochrome cred Cytochrome aox / Cytochrome ared O2 / HO

[15] [15] [15] [15] [15]

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NADH-coenzyme Q oxidoreductase (complex I)

NADH-coenzyme Q oxidoreductase, also known as NADH dehydrogenase or complex I, is the first protein in the electron transport chain.[16] Complex I is a giant enzyme with the mammalian complex I having 46 subunits and a molecular mass of about 1,000 kilodaltons (kDa).[17] The structure is known in detail only from a bacterium;[18] [19] in most organisms the complex resembles a boot with a large ball poking out from the membrane into the mitochondrion.[20] [21] The genes that encode the individual proteins are contained in both the cell nucleus and the mitochondrial genome, as is the case for many enzymes present in the mitochondrion.

Complex I or NADH-Q oxidoreductase. The abbreviations are discussed in the text. In all diagrams of respiratory complexes in this article, the matrix is at the bottom, with the intermembrane space above.

The reaction that is catalyzed by this enzyme is the two electron reduction by NADH of coenzyme Q10 or ubiquinone (represented as Q in the equation below), a lipid-soluble quinone that is found in the mitochondrion membrane:

The start of the reaction, and indeed of the entire electron chain, is the binding of a NADH molecule to complex I and the donation of two electrons. The electrons enter complex I via a prosthetic group attached to the complex, flavin mononucleotide (FMN). The addition of electrons to FMN converts it to its reduced form, FMNH2. The electrons are then transferred through a series of ironsulfur clusters: the second kind of prosthetic group present in the complex.[18] There are both [2Fe2S] and [4Fe4S] ironsulfur clusters in complex I. As the electrons pass through this complex, four protons are pumped from the matrix into the intermembrane space. Exactly how this occurs is unclear, but it seems to involve conformational changes in complex I that cause the protein to bind protons on the N-side of the membrane and release them on the P-side of the membrane.[22] Finally, the electrons are transferred from the chain of ironsulfur clusters to a ubiquinone molecule in the membrane.[16] Reduction of ubiquinone also contributes to the generation of a proton gradient, as two protons are taken up from the matrix as it is reduced to ubiquinol (QH2).

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Succinate-Q oxidoreductase (complex II)

Succinate-Q oxidoreductase, also known as complex II or succinate dehydrogenase, is a second entry point to the electron transport chain.[23] It is unusual because it is the only enzyme that is part of both the citric acid cycle and the electron transport chain. Complex II consists of four protein subunits and contains a bound flavin adenine dinucleotide (FAD) cofactor, ironsulfur clusters, and a heme group that does not participate in electron transfer to coenzyme Q, but is believed to be important in decreasing production of reactive oxygen species.[24] [25] It oxidizes succinate to fumarate and reduces ubiquinone. As this reaction releases less energy than the oxidation of NADH, complex II does not transport protons across the membrane and does not contribute to the proton gradient.
Complex II: Succinate-Q oxidoreductase.

In some eukaryotes, such as the parasitic worm Ascaris suum, an enzyme similar to complex II, fumarate reductase (menaquinol:fumarate oxidoreductase, or QFR), operates in reverse to oxidize ubiquinol and reduce fumarate. This allows the worm to survive in the anaerobic environment of the large intestine, carrying out anaerobic oxidative phosphorylation with fumarate as the electron acceptor.[26] Another unconventional function of complex II is seen in the malaria parasite Plasmodium falciparum. Here, the reversed action of complex II as an oxidase is important in regenerating ubiquinol, which the parasite uses in an unusual form of pyrimidine biosynthesis.[27]

Electron transfer flavoprotein-Q oxidoreductase

Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-Q oxidoreductase), also known as electron transferring-flavoprotein dehydrogenase, is a third entry point to the electron transport chain. It is an enzyme that accepts electrons from electron-transferring flavoprotein in the mitochondrial matrix, and uses these electrons to reduce ubiquinone.[28] This enzyme contains a flavin and a [4Fe4S] cluster, but, unlike the other respiratory complexes, it attaches to the surface of the membrane and does not cross the lipid bilayer.[29]

In mammals, this metabolic pathway is important in beta oxidation of fatty acids and catabolism of amino acids and choline, as it accepts electrons from multiple acetyl-CoA dehydrogenases.[30] [31] In plants, ETF-Q oxidoreductase is also important in the metabolic responses that allow survival in extended periods of darkness.[32]

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Q-cytochrome c oxidoreductase (complex III)

Q-cytochrome c oxidoreductase is also known as cytochrome c reductase, cytochrome bc1 complex, or simply complex III.[33] [34] In mammals, this enzyme is a dimer, with each subunit complex containing 11 protein subunits, an [2Fe-2S] ironsulfur cluster and three cytochromes: one cytochrome c1 and two b [35] cytochromes. A cytochrome is a kind of electron-transferring protein The two electron transfer steps in complex III: Q-cytochrome c oxidoreductase. After that contains at least one heme group. each step, Q (in the upper part of the figure) leaves the enzyme. The iron atoms inside complex IIIs heme groups alternate between a reduced ferrous (+2) and oxidized ferric (+3) state as the electrons are transferred through the protein. The reaction catalyzed by complex III is the oxidation of one molecule of ubiquinol and the reduction of two molecules of cytochrome c, a heme protein loosely associated with the mitochondrion. Unlike coenzyme Q, which carries two electrons, cytochrome c carries only one electron.

As only one of the electrons can be transferred from the QH2 donor to a cytochrome c acceptor at a time, the reaction mechanism of complex III is more elaborate than those of the other respiratory complexes, and occurs in two steps called the Q cycle.[36] In the first step, the enzyme binds three substrates, first, QH2, which is then oxidized, with one electron being passed to the second substrate, cytochrome c. The two protons released from QH2 pass into the intermembrane space. The third substrate is Q, which accepts the second electron from the QH2 and is reduced to Q.-, which is the ubisemiquinone free radical. The first two substrates are released, but this ubisemiquinone intermediate remains bound. In the second step, a second molecule of QH2 is bound and again passes its first electron to a cytochrome c acceptor. The second electron is passed to the bound ubisemiquinone, reducing it to QH2 as it gains two protons from the mitochondrial matrix. This QH2 is then released from the enzyme.[37] As coenzyme Q is reduced to ubiquinol on the inner side of the membrane and oxidized to ubiquinone on the other, a net transfer of protons across the membrane occurs, adding to the proton gradient.[3] The rather complex two-step mechanism by which this occurs is important, as it increases the efficiency of proton transfer. If, instead of the Q cycle, one molecule of QH2 were used to directly reduce two molecules of cytochrome c, the efficiency would be halved, with only one proton transferred per cytochrome c reduced.[3]

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Cytochrome c oxidase (complex IV)

Cytochrome c oxidase, also known as complex IV, is the final protein complex in the electron transport chain.[38] The mammalian enzyme has an extremely complicated structure and contains 13 subunits, two heme groups, as well as multiple metal ion cofactors in all three atoms of copper, one of magnesium and one of zinc.[39] This enzyme mediates the final reaction in the electron transport chain and transfers electrons to oxygen, while pumping protons across the membrane.[40] The final electron acceptor oxygen, which is also called the terminal electron acceptor, is reduced to water in this step. Both the direct pumping of protons and the consumption of matrix protons in the reduction of oxygen contribute to the proton gradient. The reaction catalyzed is the oxidation of cytochrome c and the reduction of oxygen:

Complex IV: cytochrome c oxidase.

Alternative reductases and oxidases

Many eukaryotic organisms have electron transport chains that differ from the much-studied mammalian enzymes described above. For example, plants have alternative NADH oxidases, which oxidize NADH in the cytosol rather than in the mitochondrial matrix, and pass these electrons to the ubiquinone pool.[41] These enzymes do not transport protons, and, therefore, reduce ubiquinone without altering the electrochemical gradient across the inner membrane.[42] Another example of a divergent electron transport chain is the alternative oxidase, which is found in plants, as well as some fungi, protists, and possibly some animals.[43] [44] This enzyme transfers electrons directly from ubiquinol to oxygen.[45] The electron transport pathways produced by these alternative NADH and ubiquinone oxidases have lower ATP yields than the full pathway. The advantages produced by a shortened pathway are not entirely clear. However, the alternative oxidase is produced in response to stresses such as cold, reactive oxygen species, and infection by pathogens, as well as other factors that inhibit the full electron transport chain.[46] [47] Alternative pathways might, therefore, enhance an organisms' resistance to injury, by reducing oxidative stress.[48]

Organization of complexes
The original model for how the respiratory chain complexes are organized was that they diffuse freely and independently in the mitochondrial membrane.[17] However, recent data suggest that the complexes might form higher-order structures called supercomplexes or "respirasomes."[49] In this model, the various complexes exist as organized sets of interacting enzymes.[50] These associations might allow channeling of substrates between the various enzyme complexes, increasing the rate and efficiency of electron transfer.[51] Within such mammalian supercomplexes, some components would be present in higher amounts than others, with some data suggesting a ratio between complexes I/II/III/IV and the ATP synthase of approximately 1:1:3:7:4.[52] However, the debate over this supercomplex hypothesis is not completely resolved, as some data do not appear to fit with this model.[17] [53]

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Prokaryotic electron transport chains

Further information: Microbial metabolism In contrast to the general similarity in structure and function of the electron transport chains in eukaryotes, bacteria and archaea possess a large variety of electron-transfer enzymes. These use an equally wide set of chemicals as substrates.[54] In common with eukaryotes, prokaryotic electron transport uses the energy released from the oxidation of a substrate to pump ions across a membrane and generate an electrochemical gradient. In the bacteria, oxidative phosphorylation in Escherichia coli is understood in most detail, while archaeal systems are at present poorly understood.[55] The main difference between eukaryotic and prokaryotic oxidative phosphorylation is that bacteria and archaea use many different substances to donate or accept electrons. This allows prokaryotes to grow under a wide variety of environmental conditions.[56] In E. coli, for example, oxidative phosphorylation can be driven by a large number of pairs of reducing agents and oxidizing agents, which are listed below. The midpoint potential of a chemical measures how much energy is released when it is oxidized or reduced, with reducing agents having negative potentials and oxidizing agents positive potentials.

Respiratory enzymes and substrates in E. coli.[57]

Respiratory enzyme Formate dehydrogenase Hydrogenase NADH dehydrogenase Redox pair Bicarbonate / Formate Proton / Hydrogen NAD+ / NADH Midpoint potential (Volts) 0.43 0.42 0.32 0.19 ? 0.19 ? 0.14 +0.03 +0.82 +0.42 +0.36 +0.16 +0.13 +0.03

Glycerol-3-phosphate dehydrogenase DHAP / Gly-3-P Pyruvate oxidase Lactate dehydrogenase D-amino acid dehydrogenase Glucose dehydrogenase Succinate dehydrogenase Ubiquinol oxidase Nitrate reductase Nitrite reductase Dimethyl sulfoxide reductase Trimethylamine N-oxide reductase Fumarate reductase Acetate + Carbon dioxide / Pyruvate Pyruvate / Lactate 2-oxoacid + ammonia / D-amino acid Gluconate / Glucose Fumarate / Succinate Oxygen / Water Nitrate / Nitrite Nitrite / Ammonia DMSO / DMS TMAO / TMA Fumarate / Succinate

As shown above, E. coli can grow with reducing agents such as formate, hydrogen, or lactate as electron donors, and nitrate, DMSO, or oxygen as acceptors.[56] The larger the difference in midpoint potential between an oxidizing and reducing agent, the more energy is released when they react. Out of these compounds, the succinate/fumarate pair is unusual, as its midpoint potential is close to zero. Succinate can therefore be oxidized to fumarate if a strong oxidizing agent such as oxygen is available, or fumarate can be reduced to succinate using a strong reducing agent such as formate. These alternative reactions are catalyzed by succinate dehydrogenase and fumarate reductase, respectively.[58] Some prokaryotes use redox pairs that have only a small difference in midpoint potential. For example, nitrifying bacteria such as Nitrobacter oxidize nitrite to nitrate, donating the electrons to oxygen. The small amount of energy released in this reaction is enough to pump protons and generate ATP, but not enough to produce NADH or NADPH

Oxidative phosphorylation directly for use in anabolism.[59] This problem is solved by using a nitrite oxidoreductase to produce enough proton-motive force to run part of the electron transport chain in reverse, causing complex I to generate NADH.[60]
[61]

261

Prokaryotes control their use of these electron donors and acceptors by varying which enzymes are produced, in response to environmental conditions.[62] This flexibility is possible because different oxidases and reductases use the same ubiquinone pool. This allows many combinations of enzymes to function together, linked by the common ubiquinol intermediate.[57] These respiratory chains therefore have a modular design, with easily interchangeable sets of enzyme systems. In addition to this metabolic diversity, prokaryotes also possess a range of isozymes different enzymes that catalyze the same reaction. For example, in E. coli, there are two different types of ubiquinol oxidase using oxygen as an electron acceptor. Under highly aerobic conditions, the cell uses an oxidase with a low affinity for oxygen that can transport two protons per electron. However, if levels of oxygen fall, they switch to an oxidase that transfers only one proton per electron, but has a high affinity for oxygen.[63]

ATP synthase (complex V)

Further information: ATP synthase ATP synthase, also called complex V, is the final enzyme in the oxidative phosphorylation pathway. This enzyme is found in all forms of life and functions in the same way in both prokaryotes and eukaryotes.[64] The enzyme uses the energy stored in a proton gradient across a membrane to drive the synthesis of ATP from ADP and phosphate (Pi). Estimates of the number of protons required to synthesize one ATP have ranged from three to four,[65] [66] with some suggesting cells can vary this ratio, to suit different conditions.[67]

This phosphorylation reaction is an equilibrium, which can be shifted by altering the proton-motive force. In the absence of a proton-motive force, the ATP synthase reaction will run from right to left, hydrolyzing ATP and pumping protons out of the matrix across the membrane. However, when the proton-motive force is high, the reaction is forced to run in the opposite direction; it proceeds from left to right, allowing protons to flow down their concentration gradient and turning ADP into ATP.[64] Indeed, in the closely related vacuolar type H+-ATPases, the same reaction is used to acidify cellular compartments, by pumping protons and hydrolysing ATP.[68] ATP synthase is a massive protein complex with a mushroom-like shape. The mammalian enzyme complex contains 16 subunits and has a mass of approximately 600 kilodaltons.[69] The portion embedded within the membrane is called FO and contains a ring of c subunits and the proton channel. The stalk and the ball-shaped headpiece is called F1 and is the site of ATP synthesis. The ball-shaped complex at the end of the F1 portion contains six proteins of two different kinds (three subunits and three subunits), whereas the "stalk" consists of one protein: the subunit, with the tip of the stalk extending into the ball of and subunits.[70] Both the and subunits bind nucleotides, but only the subunits catalyze the ATP synthesis reaction. Reaching along the side of the F1 portion and back into the membrane is a long rod-like subunit that anchors the and subunits into the base of the enzyme. As protons cross the membrane through the channel in the base of ATP synthase, the FO proton-driven motor rotates.[71] Rotation might be caused by changes in the ionization of amino acids in the ring of c subunits causing electrostatic interactions that propel the ring of c subunits past the proton channel.[72] This rotating ring in turn drives the rotation of the central axle (the subunit stalk) within the and subunits. The and subunits are prevented from rotating themselves by the side-arm, which acts as a stator. This movement of the tip of the subunit within the ball of and subunits provides the energy for the active sites in the subunits to undergo a cycle of movements that produces and then releases ATP.[2]

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This ATP synthesis reaction is called the binding change mechanism and involves the active site of a subunit cycling between three states.[73] In the "open" state, ADP and phosphate enter the active site (shown in brown in the diagram). The protein then closes up around the molecules and binds them loosely the "loose" state (shown in red). The enzyme then changes shape again and forces these molecules together, with the active site in the resulting "tight" state (shown in pink) binding the newly produced ATP molecule with very high affinity. Finally, the active site cycles back to the open state, releasing ATP and binding more ADP and phosphate, ready for the next cycle.
Mechanism of ATP synthase. ATP is shown in red, ADP and

In some bacteria and archaea, ATP synthesis is driven by the phosphate in pink and the rotating subunit in black. movement of sodium ions through the cell membrane, rather than the movement of protons.[74] [75] Archaea such as Methanococcus also contain the A1Ao synthase, a form of the enzyme that contains additional proteins with little similarity in sequence to other bacterial and eukaryotic ATP synthase subunits. It is possible that, in some species, the A1Ao form of the enzyme is a specialized sodium-driven ATP synthase,[76] but this might not be true in all cases.[75]

Reactive oxygen species

Further information: Oxidative stress and Antioxidant Molecular oxygen is an ideal terminal electron acceptor because it is a strong oxidizing agent. The reduction of oxygen does involve potentially harmful intermediates.[77] Although the transfer of four electrons and four protons reduces oxygen to water, which is harmless, transfer of one or two electrons produces superoxide or peroxide anions, which are dangerously reactive.

These reactive oxygen species and their reaction products, such as the hydroxyl radical, are very harmful to cells, as they oxidize proteins and cause mutations in DNA. This cellular damage might contribute to disease and is proposed as one cause of aging.[78] [79] The cytochrome c oxidase complex is highly efficient at reducing oxygen to water, and it releases very few partly reduced intermediates; however small amounts of superoxide anion and peroxide are produced by the electron transport chain.[80] Particularly important is the reduction of coenzyme Q in complex III, as a highly reactive ubisemiquinone free radical is formed as an intermediate in the Q cycle. This unstable species can lead to electron "leakage" when electrons transfer directly to oxygen, forming superoxide.[81] As the production of reactive oxygen species by these proton-pumping complexes is greatest at high membrane potentials, it has been proposed that mitochondria regulate their activity to maintain the membrane potential within a narrow range that balances ATP production against oxidant generation.[82] For instance, oxidants can activate uncoupling proteins that reduce membrane potential.[83] To counteract these reactive oxygen species, cells contain numerous antioxidant systems, including antioxidant vitamins such as vitamin C and vitamin E, and antioxidant enzymes such as superoxide dismutase, catalase, and peroxidases,[77] which detoxify the reactive species, limiting damage to the cell.

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Inhibitors
There are several well-known drugs and toxins that inhibit oxidative phosphorylation. Although any one of these toxins inhibits only one enzyme in the electron transport chain, inhibition of any step in this process will halt the rest of the process. For example, if oligomycin inhibits ATP synthase, protons cannot pass back into the mitochondrion.[84] As a result, the proton pumps are unable to operate, as the gradient becomes too strong for them to overcome. NADH is then no longer oxidized and the citric acid cycle ceases to operate because the concentration of NAD+ falls below the concentration that these enzymes can use.
Compounds Cyanide Carbonmonoxide Azide Oligomycin CCCP 2,4-Dinitrophenol Rotenone Malonate and oxaloacetate Use Poisons Effect on oxidative phosphorylation Inhibit the electron transport chain by binding more strongly than oxygen to the FeCu center in cytochrome c [85] oxidase, preventing the reduction of oxygen.

Antibiotic Inhibits ATP synthase by blocking the flow of protons through the F subunit.[84] o Poisons Ionophores that disrupt the proton gradient by carrying protons across a membrane. This ionophore uncouples [86] proton pumping from ATP synthesis because it carries protons across the inner mitochondrial membrane.

Pesticide Prevents the transfer of electrons from complex I to ubiquinone by blocking the ubiquinone-binding site.[87] Competitive inhibitors of succinate dehydrogenase (complex II). [88]

Not all inhibitors of oxidative phosphorylation are toxins. In brown adipose tissue, regulated proton channels called uncoupling proteins can uncouple respiration from ATP synthesis.[89] This rapid respiration produces heat, and is particularly important as a way of maintaining body temperature for hibernating animals, although these proteins may also have a more general function in cells' responses to stress.[90]

History
Further information: History of biochemistry and History of molecular biology The field of oxidative phosphorylation began with the report in 1906 by Arthur Harden of a vital role for phosphate in cellular fermentation, but initially only sugar phosphates were known to be involved.[91] However, in the early 1940s, the link between the oxidation of sugars and the generation of ATP was firmly established by Herman Kalckar,[92] confirming the central role of ATP in energy transfer that had been proposed by Fritz Albert Lipmann in 1941.[93] Later, in 1949, Morris Friedkin and Albert L. Lehninger proved that the coenzyme NADH linked metabolic pathways such as the citric acid cycle and the synthesis of ATP.[94] For another twenty years, the mechanism by which ATP is generated remained mysterious, with scientists searching for an elusive "high-energy intermediate" that would link oxidation and phosphorylation reactions.[95] This puzzle was solved by Peter D. Mitchell with the publication of the chemiosmotic theory in 1961.[96] At first, this proposal was highly controversial, but it was slowly accepted and Mitchell was awarded a Nobel prize in 1978.[97] [98] Subsequent research concentrated on purifying and characterizing the enzymes involved, with major contributions being made by David E. Green on the complexes of the electron-transport chain, as well as Efraim Racker on the ATP synthase.[99] A critical step towards solving the mechanism of the ATP synthase was provided by Paul D. Boyer, by his development in 1973 of the "binding change" mechanism, followed by his radical proposal of rotational catalysis in 1982.[73] [100] More recent work has included structural studies on the enzymes involved in oxidative phosphorylation by John E. Walker, with Walker and Boyer being awarded a Nobel Prize in 1997.[101]

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References
[1] Mitchell P, Moyle J (1967). "Chemiosmotic hypothesis of oxidative phosphorylation". Nature 213 (5072): 1379. doi:10.1038/213137a0. PMID4291593. [2] Dimroth P, Kaim G, Matthey U (1 January 2000). "Crucial role of the membrane potential for ATP synthesis by F(1)F(o) ATP synthases" (http:/ / jeb. biologists. org/ cgi/ reprint/ 203/ 1/ 51). J. Exp. Biol. 203 (Pt 1): 519. PMID10600673. . [3] Schultz B, Chan S (2001). "Structures and proton-pumping strategies of mitochondrial respiratory enzymes". Annu Rev Biophys Biomol Struct 30: 2365. doi:10.1146/annurev.biophys.30.1.23. PMID11340051. [4] Rich PR (2003). "The molecular machinery of Keilin's respiratory chain" (http:/ / www. biochemsoctrans. org/ bst/ 031/ 1095/ bst0311095. htm). Biochem. Soc. Trans. 31 (Pt 6): 1095105. doi:10.1042/BST0311095. PMID14641005. . [5] Porter RK, Brand MD (1995). "Mitochondrial proton conductance and H+/O ratio are independent of electron transport rate in isolated hepatocytes". Biochem. J. 310 ((Pt 2)): 37982. PMC1135905. PMID7654171. [6] Mathews FS (1985). "The structure, function and evolution of cytochromes". Prog. Biophys. Mol. Biol. 45 (1): 156. doi:10.1016/0079-6107(85)90004-5. PMID3881803. [7] Wood PM (1983). "Why do c-type cytochromes exist?". FEBS Lett. 164 (2): 2236. doi:10.1016/0014-5793(83)80289-0. PMID6317447. [8] Crane FL (1 December 2001). "Biochemical functions of coenzyme Q10" (http:/ / www. jacn. org/ cgi/ content/ full/ 20/ 6/ 591). Journal of the American College of Nutrition 20 (6): 5918. PMID11771674. . [9] Mitchell P (1979). "Keilin's respiratory chain concept and its chemiosmotic consequences". Science 206 (4423): 114859. doi:10.1126/science.388618. PMID388618. [10] Sballe B, Poole RK (1999). "Microbial ubiquinones: multiple roles in respiration, gene regulation and oxidative stress management" (http:/ / mic. sgmjournals. org/ cgi/ reprint/ 145/ 8/ 1817. pdf) (PDF). Microbiology (Reading, Engl.) 145 (Pt 8): 181730. PMID10463148. . [11] Johnson D, Dean D, Smith A, Johnson M (2005). "Structure, function, and formation of biological iron-sulfur clusters". Annu Rev Biochem 74: 24781. doi:10.1146/annurev.biochem.74.082803.133518. PMID15952888. [12] Page CC, Moser CC, Chen X, Dutton PL (1999). "Natural engineering principles of electron tunnelling in biological oxidation-reduction". Nature 402 (6757): 4752. doi:10.1038/46972. PMID10573417. [13] Leys D, Scrutton NS (2004). "Electrical circuitry in biology: emerging principles from protein structure". Curr. Opin. Struct. Biol. 14 (6): 6427. doi:10.1016/j.sbi.2004.10.002. PMID15582386. [14] Boxma B, de Graaf RM, van der Staay GW, et al. (2005). "An anaerobic mitochondrion that produces hydrogen". Nature 434 (7029): 749. doi:10.1038/nature03343. PMID15744302. [15] Medical CHEMISTRY Compendium. By Anders Overgaard Pedersen and Henning Nielsen. Aarhus University. 2008 [16] Hirst J (2005). "Energy transduction by respiratory complex Ian evaluation of current knowledge" (http:/ / www. biochemsoctrans. org/ bst/ 033/ 0525/ 0330525. pdf) (PDF). Biochem. Soc. Trans. 33 (Pt 3): 5259. doi:10.1042/BST0330525. PMID15916556. . [17] Lenaz G, Fato R, Genova M, Bergamini C, Bianchi C, Biondi A (2006). "Mitochondrial Complex I: structural and functional aspects". Biochim Biophys Acta 1757 (910): 140620. doi:10.1016/j.bbabio.2006.05.007. PMID16828051. [18] Sazanov L.A., Hinchliffe P. (2006). "Structure of the hydrophilic domain of respiratory complex I from Thermus thermophilus". Science 311 (5766): 14301436. doi:10.1126/science.1123809. PMID16469879. [19] Efremov R.G., Baradaran R., & Sazanov L.A., (2010) The arcdhitecture of respiratory complex I, Nature 465, 441-445 [20] Baranova EA, Holt PJ, Sazanov LA (2007). "Projection structure of the membrane domain of Escherichia coli respiratory complex I at 8 A resolution". J. Mol. Biol. 366 (1): 14054. doi:10.1016/j.jmb.2006.11.026. PMID17157874. [21] Friedrich T, Bttcher B (2004). "The gross structure of the respiratory complex I: a Lego System". Biochim. Biophys. Acta 1608 (1): 19. doi:10.1016/j.bbabio.2003.10.002. PMID14741580. [22] Hirst J (January 2010). "Towards the molecular mechanism of respiratory complex I". Biochem. J. 425 (2): 32739. doi:10.1042/BJ20091382. PMID20025615. [23] Cecchini G (2003). "Function and structure of complex II of the respiratory chain". Annu Rev Biochem 72: 77109. doi:10.1146/annurev.biochem.72.121801.161700. PMID14527321. [24] Yankovskaya V., Horsefield R., Tornroth S., Luna-Chavez C., Miyoshi H., Leger C., Byrne B., Cecchini G. et al. (2003). "Architecture of succinate dehydrogenase and reactive oxygen species generation". Science 299 (5607): 700704. doi:10.1126/science.1079605. PMID12560550. [25] Horsefield R, Iwata S, Byrne B (2004). "Complex II from a structural perspective". Curr. Protein Pept. Sci. 5 (2): 10718. doi:10.2174/1389203043486847. PMID15078221. [26] Kita K, Hirawake H, Miyadera H, Amino H, Takeo S (2002). "Role of complex II in anaerobic respiration of the parasite mitochondria from Ascaris suum and Plasmodium falciparum". Biochim. Biophys. Acta 1553 (12): 12339. doi:10.1016/S0005-2728(01)00237-7. PMID11803022. [27] Painter HJ, Morrisey JM, Mather MW, Vaidya AB (2007). "Specific role of mitochondrial electron transport in blood-stage Plasmodium falciparum". Nature 446 (7131): 8891. doi:10.1038/nature05572. PMID17330044. [28] Ramsay RR, Steenkamp DJ, Husain M (1987). "Reactions of electron-transfer flavoprotein and electron-transfer flavoprotein: ubiquinone oxidoreductase". Biochem. J. 241 (3): 88392. PMC1147643. PMID3593226. [29] Zhang J, Frerman FE, Kim JJ (2006). "Structure of electron transfer flavoprotein-ubiquinone oxidoreductase and electron transfer to the mitochondrial ubiquinone pool". Proc. Natl. Acad. Sci. U.S.A. 103 (44): 162127. doi:10.1073/pnas.0604567103. PMC1637562.

Oxidative phosphorylation
PMID17050691. [30] Ikeda Y, Dabrowski C, Tanaka K (25 January 1983). "Separation and properties of five distinct acyl-CoA dehydrogenases from rat liver mitochondria. Identification of a new 2-methyl branched chain acyl-CoA dehydrogenase" (http:/ / www. jbc. org/ cgi/ reprint/ 258/ 2/ 1066). J. Biol. Chem. 258 (2): 106676. PMID6401712. . [31] Ruzicka FJ, Beinert H (1977). "A new iron-sulfur flavoprotein of the respiratory chain. A component of the fatty acid beta oxidation pathway" (http:/ / www. jbc. org/ cgi/ reprint/ 252/ 23/ 8440. pdf) (PDF). J. Biol. Chem. 252 (23): 84405. PMID925004. . [32] Ishizaki K, Larson TR, Schauer N, Fernie AR, Graham IA, Leaver CJ (2005). "The Critical Role of Arabidopsis Electron-Transfer Flavoprotein:Ubiquinone Oxidoreductase during Dark-Induced Starvation". Plant Cell 17 (9): 2587600. doi:10.1105/tpc.105.035162. PMC1197437. PMID16055629. [33] Berry E, Guergova-Kuras M, Huang L, Crofts A (2000). "Structure and function of cytochrome bc complexes". Annu Rev Biochem 69: 100575. doi:10.1146/annurev.biochem.69.1.1005. PMID10966481. [34] Crofts AR (2004). "The cytochrome bc1 complex: function in the context of structure". Annu. Rev. Physiol. 66: 689733. doi:10.1146/annurev.physiol.66.032102.150251. PMID14977419. [35] Iwata S, Lee JW, Okada K, et al. (1998). "Complete structure of the 11-subunit bovine mitochondrial cytochrome bc1 complex". Science 281 (5373): 6471. doi:10.1126/science.281.5373.64. PMID9651245. [36] Trumpower BL (1990). "The protonmotive Q cycle. Energy transduction by coupling of proton translocation to electron transfer by the cytochrome bc1 complex" (http:/ / www. jbc. org/ cgi/ reprint/ 265/ 20/ 11409. pdf) (PDF). J. Biol. Chem. 265 (20): 1140912. PMID2164001. . [37] Hunte C, Palsdottir H, Trumpower BL (2003). "Protonmotive pathways and mechanisms in the cytochrome bc1 complex". FEBS Lett. 545 (1): 3946. doi:10.1016/S0014-5793(03)00391-0. PMID12788490. [38] Calhoun M, Thomas J, Gennis R (1994). "The cytochrome oxidase superfamily of redox-driven proton pumps". Trends Biochem Sci 19 (8): 32530. doi:10.1016/0968-0004(94)90071-X. PMID7940677. [39] Tsukihara T, Aoyama H, Yamashita E, Tomizaki T, Yamaguchi H, Shinzawa-Itoh K, Nakashima R, Yaono R, Yoshikawa S. (1996). "The whole structure of the 13-subunit oxidized cytochrome c oxidase at 2.8 A". Science 272 (5265): 113644. Bibcode1996Sci...272.1136T. doi:10.1126/science.272.5265.1136. PMID8638158. [40] Yoshikawa S, Muramoto K, Shinzawa-Itoh K, et al. (2006). "Proton pumping mechanism of bovine heart cytochrome c oxidase". Biochim. Biophys. Acta 1757 (910): 11106. doi:10.1016/j.bbabio.2006.06.004. PMID16904626. [41] Rasmusson AG, Soole KL, Elthon TE (2004). "Alternative NAD(P)H dehydrogenases of plant mitochondria". Annual review of plant biology 55: 2339. doi:10.1146/annurev.arplant.55.031903.141720. PMID15725055. [42] Menz RI, Day DA (1996). "Purification and characterization of a 43-kDa rotenone-insensitive NADH dehydrogenase from plant mitochondria" (http:/ / www. jbc. org/ cgi/ content/ full/ 271/ 38/ 23117). J. Biol. Chem. 271 (38): 2311720. doi:10.1074/jbc.271.38.23117. PMID8798503. . [43] McDonald A, Vanlerberghe G (2004). "Branched mitochondrial electron transport in the Animalia: presence of alternative oxidase in several animal phyla". IUBMB Life 56 (6): 33341. doi:10.1080/1521-6540400000876. PMID15370881. [44] Sluse FE, Jarmuszkiewicz W (1998). "Alternative oxidase in the branched mitochondrial respiratory network: an overview on structure, function, regulation, and role". Braz. J. Med. Biol. Res. 31 (6): 73347. doi:10.1590/S0100-879X1998000600003. PMID9698817. [45] Moore AL, Siedow JN (1991). "The regulation and nature of the cyanide-resistant alternative oxidase of plant mitochondria". Biochim. Biophys. Acta 1059 (2): 12140. doi:10.1016/S0005-2728(05)80197-5. PMID1883834. [46] Vanlerberghe GC, McIntosh L (1997). "Alternative oxidase: From Gene to Function". Annual Review of Plant Physiology and Plant Molecular Biology 48: 70334. doi:10.1146/annurev.arplant.48.1.703. PMID15012279. [47] Ito Y, Saisho D, Nakazono M, Tsutsumi N, Hirai A (1997). "Transcript levels of tandem-arranged alternative oxidase genes in rice are increased by low temperature". Gene 203 (2): 1219. doi:10.1016/S0378-1119(97)00502-7. PMID9426242. [48] Maxwell DP, Wang Y, McIntosh L (1999). "The alternative oxidase lowers mitochondrial reactive oxygen production in plant cells" (http:/ / www. pnas. org/ cgi/ content/ full/ 96/ 14/ 8271). Proc. Natl. Acad. Sci. U.S.A. 96 (14): 82716. doi:10.1073/pnas.96.14.8271. PMC22224. PMID10393984. . [49] Heinemeyer J, Braun HP, Boekema EJ, Kouril R (2007). "A structural model of the cytochrome C reductase/oxidase supercomplex from yeast mitochondria". J. Biol. Chem. 282 (16): 122408. doi:10.1074/jbc.M610545200. PMID17322303. [50] Schgger H, Pfeiffer K (2000). "Supercomplexes in the respiratory chains of yeast and mammalian mitochondria". EMBO J. 19 (8): 177783. doi:10.1093/emboj/19.8.1777. PMC302020. PMID10775262. [51] Schgger H (2002). "Respiratory chain supercomplexes of mitochondria and bacteria". Biochim. Biophys. Acta 1555 (13): 1549. doi:10.1016/S0005-2728(02)00271-2. PMID12206908. [52] Schgger H, Pfeiffer K (2001). "The ratio of oxidative phosphorylation complexes I-V in bovine heart mitochondria and the composition of respiratory chain supercomplexes" (http:/ / www. jbc. org/ cgi/ content/ full/ 276/ 41/ 37861). J. Biol. Chem. 276 (41): 378617. doi:10.1074/jbc.M106474200. PMID11483615. . [53] Gupte S, Wu ES, Hoechli L, et al. (1984). "Relationship between lateral diffusion, collision frequency, and electron transfer of mitochondrial inner membrane oxidation-reduction components". Proc. Natl. Acad. Sci. U.S.A. 81 (9): 260610. doi:10.1073/pnas.81.9.2606. PMC345118. PMID6326133. [54] Nealson KH (1999). "Post-Viking microbiology: new approaches, new data, new insights". Origins of life and evolution of the biosphere: the journal of the International Society for the Study of the Origin of Life 29 (1): 7393. doi:10.1023/A:1006515817767. PMID11536899.

265

Oxidative phosphorylation
[55] Schfer G, Engelhard M, Mller V (1999). "Bioenergetics of the Archaea". Microbiol. Mol. Biol. Rev. 63 (3): 570620. PMC103747. PMID10477309. [56] Ingledew WJ, Poole RK (1984). "The respiratory chains of Escherichia coli". Microbiol. Rev. 48 (3): 22271. PMC373010. PMID6387427. [57] Unden G, Bongaerts J (1997). "Alternative respiratory pathways of Escherichia coli: energetics and transcriptional regulation in response to electron acceptors". Biochim. Biophys. Acta 1320 (3): 21734. doi:10.1016/S0005-2728(97)00034-0. PMID9230919. [58] Cecchini G, Schrder I, Gunsalus RP, Maklashina E (2002). "Succinate dehydrogenase and fumarate reductase from Escherichia coli". Biochim. Biophys. Acta 1553 (12): 14057. doi:10.1016/S0005-2728(01)00238-9. PMID11803023. [59] Freitag A, Bock E (1990). "Energy conservation in Nitrobacter". FEMS Microbiology Letters 66 (13): 15762. doi:10.1111/j.1574-6968.1990.tb03989.x. [60] Starkenburg SR, Chain PS, Sayavedra-Soto LA, et al. (2006). "Genome Sequence of the Chemolithoautotrophic Nitrite-Oxidizing Bacterium Nitrobacter winogradskyi Nb-255" (http:/ / aem. asm. org/ cgi/ content/ full/ 72/ 3/ 2050?view=long& pmid=16517654). Appl. Environ. Microbiol. 72 (3): 205063. doi:10.1128/AEM.72.3.2050-2063.2006. PMC1393235. PMID16517654. . [61] Yamanaka T, Fukumori Y (1988). "The nitrite oxidizing system of Nitrobacter winogradskyi". FEMS Microbiol. Rev. 4 (4): 25970. PMID2856189. [62] Iuchi S, Lin EC (1993). "Adaptation of Escherichia coli to redox environments by gene expression". Mol. Microbiol. 9 (1): 915. doi:10.1111/j.1365-2958.1993.tb01664.x. PMID8412675. [63] Calhoun MW, Oden KL, Gennis RB, de Mattos MJ, Neijssel OM (1993). "Energetic efficiency of Escherichia coli: effects of mutations in components of the aerobic respiratory chain" (http:/ / jb. asm. org/ cgi/ reprint/ 175/ 10/ 3020. pdf) (PDF). J. Bacteriol. 175 (10): 30205. PMC204621. PMID8491720. . [64] Boyer PD (1997). "The ATP synthasea splendid molecular machine". Annu. Rev. Biochem. 66: 71749. doi:10.1146/annurev.biochem.66.1.717. PMID9242922. [65] Van Walraven HS, Strotmann H, Schwarz O, Rumberg B (1996). "The H+/ATP coupling ratio of the ATP synthase from thiol-modulated chloroplasts and two cyanobacterial strains is four". FEBS Lett. 379 (3): 30913. doi:10.1016/0014-5793(95)01536-1. PMID8603713. [66] Yoshida M, Muneyuki E, Hisabori T (2001). "ATP synthasea marvellous rotary engine of the cell". Nat. Rev. Mol. Cell Biol. 2 (9): 66977. doi:10.1038/35089509. PMID11533724. [67] Schemidt RA, Qu J, Williams JR, Brusilow WS (1998). "Effects of Carbon Source on Expression of Fo Genes and on the Stoichiometry of the c Subunit in the F1Fo ATPase of Escherichia coli". J. Bacteriol. 180 (12): 32058. PMC107823. PMID9620972. [68] Nelson N, Perzov N, Cohen A, Hagai K, Padler V, Nelson H (1 January 2000). "The cellular biology of proton-motive force generation by V-ATPases" (http:/ / jeb. biologists. org/ cgi/ reprint/ 203/ 1/ 89). J. Exp. Biol. 203 (Pt 1): 8995. PMID10600677. . [69] Rubinstein JL, Walker JE, Henderson R (2003). "Structure of the mitochondrial ATP synthase by electron cryomicroscopy". EMBO J. 22 (23): 618292. doi:10.1093/emboj/cdg608. PMC291849. PMID14633978. [70] Leslie AG, Walker JE (2000). "Structural model of F1-ATPase and the implications for rotary catalysis". Philos. Trans. R. Soc. Lond., B, Biol. Sci. 355 (1396): 46571. doi:10.1098/rstb.2000.0588. PMC1692760. PMID10836500. [71] Noji H, Yoshida M (2001). "The rotary machine in the cell, ATP synthase" (http:/ / www. jbc. org/ cgi/ content/ full/ 276/ 3/ 1665). J. Biol. Chem. 276 (3): 16658. doi:10.1074/jbc.R000021200. PMID11080505. . [72] Capaldi R, Aggeler R (2002). "Mechanism of the F(1)F(0)-type ATP synthase, a biological rotary motor". Trends Biochem Sci 27 (3): 15460. doi:10.1016/S0968-0004(01)02051-5. PMID11893513. [73] Gresser MJ, Myers JA, Boyer PD (25 October 1982). "Catalytic site cooperativity of beef heart mitochondrial F1 adenosine triphosphatase. Correlations of initial velocity, bound intermediate, and oxygen exchange measurements with an alternating three-site model" (http:/ / www. jbc. org/ cgi/ reprint/ 257/ 20/ 12030). J. Biol. Chem. 257 (20): 120308. PMID6214554. . [74] Dimroth P (1994). "Bacterial sodium ion-coupled energetics". Antonie Van Leeuwenhoek 65 (4): 38195. doi:10.1007/BF00872221. PMID7832594. [75] Becher B, Mller V (1994). "Delta mu Na+ drives the synthesis of ATP via an delta mu Na(+)-translocating F1F0-ATP synthase in membrane vesicles of the archaeon Methanosarcina mazei G1". J. Bacteriol. 176 (9): 254350. PMC205391. PMID8169202. [76] Mller V (2004). "An exceptional variability in the motor of archaeal A1A0 ATPases: from multimeric to monomeric rotors comprising 613 ion binding sites". J. Bioenerg. Biomembr. 36 (1): 11525. doi:10.1023/B:JOBB.0000019603.68282.04. PMID15168615. [77] Davies K (1995). "Oxidative stress: the paradox of aerobic life". Biochem Soc Symp 61: 131. PMID8660387. [78] Rattan SI (2006). "Theories of biological aging: genes, proteins, and free radicals". Free Radic. Res. 40 (12): 12308. doi:10.1080/10715760600911303. PMID17090411. [79] Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J (2007). "Free radicals and antioxidants in normal physiological functions and human disease". Int. J. Biochem. Cell Biol. 39 (1): 4484. doi:10.1016/j.biocel.2006.07.001. PMID16978905. [80] Raha S, Robinson B (2000). "Mitochondria, oxygen free radicals, disease and ageing". Trends Biochem Sci 25 (10): 5028. doi:10.1016/S0968-0004(00)01674-1. PMID11050436. [81] Finkel T, Holbrook NJ (2000). "Oxidants, oxidative stress and the biology of ageing". Nature 408 (6809): 23947. doi:10.1038/35041687. PMID11089981. [82] Kadenbach B, Ramzan R, Wen L, Vogt S (May 2009). "New extension of the Mitchell Theory for oxidative phosphorylation in mitochondria of living organisms". Biochim. Biophys. Acta 1800 (3): 205212. doi:10.1016/j.bbagen.2009.04.019. PMID19409964.

266

Oxidative phosphorylation
[83] Echtay KS, Roussel D, St-Pierre J, et al. (January 2002). "Superoxide activates mitochondrial uncoupling proteins". Nature 415 (6867): 969. doi:10.1038/415096a. PMID11780125. [84] Joshi S, Huang YG (1991). "ATP synthase complex from bovine heart mitochondria: the oligomycin sensitivity conferring protein is essential for dicyclohexyl carbodiimide-sensitive ATPase". Biochim. Biophys. Acta 1067 (2): 2558. doi:10.1016/0005-2736(91)90051-9. PMID1831660. [85] Tsubaki M; Yoshikawa, Shinya (1993). "Fourier-transform infrared study of cyanide binding to the Fea3-CuB binuclear site of bovine heart cytochrome c oxidase: implication of the redox-linked conformational change at the binuclear site". Biochemistry 32 (1): 16473. doi:10.1021/bi00052a022. PMID8380331. [86] Heytler PG (1979). "Uncouplers of oxidative phosphorylation". Meth. Enzymol.. Methods in Enzymology 55: 46242. doi:10.1016/0076-6879(79)55060-5. ISBN9780121819552. PMID156853. [87] Lambert AJ, Brand MD (2004). "Inhibitors of the quinone-binding site allow rapid superoxide production from mitochondrial NADH: ubiquinone oxidoreductase (complex I)" (http:/ / www. jbc. org/ cgi/ content/ full/ 279/ 38/ 39414). J. Biol. Chem. 279 (38): 3941420. doi:10.1074/jbc.M406576200. PMID15262965. . [88] Dervartanian DV, Veeger C. (November 1964). "Studies on succinate dehydrogenase. I. Spectral properties of the purified enzyme and formation of enzyme-competitive inhibitor complexes". Biochim. Biophys. Acta 92: 23347. PMID14249115. [89] Ricquier D, Bouillaud F (2000). "The uncoupling protein homologues: UCP1, UCP2, UCP3, StUCP and AtUCP". Biochem. J. 345 Pt 2 (2): 16179. doi:10.1042/0264-6021:3450161. PMC1220743. PMID10620491. [90] Boreck J, Vercesi AE (2005). "Plant uncoupling mitochondrial protein and alternative oxidase: energy metabolism and stress". Biosci. Rep. 25 (34): 27186. doi:10.1007/s10540-005-2889-2. PMID16283557. [91] Harden A, Young WJ. (1906). "The alcoholic ferment of yeast-juice". Proc. R. Soc. (Lond.) B (77): 40520. doi:10.1098/rspb.1906.0029. [92] Kalckar HM (1974). "Origins of the concept oxidative phosphorylation". Mol. Cell. Biochem. 5 (12): 5563. doi:10.1007/BF01874172. PMID4279328. [93] Lipmann F, (1941). "Metabolic generation and utilization of phosphate bond energy". Adv Enzymol 1: 99162. [94] Friedkin M, Lehninger AL. (1 April 1949). "Esterification of inorganic phosphate coupled to electron transport between dihydrodiphosphopyridine nucleotide and oxygen" (http:/ / www. jbc. org/ cgi/ reprint/ 178/ 2/ 611). J. Biol. Chem. 178 (2): 61123. PMID18116985. . [95] Slater EC. (1953). "Mechanism of Phosphorylation in the Respiratory Chain". Nature 172 (4387): 9758. doi:10.1038/172975a0. PMID13111237. [96] Mitchell P. (1961). "Coupling of Phosphorylation to Electron and Hydrogen Transfer by a Chemi-Osmotic type of Mechanism". Nature 191 (4784): 1448. Bibcode1961Natur.191..144M. doi:10.1038/191144a0. PMID13771349. [97] Milton H. Saier Jr. "Peter Mitchell and the Vital Force" (http:/ / www-biology. ucsd. edu/ ~msaier/ transport/ petermitchell/ MitchellFrame-1. html). . Retrieved 2007-08-23. [98] Mitchell, Peter (1978). "David Keilin's Respiratory Chain Concept and Its Chemiosmotic Consequences" (http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1978/ mitchell-lecture. pdf) (Pdf). Nobel lecture. Nobel Foundation. . Retrieved 2007-07-21. [99] Pullman ME, Penefsky HS, Datta A, and Racker E. (1 November 1960). "Partial Resolution of the Enzymes Catalyzing Oxidative Phosphorylation. I. Purification and Properties of Soluble, Dinitrophenol-stimulated Adenosine Triphosphatase" (http:/ / www. jbc. org/ cgi/ reprint/ 235/ 11/ 3322). J. Biol. Chem. 235 (11): 33229. PMID13738472. . [100] Boyer PD, Cross RL, Momsen W (1973). "A New Concept for Energy Coupling in Oxidative Phosphorylation Based on a Molecular Explanation of the Oxygen Exchange Reactions". Proc. Natl. Acad. Sci. U.S.A. 70 (10): 28379. doi:10.1073/pnas.70.10.2837. PMC427120. PMID4517936. [101] "The Nobel Prize in Chemistry 1997" (http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 1997/ ). Nobel Foundation. . Retrieved 2007-07-21.

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Further reading
Introductory Nelson DL; Cox MM (2004). Lehninger Principles of Biochemistry (4th ed.). W. H. Freeman. ISBN0-7167-4339-6. Schneider ED; Sagan D (2006). Into the Cool: Energy Flow, Thermodynamics and Life (1st ed.). University of Chicago Press. ISBN0-226-73937-6. Lane N (2006). Power, Sex, Suicide: Mitochondria and the Meaning of Life (1st ed.). Oxford University Press, USA. ISBN0-19-920564-7. Advanced Nicholls DG; Ferguson SJ (2002). Bioenergetics 3 (1st ed.). Academic Press. ISBN0-12-518121-3. Haynie D (2001). Biological Thermodynamics (1st ed.). Cambridge University Press. ISBN0-521-79549-4. Rajan SS (2003). Introduction to Bioenergetics (1st ed.). Anmol. ISBN8-126-11364-2.

Oxidative phosphorylation Wikstrom M (Ed) (2005). Biophysical and Structural Aspects of Bioenergetics (1st ed.). Royal Society of Chemistry. ISBN0-85404-346-2.

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External links
General resources Animated diagrams illustrating oxidative phosphorylation (http://www.wiley.com/legacy/college/boyer/ 0470003790/animations/electron_transport/electron_transport.htm) Wiley and Co Concepts in Biochemistry ATP synthase - the rotary engine in the cell (http://www.res.titech.ac.jp/~seibutu/) Brief introduction, including videos of microscope images of the enzyme rotating, at Tokyo Institute of Technology On-line biophysics lectures (http://www.life.uiuc.edu/crofts/bioph354/) Antony Crofts, University of Illinois at Urbana-Champaign Structural resources Animations of the ATP synthase (http://nature.berkeley.edu/~hongwang/Project/ATP_synthase/) Hongyun Wang and George Oster, University of California, Berkeley PDB molecule of the month: ATP synthase (http://www.rcsb.org/pdb/static.do?p=education_discussion/molecule_of_the_month/ pdb72_1.html) Cytochrome c (http://www.rcsb.org/pdb/static.do?p=education_discussion/molecule_of_the_month/ pdb36_1.html) Cytochrome c oxidase (http://www.rcsb.org/pdb/static.do?p=education_discussion/ molecule_of_the_month/pdb5_1.html) Interactive molecular models at Universidade Fernando Pessoa: NADH dehydrogenase (http://www2.ufp.pt/~pedros/anim/2frame-ien.htm) succinate dehydrogenase (http://www2.ufp.pt/~pedros/anim/2frame-iien.htm) Coenzyme Q - cytochrome c reductase (http://www2.ufp.pt/~pedros/anim/2frame-iiien.htm) cytochrome c oxidase (http://www2.ufp.pt/~pedros/anim/2frame-iven.htm)

269

Photosynthesis
Photosynthesis
Photosynthesis (English pronunciation: /fotosnss/; from the Greek - [photo-], "light," and [synthesis], "putting together", "composition") is a chemical process that converts carbon dioxide into organic compounds, especially sugars, using the energy from sunlight.[1] Photosynthesis occurs in plants, algae, and many species of bacteria, but not in archaea. Photosynthetic organisms are called photoautotrophs, since they can create their own food. In plants, Composite image showing the global distribution of photosynthesis, including both algae, and cyanobacteria, oceanic phytoplankton and vegetation photosynthesis uses carbon dioxide and water, releasing oxygen as a waste product. Photosynthesis is vital for all aerobic life on Earth. In addition to maintaining normal levels of oxygen in the atmosphere, photosynthesis is the Overall equation for the type of photosynthesis that occurs in plants source of energy for nearly all life on earth, either directly, through primary production, or indirectly, as the ultimate source of the energy in their food,[2] the exceptions being chemoautotrophs that live in rocks or around deep sea hydrothermal vents. The rate of energy capture by photosynthesis is immense, approximately 100terawatts,[3] which is about six times larger than the power consumption of human civilization.[4] As well as energy, photosynthesis is also the source of the carbon in all the organic compounds within organisms' bodies. In all, photosynthetic organisms convert around 100115 petagrams of carbon into biomass per year.[5] [6] Although photosynthesis can happen in different ways in different species, some features are always the same. For example, the process always begins when energy from light is absorbed by proteins called photosynthetic reaction centers that contain chlorophylls. In plants, these proteins are held inside organelles called chloroplasts, while in bacteria they are embedded in the plasma membrane. Some of the light energy gathered by chlorophylls is stored in the form of adenosine triphosphate (ATP). The rest of the energy is used to remove electrons from a substance such as water. These electrons are then used in the reactions that turn carbon dioxide into organic compounds. In plants, algae and cyanobacteria, this is done by a sequence of reactions called the Calvin cycle, but different sets of reactions are found in some bacteria, such as the reverse Krebs cycle in Chlorobium. Many photosynthetic organisms have adaptations that concentrate or store carbon dioxide. This helps reduce a wasteful process called photorespiration that can consume part of the sugar produced during photosynthesis.

Photosynthesis The first photosynthetic organisms probably evolved about 3500 [7] million years ago, early in the evolutionary history of life, when all forms of life on Earth were microorganisms and the atmosphere had much more carbon dioxide. They most likely used hydrogen or hydrogen sulfide as sources of electrons, rather than water.[8] Cyanobacteria appeared later, around 3000 [9] million years ago, and drastically changed the Earth when they began to oxygenate the atmosphere, beginning about 2400 [10] million years ago.[11] This new atmosphere allowed the evolution of complex life such as protists. Eventually, no later than a billion years ago, one of these protists formed a symbiotic relationship with a cyanobacterium, producing the ancestor of many plants and algae.[12] The chloroplasts in modern plants are the descendants of these ancient symbiotic cyanobacteria.[13]

270

Schematic of photosynthesis in plants

Overview
Photosynthetic organisms are photoautotrophs, which means that they are repositories of energy, they are able to synthesize food directly from carbon dioxide, water, and using energy from light. They accrue it as part of their potential energy. However, not all organisms that use light as a source of energy carry out photosynthesis, since photoheterotrophs use organic compounds, rather than carbon dioxide, as a source of carbon.[2] In plants, algae and cyanobacteria, photosynthesis releases oxygen. This is called oxygenic photosynthesis. Although there are some differences between oxygenic photosynthesis in plants, algae and cyanobacteria, the overall process is quite similar in these organisms. However, there are some types of bacteria that carry out anoxygenic photosynthesis, which consumes carbon dioxide but does not release oxygen. Carbon dioxide is converted into sugars in a process called carbon Photosynthesis changes sunlight into chemical fixation. Carbon fixation is a redox reaction, so photosynthesis needs energy, splits water to liberate O2, and fixes CO2 into sugar. to supply both a source of energy to drive this process, and the electrons needed to convert carbon dioxide into a carbohydrate, which is a reduction reaction. In general outline, photosynthesis is the opposite of cellular respiration, where glucose and other compounds are oxidized to produce carbon dioxide, water, and release chemical energy. However, the two processes take place through a different sequence of chemical reactions and in different cellular compartments. The general equation for photosynthesis is therefore: 2n CO2 + 2n DH2 + photons 2(CH2O)n + 2n DO Carbon dioxide + electron donor + light energy carbohydrate + oxidized electron donor In oxygenic photosynthesis water is the electron donor and, since its hydrolysis releases oxygen, the equation for this process is:

Photosynthesis 2n CO2 + 4n H2O + photons 2(CH2O)n + 2n O2 + 2n H2O carbon dioxide + water + light energy carbohydrate + oxygen + water Often 2n water molecules are cancelled on both sides, yielding: 2n CO2 + 2n H2O + photons 2(CH2O)n + 2n O2 carbon dioxide + water + light energy carbohydrate + oxygen Other processes substitute other compounds (such as arsenite) for water in the electron-supply role; the microbes use sunlight to oxidize arsenite to arsenate:[14] The equation for this reaction is: CO2 + (AsO33) + photons (AsO43) + CO [15] carbon dioxide + arsenite + light energy arsenate + carbon monoxide (used to build other compounds in subsequent reactions) Photosynthesis occurs in two stages. In the first stage, light-dependent reactions or light reactions capture the energy of light and use it to make the energy-storage molecules ATP and NADPH. During the second stage, the light-independent reactions use these products to capture and reduce carbon dioxide. Most organisms that utilize photosynthesis to produce oxygen use visible light to do so, although at least three use infrared radiation.[16]

271

Photosynthetic membranes and organelles

The proteins that gather light for photosynthesis are embedded within cell membranes. The simplest way these are arranged is in photosynthetic bacteria, where these proteins are held within the plasma membrane.[17] However, this membrane may be tightly folded into cylindrical sheets called thylakoids,[18] or bunched up into round vesicles called intracytoplasmic membranes.[19] These structures can fill most of the interior of a cell, giving the membrane a very large surface area and therefore increasing the amount of light that the bacteria can absorb.[18]

Chloroplast ultrastructure: 1. outer membrane 2. intermembrane space3. inner membrane (1+2+3: envelope) 4. stroma (aqueous fluid) 5. thylakoid lumen (inside of thylakoid) 6. thylakoid membrane 7. granum (stack of thylakoids) 8. thylakoid (lamella) 9. starch 10. ribosome 11. plastidial DNA 12. plastoglobule (drop of lipids)

In plants and algae, photosynthesis takes place in organelles called chloroplasts. A typical plant cell contains about 10 to 100 chloroplasts. The chloroplast is enclosed by a membrane. This membrane is composed of a phospholipid inner membrane, a phospholipid outer membrane, and an intermembrane space between them. Within the membrane is an aqueous fluid called the stroma. The stroma contains stacks (grana) of thylakoids, which are the site of photosynthesis. The thylakoids are flattened disks, bounded by a membrane with a lumen or thylakoid space within it. The site of photosynthesis is the thylakoid membrane, which contains integral and peripheral membrane protein complexes, including the pigments that absorb light energy, which form the photosystems. Plants absorb light primarily using the pigment chlorophyll, which is the reason that most plants have a green color. Besides chlorophyll, plants also use pigments such as carotenes and xanthophylls.[20] Algae also use chlorophyll, but various other pigments are present as phycocyanin, carotenes, and xanthophylls in green algae, phycoerythrin in red algae (rhodophytes) and fucoxanthin in brown algae and diatoms resulting in a wide variety of colors. These pigments are embedded in plants and algae in special antenna-proteins. In such proteins all the pigments are ordered to work well together. Such a protein is also called a light-harvesting complex.

Photosynthesis Although all cells in the green parts of a plant have chloroplasts, most of the energy is captured in the leaves. The cells in the interior tissues of a leaf, called the mesophyll, can contain between 450,000 and 800,000 chloroplasts for every square millimeter of leaf. The surface of the leaf is uniformly coated with a water-resistant waxy cuticle that protects the leaf from excessive evaporation of water and decreases the absorption of ultraviolet or blue light to reduce heating. The transparent epidermis layer allows light to pass through to the palisade mesophyll cells where most of the photosynthesis takes place.

272

Light reactions
In the light reactions, one molecule of the pigment chlorophyll absorbs one photon and loses one electron. This electron is passed to a modified form of chlorophyll called pheophytin, which passes the electron to a quinone molecule, allowing the start of a flow of electrons down an electron transport chain that leads to the ultimate reduction of NADP to NADPH. In addition, this creates a proton gradient across the chloroplast membrane; its Light-dependent reactions of photosynthesis at the thylakoid membrane dissipation is used by ATP synthase for the concomitant synthesis of ATP. The chlorophyll molecule regains the lost electron from a water molecule through a process called photolysis, which releases a dioxygen (O2) molecule. The overall equation for the light-dependent reactions under the conditions of non-cyclic electron flow in green plants is:[21] 2 H2O + 2 NADP+ + 3 ADP + 3 Pi + light 2 NADPH + 2 H+ + 3 ATP + O2 Not all wavelengths of light can support photosynthesis. The photosynthetic action spectrum depends on the type of accessory pigments present. For example, in green plants, the action spectrum resembles the absorption spectrum for chlorophylls and carotenoids with peaks for violet-blue and red light. In red algae, the action spectrum overlaps with the absorption spectrum of phycobilins for red blue-green light, which allows these algae to grow in deeper waters that filter out the longer wavelengths used by green plants. The non-absorbed part of the light spectrum is what gives photosynthetic organisms their color (e.g., green plants, red algae, purple bacteria) and is the least effective for photosynthesis in the respective organisms.

Photosynthesis

273

Z scheme

The "Z scheme"

In plants, light-dependent reactions occur in the thylakoid membranes of the chloroplasts and use light energy to synthesize ATP and NADPH. The light-dependent reaction has two forms: cyclic and non-cyclic. In the non-cyclic reaction, the photons are captured in the light-harvesting antenna complexes of photosystem II by chlorophyll and other accessory pigments (see diagram at right). When a chlorophyll molecule at the core of the photosystem II reaction center obtains sufficient excitation energy from the adjacent antenna pigments, an electron is transferred to the primary electron-acceptor molecule, pheophytin, through a process called photoinduced charge separation. These electrons are shuttled through an electron transport chain, the so-called Z-scheme shown in the diagram, that initially functions to generate a chemiosmotic potential across the membrane. An ATP synthase enzyme uses the chemiosmotic potential to make ATP during photophosphorylation, whereas NADPH is a product of the terminal redox reaction in the Z-scheme. The electron enters a chlorophyll molecule in Photosystem I. The electron is excited due to the light absorbed by the photosystem. A second electron carrier accepts the electron, which again is passed down lowering energies of electron acceptors. The energy created by the electron acceptors is used to move hydrogen ions across the thylakoid membrane into the lumen. The electron is used to reduce the co-enzyme NADP, which has functions in the light-independent reaction. The cyclic reaction is similar to that of the non-cyclic, but differs in the form that it generates only ATP, and no reduced NADP (NADPH) is created. The cyclic reaction takes place only at photosystem I. Once the electron is displaced from the photosystem, the electron is passed down the electron acceptor molecules and returns to photosystem I, from where it was emitted, hence the name cyclic reaction.

Water photolysis
The NADPH is the main reducing agent in chloroplasts, providing a source of energetic electrons to other reactions. Its production leaves chlorophyll with a deficit of electrons (oxidized), which must be obtained from some other reducing agent. The excited electrons lost from chlorophyll in photosystem I are replaced from the electron transport chain by plastocyanin. However, since photosystem II includes the first steps of the Z-scheme, an external source of electrons is required to reduce its oxidized chlorophyll a molecules. The source of electrons in green-plant and cyanobacterial photosynthesis is water. Two water molecules are oxidized by four successive charge-separation reactions by photosystem II to yield a molecule of diatomic oxygen and four hydrogen ions; the electron yielded in each step is transferred to a redox-active tyrosine residue that then reduces the photoxidized paired-chlorophyll a species called P680 that serves as the primary (light-driven) electron donor in the photosystem II reaction center. The oxidation of water is catalyzed in photosystem II by a redox-active structure that contains four manganese ions and a calcium ion; this oxygen-evolving complex binds two water molecules and stores the four oxidizing equivalents that are required to drive the water-oxidizing reaction. Photosystem II is the only known biological enzyme that carries out this oxidation of water. The hydrogen ions contribute to the transmembrane chemiosmotic potential that leads to ATP synthesis. Oxygen is a waste product of light-dependent reactions, but the majority of organisms on Earth use

Photosynthesis oxygen for cellular respiration, including photosynthetic organisms.[22] [23]

274

Light-independent reactions
The Calvin Cycle
In the light-independent or dark reactions the enzyme RuBisCO captures CO2 from the atmosphere and in a process that requires the newly formed NADPH, called the Calvin-Benson Cycle, releases three-carbon sugars, which are later combined to form sucrose and starch. The overall equation for the light-independent reactions in green plants is:[21] 3 CO2 + 9 ATP + 6 NADPH + 6 H+ C3H6O3-phosphate + 9 ADP + 8 Pi + 6 NADP+ + 3 H2O To be more specific, carbon fixation produces an intermediate product, which is then converted to the final carbohydrate products. The carbon skeletons produced by photosynthesis are then variously used to form other organic compounds, such as the building material cellulose, as precursors for lipid and amino acid biosynthesis, or as a fuel in cellular respiration. The latter occurs not only in plants but also in animals when the energy from plants gets passed through a food chain. The fixation or reduction of carbon dioxide is a process in which carbon dioxide combines with a five-carbon sugar, ribulose 1,5-bisphosphate Overview of the Calvin cycle and carbon fixation (RuBP), to yield two molecules of a three-carbon compound, glycerate 3-phosphate (GP), also known as 3-phosphoglycerate (PGA). GP, in the presence of ATP and NADPH from the light-dependent stages, is reduced to glyceraldehyde 3-phosphate (G3P). This product is also referred to as 3-phosphoglyceraldehyde (PGAL) or even as triose phosphate. Triose is a 3-carbon sugar (see carbohydrates). Most (5 out of 6 molecules) of the G3P produced is used to regenerate RuBP so the process can continue (see Calvin-Benson cycle). The 1 out of 6 molecules of the triose phosphates not "recycled" often condense to form hexose phosphates, which ultimately yield sucrose, starch and cellulose. The sugars produced during carbon metabolism yield carbon skeletons that can be used for other metabolic reactions like the production of amino acids and lipids.

Photosynthesis

275

Carbon concentrating mechanisms

On land In hot and dry conditions, plants close their stomata to prevent the loss of water. Under these conditions, CO2 will decrease, and oxygen gas, produced by the light reactions of photosynthesis, will decrease in the stem, not leaves, causing an increase of photorespiration by the oxygenase activity of ribulose-1,5-bisphosphate carboxylase/oxygenase and decrease in carbon fixation. Some plants have evolved mechanisms to increase the CO2 concentration in the leaves under these conditions. C4 plants chemically fix carbon dioxide in the cells of the mesophyll by adding it to the three-carbon molecule phosphoenolpyruvate (PEP), a reaction catalyzed by an enzyme called PEP carboxylase, creating the four-carbon organic acid oxaloacetic acid. Oxaloacetic acid or malate synthesized by this process is then translocated to specialized bundle sheath cells where the enzyme RuBisCO and other Calvin cycle enzymes are located, and where CO2 released by decarboxylation of the four-carbon acids is then fixed by RuBisCO activity to the three-carbon sugar 3-phosphoglyceric acids. The physical separation of RuBisCO from the Overview of C4 carbon fixation oxygen-generating light reactions reduces photorespiration and increases CO2 fixation and, thus, photosynthetic capacity of the leaf.[24] C4 plants can produce more sugar than C3 plants in conditions of high light and temperature. Many important crop plants are C4 plants, including maize, sorghum, sugarcane, and millet. Plants that do not use PEP-carboxylase in carbon fixation are called C3 plants because the primary carboxylation reaction, catalyzed by RuBisCO, produces the three-carbon sugar 3-phosphoglyceric acids directly in the Calvin-Benson cycle. Over 90% of plants use C3 carbon fixation, compared to 3% that use C4 carbon fixation.[25] Xerophytes, such as cacti and most succulents, also use PEP carboxylase to capture carbon dioxide in a process called Crassulacean acid metabolism (CAM). In contrast to C4 metabolism, which physically separates the CO2 fixation to PEP from the Calvin cycle, CAM temporally separates these two processes. CAM plants have a different leaf anatomy from C3 plants, and fix the CO2 at night, when their stomata are open. CAM plants store the CO2 mostly in the form of malic acid via carboxylation of phosphoenolpyruvate to oxaloacetate, which is then reduced to malate. Decarboxylation of malate during the day releases CO2 inside the leaves, thus allowing carbon fixation to 3-phosphoglycerate by RuBisCO. Sixteen thousand species of plants use CAM.[26]

Photosynthesis In water Cyanobacteria possess carboxysomes, which increase the concentration of CO2 around RuBisCO to increase the rate of photosynthesis. This operates by carbonic anhydrase, producing hydrocarbonate ions (HCO3), which are then pumped into the carboxysome, before being processed by a different carbonic anhydrase to produce CO2.[27] Pyrenoids in algae and hornworts also act to concentrate CO2 around rubisco.[28]

276

Order and kinetics

The overall process of photosynthesis takes place in four stages:[6]
Stage 1 2 3 4 Description Energy transfer in antenna chlorophyll (thylakoid membranes) Time scale femtosecond to picosecond

Transfer of electrons in photochemical reactions (thylakoid membranes) picosecond to nanosecond Electron transport chain and ATP synthesis (thylakoid membranes) Carbon fixation and export of stable products microsecond to millisecond millisecond to second

Efficiency
Plants usually convert light into chemical energy with a photosynthetic efficiency of 36%.[29] Actual plants' photosynthetic efficiency varies with the frequency of the light being converted, light intensity, temperature and proportion of carbon dioxide in the atmosphere, and can vary from 0.1% to 8%.[30] By comparison, solar panels convert light into electric energy at an efficiency of approximately 620% for mass-produced panels, and above 40% in laboratory devices.

Evolution
Early photosynthetic systems, such as those from green and purple sulfur and green and purple nonsulfur bacteria, are thought to have been anoxygenic, using various molecules as electron donors. Green and purple sulfur bacteria are thought to have used hydrogen and sulfur as an electron donor. Green nonsulfur bacteria used various amino and other organic acids. Purple nonsulfur bacteria used a variety of nonspecific organic molecules. The use of these molecules is consistent with the geological evidence that the atmosphere was highly reduced at that time. Fossils of what are thought to be filamentous photosynthetic organisms have been dated at 3.4 billion years old.[31] [32]
Plant cells with visible chloroplasts (from a moss, Plagiomnium affine)

The main source of oxygen in the atmosphere is oxygenic photosynthesis, and its first appearance is sometimes referred to as the oxygen catastrophe. Geological evidence suggests that oxygenic photosynthesis, such as that in cyanobacteria, became important during the Paleoproterozoic era around 2 billion years ago. Modern photosynthesis

Photosynthesis in plants and most photosynthetic prokaryotes is oxygenic. Oxygenic photosynthesis uses water as an electron donor, which is oxidized to molecular oxygen (O2) in the photosynthetic reaction center.

277

Symbiosis and the origin of chloroplasts

Several groups of animals have formed symbiotic relationships with photosynthetic algae. These are most common in corals, sponges and sea anemones. It is presumed that this is due to the particularly simple body plans and large surface areas of these animals compared to their volumes.[33] In addition, a few marine mollusks Elysia viridis and Elysia chlorotica also maintain a symbiotic relationship with chloroplasts they capture from the algae in their diet and then store in their bodies. This allows the mollusks to survive solely by photosynthesis for several months at a time.[34] [35] Some of the genes from the plant cell nucleus have even been transferred to the slugs, so that the chloroplasts can be supplied with proteins that they need to survive.[36] An even closer form of symbiosis may explain the origin of chloroplasts. Chloroplasts have many similarities with photosynthetic bacteria, including a circular chromosome, prokaryotic-type ribosomes, and similar proteins in the photosynthetic reaction center.[37] [38] The endosymbiotic theory suggests that photosynthetic bacteria were acquired (by endocytosis) by early eukaryotic cells to form the first plant cells. Therefore, chloroplasts may be photosynthetic bacteria that adapted to life inside plant cells. Like mitochondria, chloroplasts still possess their own DNA, separate from the nuclear DNA of their plant host cells and the genes in this chloroplast DNA resemble those in cyanobacteria.[39] DNA in chloroplasts codes for redox proteins such as photosynthetic reaction centers. The CoRR Hypothesis proposes that this Co-location is required for Redox Regulation.

Cyanobacteria and the evolution of photosynthesis

The biochemical capacity to use water as the source for electrons in photosynthesis evolved once, in a common ancestor of extant cyanobacteria. The geological record indicates that this transforming event took place early in Earth's history, at least 24502320 million years ago (Ma), and, it is speculated, much earlier.[40] Available evidence from geobiological studies of Archean (>2500 Ma) sedimentary rocks indicates that life existed 3500 Ma, but the question of when oxygenic photosynthesis evolved is still unanswered. A clear paleontological window on cyanobacterial evolution opened about 2000 Ma, revealing an already-diverse biota of blue-greens. Cyanobacteria remained principal primary producers throughout the Proterozoic Eon (2500543 Ma), in part because the redox structure of the oceans favored photoautotrophs capable of nitrogen fixation. Green algae joined blue-greens as major primary producers on continental shelves near the end of the Proterozoic, but only with the Mesozoic (25165 Ma) radiations of dinoflagellates, coccolithophorids, and diatoms did primary production in marine shelf waters take modern form. Cyanobacteria remain critical to marine ecosystems as primary producers in oceanic gyres, as agents of biological nitrogen fixation, and, in modified form, as the plastids of marine algae.[41] A 2010 study by researchers at Tel Aviv University discovered that the Oriental hornet (Vespa orientalis) converts sunlight into electric power using a pigment called xanthopterin. This is the first scientific evidence of a member of the animal kingdom engaging in photosynthesis.[42]

Discovery
Although some of the steps in photosynthesis are still not completely understood, the overall photosynthetic equation has been known since the 19th century. Jan van Helmont began the research of the process in the mid-17th century when he carefully measured the mass of the soil used by a plant and the mass of the plant as it grew. After noticing that the soil mass changed very little, he hypothesized that the mass of the growing plant must come from the water, the only substance he added to the potted plant. His hypothesis was partially accurate much of the gained mass also comes from carbon dioxide as well as water. However, this was a signaling point to the idea that the bulk of a plant's biomass comes from the inputs of photosynthesis, not the soil itself.

Photosynthesis Joseph Priestley, a chemist and minister, discovered that, when he isolated a volume of air under an inverted jar, and burned a candle in it, the candle would burn out very quickly, much before it ran out of wax. He further discovered that a mouse could similarly "injure" air. He then showed that the air that had been "injured" by the candle and the mouse could be restored by a plant. In 1778, Jan Ingenhousz, court physician to the Austrian Empress, repeated Priestley's experiments. He discovered that it was the influence of sunlight on the plant that could cause it to revive a mouse in a matter of hours. In 1796, Jean Senebier, a Swiss pastor, botanist, and naturalist, demonstrated that green plants consume carbon dioxide and release oxygen under the influence of light. Soon afterward, Nicolas-Thodore de Saussure showed that the increase in mass of the plant as it grows could not be due only to uptake of CO2 but also to the incorporation of water. Thus, the basic reaction by which photosynthesis is used to produce food (such as glucose) was outlined. Cornelis Van Niel made key discoveries explaining the chemistry of photosynthesis. By studying purple sulfur bacteria and green bacteria he was the first scientist to demonstrate that photosynthesis is a light-dependent redox reaction, in which hydrogen reduces carbon dioxide. Robert Emerson discovered two light reactions by testing plant productivity using different wavelengths of light. With the red alone, the light reactions were suppressed. When blue and red were combined, the output was much more substantial. Thus, there were two photosystems, one absorbing up to 600nm wavelengths, the other up to 700 nm. The former is known as PSII, the latter is PSI. PSI contains only chlorophyll a, PSII contains primarily chlorophyll a with most of the available chlorophyll b, among other pigment. These include phycobilins, which are the red and blue pigments of red and blue algae respectively, and fucoxanthol for brown algae and diatoms. The process is most productive when absorption of quanta are equal in both the PSII and PSI, assuring that input energy from the antenna complex is divided between the PSI and PSII system, which in turn powers the photochemistry.[6] Robert Hill thought that a complex of reactions consisting of an intermediate to cytochrome b6 (now a plastoquinone), another is from cytochrome f to a step in the carbohydrate-generating mechanisms. These are linked by plastoquinone, which does require energy to reduce cytochrome f for it is a sufficient reductant. Further experiments to prove that the oxygen developed during the photosynthesis of green plants came from water, were performed by Hill in 1937 and 1939. He showed that isolated chloroplasts give off oxygen in the presence of unnatural reducing agents like iron oxalate, ferricyanide or benzoquinone after exposure to light. The Hill reaction is as follows: 2 H2O + 2 A + (light, chloroplasts) 2 AH2 + O2 where A is the electron acceptor. Therefore, in light, the electron acceptor is reduced and oxygen is evolved. Samuel Ruben and Martin Kamen used radioactive isotopes to determine that the oxygen liberated in photosynthesis came from the water. Melvin Calvin and Andrew Benson, along with James Bassham, elucidated the path of carbon assimilation (the photosynthetic carbon reduction cycle) in plants. The carbon reduction cycle is known as the Calvin cycle, which ignores the contribution of Bassham and Benson. Many scientists refer to the cycle as the Calvin-Benson Cycle, Benson-Calvin, and some even call it the Calvin-Benson-Bassham (or CBB) Cycle. Nobel Prize-winning scientist Rudolph A. Marcus was able to discover the function and significance of the electron transport chain. Otto Heinrich Warburg and Dean Burk discovered the I-quantum photosynthesis reaction that splits the CO2, activated by the respiration.[43] Louis N.M. Duysens and Jan Amesz discovered that chlorophyll a will absorb one light, oxidize cytochrome f, chlorophyll a (and other pigments) will absorb another light, but will reduce this same oxidized cytochrome, stating the two light reactions are in series.

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Photosynthesis

279

Factors
There are three main factors affecting photosynthesis and several corollary factors. The three main are: Light irradiance and wavelength Carbon dioxide concentration Temperature.

Light intensity (irradiance), wavelength and temperature

In the early 20th century, Frederick Frost Blackman along with Albert Einstein investigated the effects of light intensity (irradiance) and temperature on the rate of carbon assimilation.
The leaf is the primary site of photosynthesis in plants.

At constant temperature, the rate of carbon assimilation varies with irradiance, initially increasing as the irradiance increases. However, at higher irradiance, this relationship no longer holds and the rate of carbon assimilation reaches a plateau. At constant irradiance, the rate of carbon assimilation increases as the temperature is increased over a limited range. This effect is seen only at high irradiance levels. At low irradiance, increasing the temperature has little influence on the rate of carbon assimilation. These two experiments illustrate vital points: First, from research it is known that, in general, photochemical reactions are not affected by temperature. However, these experiments clearly show that temperature affects the rate of carbon assimilation, so there must be two sets of reactions in the full process of carbon assimilation. These are, of course, the light-dependent 'photochemical' stage and the light-independent, temperature-dependent stage. Second, Blackman's experiments illustrate the concept of limiting factors. Another limiting factor is the wavelength of light. Cyanobacteria, which reside several meters underwater, cannot receive the correct wavelengths required to cause photoinduced charge separation in conventional photosynthetic pigments. To combat this problem, a series of proteins with different pigments surround the reaction center. This unit is called a phycobilisome.

Carbon dioxide levels and photorespiration

As carbon dioxide concentrations rise, the rate at which sugars are made by the light-independent reactions increases until limited by other factors. RuBisCO, the enzyme that captures carbon dioxide in the light-independent reactions, has a binding affinity for both carbon dioxide and oxygen. When the concentration of carbon dioxide is high, RuBisCO will fix carbon dioxide. However, if the carbon dioxide concentration is low, RuBisCO will bind oxygen instead of carbon dioxide. This process, called photorespiration, uses energy, but does not produce sugars. RuBisCO oxygenase activity is disadvantageous to plants for several reasons: 1. One product of oxygenase activity is phosphoglycolate (2 carbon) instead of 3-phosphoglycerate (3 carbon). Phosphoglycolate cannot be metabolized by the Calvin-Benson cycle and represents carbon lost from the cycle. A high oxygenase activity, therefore, drains the sugars that are required to recycle ribulose 5-bisphosphate and for the continuation of the Calvin-Benson cycle. 2. Phosphoglycolate is quickly metabolized to glycolate that is toxic to a plant at a high concentration; it inhibits photosynthesis. 3. Salvaging glycolate is an energetically expensive process that uses the glycolate pathway, and only 75% of the carbon is returned to the Calvin-Benson cycle as 3-phosphoglycerate. The reactions also produce ammonia (NH3), which is able to diffuse out of the plant, leading to a loss of nitrogen. A highly simplified summary is:

Photosynthesis 2 glycolate + ATP 3-phosphoglycerate + carbon dioxide + ADP + NH3 The salvaging pathway for the products of RuBisCO oxygenase activity is more commonly known as photorespiration, since it is characterized by light-dependent oxygen consumption and the release of carbon dioxide.

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References
[1] Smith, A. L. (1997). Oxford dictionary of biochemistry and molecular biology. Oxford [Oxfordshire]: Oxford University Press. p.508. ISBN0-19-854768-4. "Photosynthesis the synthesis by organisms of organic chemical compounds, esp. carbohydrates, from carbon dioxide using energy obtained from light rather than the oxidation of chemical compounds." [2] D.A. Bryant & N.-U. Frigaard (2006). "Prokaryotic photosynthesis and phototrophy illuminated". Trends Microbiol 14 (11): 48896. doi:10.1016/j.tim.2006.09.001. PMID16997562. [3] Nealson KH, Conrad PG (1999). "Life: past, present and future". Philos. Trans. R. Soc. Lond., B, Biol. Sci. 354 (1392): 192339. doi:10.1098/rstb.1999.0532. PMC1692713. PMID10670014. [4] "World Consumption of Primary Energy by Energy Type and Selected Country Groups, 19802004" (http:/ / www. eia. doe. gov/ pub/ international/ iealf/ table18. xls) (XLS). Energy Information Administration. July 31, 2006. . Retrieved 2007-01-20. [5] Field CB, Behrenfeld MJ, Randerson JT, Falkowski P (1998). "Primary production of the biosphere: integrating terrestrial and oceanic components". Science 281 (5374): 23740. doi:10.1126/science.281.5374.237. PMID9657713. [6] Photosynthesis, McGraw-Hill Encyclopedia of Science and Technology, Vol. 13, 2007 [7] http:/ / toolserver. org/ ~verisimilus/ Timeline/ Timeline. php?Ma=3500 [8] Olson JM (2006). "Photosynthesis in the Archean era". Photosyn. Res. 88 (2): 10917. doi:10.1007/s11120-006-9040-5. PMID16453059. [9] http:/ / toolserver. org/ ~verisimilus/ Timeline/ Timeline. php?Ma=3000 [10] http:/ / toolserver. org/ ~verisimilus/ Timeline/ Timeline. php?Ma=2400 [11] Buick R (2008). "When did oxygenic photosynthesis evolve?". Philos. Trans. R. Soc. Lond., B, Biol. Sci. 363 (1504): 273143. doi:10.1098/rstb.2008.0041. PMC2606769. PMID18468984. [12] Rodrguez-Ezpeleta, Naiara; Henner Brinkmann, Suzanne C Burey, Batrice Roure, Gertraud Burger, Wolfgang Lffelhardt, Hans J Bohnert, Herv Philippe, B Franz Lang (2005-07-26). "Monophyly of primary photosynthetic eukaryotes: green plants, red algae, and glaucophytes" (http:/ / www. ncbi. nlm. nih. gov/ pubmed/ 16051178). Current Biology: CB 15 (14): 13251330. doi:10.1016/j.cub.2005.06.040. ISSN0960-9822. PMID16051178. . Retrieved 2009-08-26. [13] Gould SB, Waller RF, McFadden GI (2008). "Plastid evolution". Annu Rev Plant Biol 59: 491517. doi:10.1146/annurev.arplant.59.032607.092915. PMID18315522. [14] Anaerobic Photosynthesis, Chemical & Engineering News, 86, 33, August 18, 2008, p. 36 [15] Kulp TR, Hoeft SE, Asao M, Madigan MT, Hollibaugh JT, Fisher JC, Stolz JF, Culbertson CW, Miller LG, Oremland RS (2008). "Arsenic(III) fuels anoxygenic photosynthesis in hot spring biofilms from Mono Lake, California". Science 321 (5891): 96770. doi:10.1126/science.1160799. PMID18703741. [16] "Scientists discover unique microbe in California's largest lake" (http:/ / www. bio-medicine. org/ biology-news/ Scientists-discover-unique-microbe-in-Californias-largest-lake-203-1/ ). . Retrieved 2009-07-20. [17] Tavano CL, Donohue TJ (2006). "Development of the bacterial photosynthetic apparatus". Curr. Opin. Microbiol. 9 (6): 62531. doi:10.1016/j.mib.2006.10.005. PMC2765710. PMID17055774. [18] Mullineaux CW (1999). "The thylakoid membranes of cyanobacteria: structure, dynamics and function". Australian Journal of Plant Physiology 26 (7): 671677. doi:10.1071/PP99027. [19] Sener MK, Olsen JD, Hunter CN, Schulten K (2007). "Atomic-level structural and functional model of a bacterial photosynthetic membrane vesicle". Proc. Natl. Acad. Sci. U.S.A. 104 (40): 157238. doi:10.1073/pnas.0706861104. PMC2000399. PMID17895378. [20] Campbell, Neil A.; Brad Williamson; Robin J. Heyden (2006). Biology: Exploring Life (http:/ / www. phschool. com/ el_marketing. html). Boston, Massachusetts: Pearson Prentice Hall. ISBN0-13-250882-6. . [21] Raven, Peter H.; Ray F. Evert, Susan E. Eichhorn (2005). Biology of Plants, 7th Edition. New York: W.H. Freeman and Company Publishers. pp.124127. ISBN0-7167-1007-2. [22] "Yachandra Group Home page" (http:/ / www. lbl. gov/ ~vkyachan/ index. html). . [23] Pushkar Y, Yano J, Sauer K, Boussac A, Yachandra VK (2008). "Structural changes in the Mn4Ca cluster and the mechanism of photosynthetic water splitting". Proc. Natl. Acad. Sci. U.S.A. 105 (6): 187984. doi:10.1073/pnas.0707092105. PMC2542863. PMID18250316. [24] L. Taiz, E. Zeiger (2006). Plant Physiology (4 ed.). Sinauer Associates. ISBN978-0878938568. [25] Sage, Rowan; Russell Monson (1999). "16" (http:/ / books. google. com/ ?id=H7Wv9ZImW-QC& pg=PA551). C4 Plant Biology. pp.551580. ISBN0126144400. . [26] Dodd, A. N.; Borland, AM; Haslam, RP; Griffiths, H; Maxwell, K (2002). "Crassulacean acid metabolism: plastic, fantastic". Journal of Experimental Botany 53 (369): 56980. doi:10.1093/jexbot/53.369.569. PMID11886877. [27] Badger, M. R.; Price, GD (2003). "CO2 concentrating mechanisms in cyanobacteria: molecular components, their diversity and evolution". Journal of Experimental Botany 54 (383): 60922. doi:10.1093/jxb/erg076. PMID12554704.

Photosynthesis
[28] Badger, Murray R.; Andrews, T. John; Whitney, S.M.; Ludwig, Martha; Yellowlees, David C.; Leggat, W.; Price, G. Dean (1998). "The diversity and coevolution of Rubisco, plastids, pyrenoids, and chloroplast-based CO2-concentrating mechanisms in algae". Canadian Journal of Botany 76 (6): 1052. doi:10.1139/cjb-76-6-1052. [29] Chapter 1 Biological energy production">Miyamoto K. "Chapter 1 Biological energy production" (http:/ / www. fao. org/ docrep/ w7241e/ w7241e05. htm#1. 2. 1 photosynthetic efficiency). Renewable biological systems for alternative sustainable energy production (FAO Agricultural Services Bulletin 128). Food and Agriculture Organization of the United Nations. . Retrieved 2009-01-04. [30] Govindjee, What is photosynthesis? (http:/ / www. life. uiuc. edu/ govindjee/ whatisit. htm) [31] Photosynthesis got a really early start (http:/ / www. newscientist. com/ article/ mg18424671. 600-photosynthesis-got-a-really-early-start. html), New Scientist, 2 October 2004 [32] Revealing the dawn of photosynthesis (http:/ / www. newscientist. com/ article/ mg19125654. 200-revealing-the-dawn-of-photosynthesis. html), New Scientist, 19 August 2006 [33] Venn AA, Loram JE, Douglas AE (2008). "Photosynthetic symbioses in animals". J. Exp. Bot. 59 (5): 106980. doi:10.1093/jxb/erm328. PMID18267943. [34] Rumpho ME, Summer EJ, Manhart JR (2000). "Solar-Powered Sea Slugs. Mollusc/Algal Chloroplast Symbiosis". Plant Physiol. 123 (1): 2938. doi:10.1104/pp.123.1.29. PMC1539252. PMID10806222. [35] Muscatine L, Greene RW (1973). "Chloroplasts and algae as symbionts in molluscs". Int. Rev. Cytol. 36: 13769. doi:10.1016/S0074-7696(08)60217-X. PMID4587388. [36] Rumpho ME, Worful JM, Lee J et al. (2008). "Horizontal gene transfer of the algal nuclear gene psbO to the photosynthetic sea slug Elysia chlorotica". Proc. Natl. Acad. Sci. U.S.A. 105 (46): 1786717871. doi:10.1073/pnas.0804968105. PMC2584685. PMID19004808. [37] Douglas SE (1998). "Plastid evolution: origins, diversity, trends". Curr. Opin. Genet. Dev. 8 (6): 65561. doi:10.1016/S0959-437X(98)80033-6. PMID9914199. [38] Reyes-Prieto A, Weber AP, Bhattacharya D (2007). "The origin and establishment of the plastid in algae and plants". Annu. Rev. Genet. 41: 14768. doi:10.1146/annurev.genet.41.110306.130134. PMID17600460. [39] Raven JA, Allen JF (2003). "Genomics and chloroplast evolution: what did cyanobacteria do for plants?". Genome Biol. 4 (3): 209. doi:10.1186/gb-2003-4-3-209. PMC153454. PMID12620099. [40] "Cyanobacteria: Fossil Record" (http:/ / www. ucmp. berkeley. edu/ bacteria/ cyanofr. html). Ucmp.berkeley.edu. . Retrieved 2010-08-26. [41] Herrero A and Flores E (editor). (2008). The Cyanobacteria: Molecular Biology, Genomics and Evolution (1st ed.). Caister Academic Press. ISBN978-1-904455-15-8. [42] Plotkin, M.; Hod, I.; Zaban, A.; Boden, S. A.; Bagnall, D. M.; Galushko, D.; Bergman, D. J. (2010). "Solar energy harvesting in the epicuticle of the oriental hornet (Vespa orientalis)". Naturwissenschaften 97 (12): 1067. Bibcode2010NW.....97.1067P. doi:10.1007/s00114-010-0728-1. PMID21052618. [43] Otto Warburg Biography (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1931/ warburg. html). Nobelprize.org (1970-08-01). Retrieved on 2011-11-03.

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Further reading
Books
Asimov, Isaac (1968). Photosynthesis. New York, London: Basic Books, Inc.. ISBN0-465-05703-9. Bidlack JE; Stern KR, Jansky S (2003). Introductory plant biology. New York: McGraw-Hill. ISBN0-07-290941-2. Blankenship RE (2008). Molecular Mechanisms of Photosynthesis (2nd ed.). John Wiley & Sons Inc. ISBN0-470-71451-4. Govindjee (1975). Bioenergetics of photosynthesis. Boston: Academic Press. ISBN0-12-294350-3. Govindjee Beatty JT,Gest H, Allen JF (2006). Discoveries in Photosynthesis. Advances in Photosynthesis and Respiration. 20. Berlin: Springer. ISBN1-4020-3323-0. Gregory RL (1971). Biochemistry of photosynthesis. New York: Wiley-Interscience. ISBN0-471-32675-5. Rabinowitch E, Govindjee (1969). Photosynthesis. London: J. Wiley. ISBN0-471-70424-5. Reece, J, Campbell, N (2005). Biology. San Francisco: Pearson, Benjamin Cummings. ISBN0-8053-7146-X.

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282

Papers
Evolutionary relationships among photosynthetic prokaryotes : implications regarding the origin of photosynthesis. (http://www.ncbi.nlm.nih.gov/pubmed/10361294) Origin and early evolution of photosynthesis. (http://www.ncbi.nlm.nih.gov/pubmed/11538390) Photosystem II: evolutionary perspectives (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1693113/ ?tool=pmcentrez)

External links
A collection of photosynthesis pages for all levels from a renowned expert (Govindjee) (http://www.life.uiuc. edu/govindjee/linksPSed.htm) In depth, advanced treatment of photosynthesis, also from Govindjee (http://www.life.uiuc.edu/govindjee/ paper/gov.html) Science Aid: Photosynthesis (http://scienceaid.co.uk/biology/biochemistry/photosynthesis.html) Article appropriate for high school science Metabolism, Cellular Respiration and Photosynthesis The Virtual Library of Biochemistry and Cell Biology (http://www.biochemweb.org/metabolism.shtml) Overall examination of Photosynthesis at an intermediate level (http://www.chemsoc.org/networks/learnnet/ cfb/Photosynthesis.htm) Overall Energetics of Photosynthesis (http://www.life.uiuc.edu/govindjee/photosynBook.html) Photosynthesis Discovery Milestones (http://www.juliantrubin.com/bigten/photosynthesisexperiments.html) experiments and background The source of oxygen produced by photosynthesis (http://bcs.whfreeman.com/thelifewire/content/chp08/ 0802001.html) Interactive animation, a textbook tutorial Jessica Marshall (2011-03-29). "First practical artificial leaf makes debut" (http://news.discovery.com/earth/ artificial-leaf-technology-solar-110329.html). Discovery News. Photosynthesis Light Dependent & Light Independent Stages (http://www.biology-innovation.co.uk/pages/ plant-biology-ecology/photosynthesis/) Khan Academy, video introduction (http://www.khanacademy.org/video/photosynthesis?playlist=Biology)

283

Lipid metabolism
Fatty acid synthesis
Fatty acid synthesis is the creation of fatty acids from acetyl-CoA and malonyl-CoA precursors through action of enzymes called fatty acid synthases. It is an important part of the lipogenesis process, which - together with glycolysis - stands behind creating fats from blood sugar in living organisms.

Straight-Chain Fatty Acids

Straight-chain fatty acids occur in two types; saturated and unsaturated.

Saturated Straight-Chain Fatty Acids

Much like -oxidation, straight-chain fatty acid synthesis occurs via the six recurring reactions shown below, until the 16 carbon palmitic acid is produced.[1] The diagrams presented show how fatty acids are synthesized in microorganisms and list the enzymes found in Escherichia coli.[1] These reactions are performed by fatty acid synthase II (FASII), which contain multiple enzymes that generally act as one complex. FASII is present in prokaryotes, plants, fungi, parasites, as well as in mitochondria.[2] In animals, as well as yeast and some fungi, these same reactions occur on fatty acid synthase I (FASI), a large dimeric protein, which has all of the enzymatic activities required to create a fatty acid. FASI is less efficient than FASII, however, it allows for the formation of more molecules, including medium-chain fatty acids via early chain termination.[2] Once a 16:0 carbon fatty acid has been formed it can undergo a number of modifications; particularly, by fatty acid synthase III (FASIII), which uses 2 carbon molecules to elongate preformed fatty acids.[2]
Step Enzyme Reaction Description Synthesis of saturated fatty acids via Fatty Acid Synthase II in E. coli

(a)

Acetyl CoA:ACP transacylase

Activates acetyl CoA for reaction with malonyl-ACP

(b)

Malonyl CoA:ACP transacylase

Activates malonyl CoA for reaction with acetyl-ACP

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284
Reacts priming acetyl-ACP with chain extending malonyl-ACP.

(c)

3-ketoacyl-ACP synthetase

(d)

3-ketoacyl-ACP reductase

Reduces the carbon 3 ketone to a hydroxyl group

(e)

3-hydroxyacyl-ACP dehydrase

Removes water

(f)

Enoyl-ACP reductase

Reduces the C3-C4 double bond.

Regulation Acetyl-CoA is formed into malonyl-CoA by acetyl-CoA carboxylase, at which point malonyl-CoA is destined to feed into the fatty acid synthesis pathway. Acetyl-CoA carboxylase is the point of regulation in saturated straight-chain fatty acid synthesis, by both phosphorylation and allosteric regulation. Regulation by phosphorylation occurs mostly in mammals, while allosteric regulation occurs in most organisms. Allosteric control occurs as feedback inhibition by palmitol-CoA and activation by citrate. When there are high levels of palmitol-CoA, the final product of saturated fatty acid synthesis, it allosterically inactivates acetyl-CoA carboxylase to prevent a build up of fatty acids in cells. Citrate acts to activate acetyl-CoA carboxylase under high levels, because high levels indicate there is enough acetyl-CoA to feed into the Krebs cycle and produce energy.[3] De Novo Synthesis in Humans In humans, fatty acids are predominantly formed in the liver and lactating mammary glands, and, to a lesser extent, the adipose tissue. Most acetyl-CoA is formed from pyruvate by pyruvate dehydrogenase in the mitochondria. Acetyl-CoA produced in the mitochondria is condensed with oxaloacetate by citrate synthase to form citrate, which is then transported into the cytosol and broken down to yield acetyl-CoA and oxaloacetate by ATP citrate lyase. Oxaloacetate in the cytosol is reduced to malate by cytoplasmic malate dehydrogenase, and malate is transported back into the mitochondria to participate in the Citric acid cycle.[4]

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285

Desaturation
Unsaturated fatty acids are essential components to prokaryotic and eukaryotic cell membranes. These fatty acids primarily function in maintaining membrane fluidity.[5] They have also been associated with serving as signaling molecules in other processes such as cell differentiation and DNA replication.[5] There are two pathways organisms use for desaturation: Aerobic and Anaerobic. Anaerobic Desaturation Many bacteria use the anaerobic pathway for synthesizing unsaturated fatty acids. This pathway does not utilize oxygen and is dependent on enzymes to insert the double bond before elongation utilizing the normal fatty acid synthesis machinery. In Escherichia coli, this pathway is well understood. FabA is a -hydroxydecanoyl-ACP dehydrase- it is specific for the 10 carbon saturated fatty acid synthesis intermediate (-hydroxydecanoyl-ACP). FabA catalyzes the dehydration of -hydroxydecanoyl-ACP causing the release of water and insertion of the double bond between C7 and C8 counting from the methyl end. This creates the trans-2-decenoyl intermediate. The trans-2-decenoyl intermediate can either be shunted to the normal saturated fatty acid synthesis pathway by FabB, where the double bond will be hydrolyzed and the final product will be a saturated fatty acid, or FabA will catalyze the isomerization into the Synthesis of unsaturated fatty acids via anaerobic cis-3-decenoyl intermediate. desaturation FabB is a -ketoacyl-ACP synthase which elongates and channels intermediates into the mainstream fatty acid synthesis pathway. When FabB reacts with the cis-decenoyl intermediate the final product after elongation will be an unsaturated fatty acid.[6] The two main unsaturated fatty acids made are Palmitoleoyl-ACP (16:17) and cis-vaccenoyl-ACP (18:17).[7] Most bacteria that undergo anaerobic desaturation contain homologues of FabA and FabB.[8] Clostridia are the main exception; they have a novel enzyme, yet to be identified, that catalyzes the formation of the cis double bond.[7] Regulation This pathway undergoes transcriptional regulation by FadR and FabR. FadR is the more extensively studied protein and has been attributed bifunctional characteristics. It acts as an activator of fabA and fabB transcription and as a repressor for the -oxidation regulon. FabR, conversely, acts as a repressor for the transcription of fabA and fabB.[6]

Fatty acid synthesis Aerobic Desaturation Aerobic desaturation is the most widespread pathway for the synthesis of unsaturated fatty acids. It is utilized in all eukaryotes and some prokaryotes. This pathway utilizes desaturases to synthesize unsaturated fatty acids from full length saturated fatty acid substrates.[9] All desaturases require oxygen and reducing equivalents acquired from the electron transport chain. Desaturases are specific for the double bond they induce in the substrate. In Bacillus subtilis, the desaturase, 5-Des, is specific for inducing a cis-double bond at the 5 position.[5] [9] Saccharomyces cerevisiae contains one desaturase, Ole1p, which induces the cis-double bond at 9.[5] Regulation In B. subtilis, this pathway is regulated by a two-component system: DesK and DesR. DesK is a membrane associated kinase and DesR is a Synthesis of unsaturated fatty acids via aerobic transcriptional regulator of the des gene.[5] [9] The regulation responds desaturation to temperature; when there is a drop in temperature this gene is upregulated. Unsaturated fatty acids decrease the fluidity of the membrane and stabilize it under lower temperatures. DesK is the sensor protein that when there is a decrease in temperature, will autophosphorylate. DesK-P will transfer its phosphoryl group to DesR. Two DesR-P proteins will dimerize and bind to the DNA promoters of the des gene and recruit RNA polymerase to begin transcription.[5] [9] Pseudomonas aeruginosa Generally, both anaerobic and aerobic unsaturated fatty acid synthesis will not occur within the same system, however Pseudomonas aeruginosa and Vibrio ABE-1 are exceptions.[10] [11] [12] While, P. aeruginosa primarily undergoes anaerobic desaturation, it also undergoes two aerobic pathways. One pathway utilizes a 9 desaturase (DesA) that catalyzes a double bond formation in membrane lipids. Another pathway uses two proteins, DesC and DesB, together to act as a 9 desaturase which inserts a double bond into a saturated fatty acid-CoA molecule. This second pathway is regulated by repressor protein, DesT. DesT is also a repressor of fabAB expression for anaerobic desaturation when in presence of exogenous unsaturated fatty acids. This functions to coordinate the expression of the two pathways within the organism.[11] [13]

286

Branched chain fatty acids

Branched-chain fatty acids are usually saturated and are found in two distinct families; the iso-series and anteiso-series. It has been found that Actinomycetales contain unique branch chain fatty acid synthesis mechanisms, including that which forms tuberculosteric acid.

Branch-Chain Fatty Acid Synthesizing System

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287

Valine primer

Leucine primer

Isoleucine primer

The branched-chain fatty acid synthesizing system uses -keto acids as primers. This system is distinct from the branched-chain fatty acid synthetase which utilizes short-chain acyl-CoA esters as primers.[14] -keto acid primers are derived from the transamination and decarboxylation of valine, leucine, and isoleucine to form 2-methylpropanyl-CoA, 3-methylbutyryl-CoA, and 2-Methylbutyryl-CoA, respectively.[15] 2-methylpropanyl-CoA primers derived from valine are elongated to produce even-numbered iso-series fatty acids such as 14-methyl-pentadecanoic (isopalmitic) acid, and 3-methylbutyryl-CoA primers from leucine may be used to form odd numbered iso-series fatty acids such as 13-methyl-tetradecanoic acid. 2-Methylbutyryl-CoA primers from isoleucine are elongated to form anteiso-series fatty acids containing an odd number of carbon atoms such as 12-Methyl tetradecanoic acid.[16] Decarboxylation of the primer precursors occurs through the branched-chain -keto acid decarboxylase (BCKA) enzyme. Elongation of the fatty acid follows the same biosynthetic pathway in Escherichia coli used to produce straight-chain fatty acids where malonyl-CoA is used as a chain extender.[17] The major end products are 12-17 carbon branched-chain fatty acids and their composition tends to be uniform and characteristic for many bacterial species.[16] BCKA decarboxylase and relative activities of -keto acid substrates The BCKA decarboxylase enzyme is composed of two subunits in a tetrameric structure (A2B2) and is essential for the synthesis of branched-chain fatty acids. It is responsible for the decarboxylation of -keto acids formed by the transamination of valine, leucine, and isoleucine and produces the primers used for branched-chain fatty acid synthesis. The activity of this enzyme is much higher with branched-chain -keto acid substrates than straight-chain substrates, and in Bacillus species its specificity is highest for the isoleucine-derived -keto--methylvaleric acid, followed by -ketoisocaproate and -ketoisovalerate.[16] [17] The enzymes high affinity toward branched-chain

Fatty acid synthesis -keto acids allows it to function as the primer donating system for branched-chain fatty acid synthetase.[17]
Substrate BCKA activity CO2 Produced (nmol/min mg) 19.7 12.4 7.4 4.9 Km (M) <1 <1 <1 51.1 Vmax (nmol/min mg) 17.8 13.3 5.6 15.2

288

L--keto--methyl-valerate 100% -Ketoisovalerate -Ketoisocaproate Pyruvate 63% 38% 25%

Factors affecting chain length and pattern distribution -keto acid primers are used to produce branched-chain fatty acids that are generally between 12 and 17 carbons in length. The proportions of these branched-chain fatty acids tend to be uniform and consistent among a particular bacterial species but may be altered due to changes in malonyl-CoA concentration, temperature, or heat-stable factors (HSF) present.[16] All of these factors may affect chain length, and HSFs have been demonstrated to alter the specificity of BCKA decarboxylase for a particular -keto acid substrate, thus shifting the ratio of branched-chain fatty acids produced.[16] An increase in malonyl-CoA concentration has been shown to result in a larger proportion of C17 fatty acids produced, up until the optimal concentration (20M) of malonyl-CoA is reached. Decreased temperatures also tend to shift the fatty-acid distribution slightly toward C17 fatty-acids in Bacillus species.[14] [16]

Branch-Chain Fatty Acid Synthase

This system functions similarly to the branch-chain fatty acid synthesizing system, however it uses short chain carboxylic acids as primers instead of alpha-keto acids. This method is generally used by bacteria that do not have the ability to perform the branch-chain fatty acid system using alpha-keto primers. Typical short chain primers include isovalerate, isobutyrate and 2-methyl butyrate. The acids needed for these primers are generally taken up from the environment; this is often seen in ruminal bacteria.[18] The over all reaction is: Isobutyryl-CoA + 6 malonyl-CoA +12 NADPH + 12H+ = Isoplmitic acid + 6CO2 12NADP + 5H2O + 7CoA[14] The difference between (straight-chain) fatty acid synthase and branch-chain fatty acid synthase is substrate specificity of the enzyme that catalyzes the reaction of acyl-CoA to acyl-ACP.[14]

Omega-alicyclic fatty acids

Omega-alicyclic fatty acids typically contain an omega-terminal propyl or butyryl cyclic group and are some of the major membrane fatty acids found in several species of bacteria. The fatty acid synthetase used to produce omega-alicyclic fatty acids is also used to produce membrane branched-chain fatty acids. In bacteria with membranes composed mainly of omega-alicyclic fatty acids, the supply of cyclic carboxylic acid-CoA esters is much greater than that of branched-chain primers.[14] The synthesis of cyclic primers is not well understood but it has been suggested that mechanism involves the conversion of sugars to shikimic acid which is then converted to cyclohexylcarboxylic acid-CoA esters that serve as primers for omega-alicyclic fatty acid synthesis [18]

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Tuberculostearic Acid Synthesis

Tuberculostearic acid (D-10-Methylstearic acid) is a saturated fatty acid that is known to be produced by Mycobacterium spp. and two species of Streptomyces. It is formed from the precursor oleic acid (a monosaturated fatty acid).[19] After oleic acid is esterified to a phospholipid, S-adenosyl-methionine donates a methyl group to the double bond of oleic acid.[20] This methylation reaction forms the intermediate 10-methylene-octadecanoyal. Successive reduction of the residue, with NADPH as a cofactor, results in 10-methylstearic acid
[15]

References
[1] Dijkstra, Albert J., R. J. Hamilton, and Wolf Hamm. "Fatty Acid Biosynthesis." Trans Fatty Acids. Oxford: Blackwell Pub., 2008. 12. Print. [2] "Fatty Acids: Straight-chain Saturated, Structure, Occurrence and Biosynthesis." Lipid Library - Lipid Chemistry, Biology, Technology and Analysis. Web. 30 Apr. 2011. <http://lipidlibrary.aocs.org/lipids/fa_sat/index.htm>. Mechanism of the synthesis of Tuberculostearic acid

[3] Diwan, Joyce J. "Fatty Acid Synthesis." Rensselaer Polytechnic Institute (RPI) :: Architecture, Business, Engineering, IT, Humanities, Science. Web. 30 Apr. 2011. <http://rpi.edu/dept/bcbp/molbiochem/MBWeb/mb2/part1/fasynthesis.htm>. [4] Ferre, P.; F. Foufelle (2007). "SREBP-1c Transcription Factor and Lipid Homeostasis: Clinical Perspective" (http:/ / content. karger. com/ ProdukteDB/ produkte. asp?Aktion=ShowFulltext& ArtikelNr=100426& Ausgabe=232805& ProduktNr=224036). Hormone Research 68 (2): 7282. doi:10.1159/000100426. PMID17344645. . Retrieved 2010-08-30. "this process is outlined graphically in page 73" [5] Aguilar, Pablo S, and Diegode Mendoza. "Control of fatty acid desaturation: a mechanism conserved from bacteria to humans." Molecular microbiology 62.6 (2006):1507-14. [6] Feng, Youjun, and John ECronan. "Complex binding of the FabR repressor of bacterial unsaturated fatty acid biosynthesis to its cognate promoters." Molecular microbiology 80.1 (2011):195-218. [7] Zhu, Lei, et al. "Functions of the Clostridium acetobutylicium FabF and FabZ proteins in unsaturated fatty acid biosynthesis." BMC microbiology 9(2009):119. [8] Wang, Haihong, and John ECronan. "Functional replacement of the FabA and FabB proteins of Escherichia coli fatty acid synthesis by Enterococcus faecalis FabZ and FabF homologues." Journal of biological chemistry 279.33 (2004):34489-95. [9] Mansilla, Mara C, and Diegode Mendoza. "The Bacillus subtilis desaturase: a model to understand phospholipid modification and temperature sensing." Archives of microbiology 183.4 (2005):229-35. [10] Wada, M, N. Fukunaga, and S. Sasaki. "Mechanism of biosynthesis of unsaturated fatty acids in Pseudomonas sp. strain E-3, a psychrotrophic bacterium." Journal of bacteriology 171.8 (1989):4267-71. [11] Subramanian, Chitra, Charles ORock, and Yong-MeiZhang. "DesT coordinates the expression of anaerobic and aerobic pathways for unsaturated fatty acid biosynthesis in Pseudomonas aeruginosa." Journal of bacteriology 192.1 (2010):280-5. [12] Morita, N, et al. "Both the anaerobic pathway and aerobic desaturation are involved in the synthesis of unsaturated fatty acids in Vibrio sp. strain ABE-1." FEBS letters 297.1-2 (1992):9-12. [13] Zhu, Kun, et al. "Two aerobic pathways for the formation of unsaturated fatty acids in Pseudomonas aeruginosa." Molecular microbiology 60.2 (2006):260-73. [14] Kaneda, Toshi. "Iso- and Anteiso-Fatty Acids in Bacteria: Biosynthesis, Function, and Taxonomic Significance." Microbiological Reviews 55.2 (1991): 288-302 [15] "Branched-chain Fatty Acids, Phytanic Acid, Tuberculostearic Acid Iso/anteiso- Fatty Acids." Lipid Library - Lipid Chemistry, Biology, Technology and Analysis. Web. 01 May 2011. http:/ / lipidlibrary. aocs. org/ lipids/ fa_branc/ index. htm. [16] Naik, Devaray N., and Toshi Kaneda. "Biosynthesis of Branched Long-chain Fatty Acids by Species of Bacillus: Relative Activity of Three -keto Acid Substrates and Factors Affecting Chain Length." Can. J. Microbiol. 20 (1974): 1701-708. [17] Oku, Hirosuke, and Toshi Kaneda. "Biosynthesis of Branched-chain Fatty Acids in Bacillis Subtilis." The Journal of Biological Chemistry 263.34 (1988): 18386-8396. [18] Christie, William W. "Fatty Acids: Natural Alicyclic Structures, Occurrence, and Biochemistry." The AOCS Lipid Library. 5 Apr. 2011. Web. 24 Apr. 2011. <http://lipidlibrary.aocs.org/lipids/fa_cycl/file.pdf>. [19] Ratledge, Colin, and John Stanford. The Biology of the Mycobacteria. London: Academic, 1982. Print. [20] Kubica, George P., and Lawrence G. Wayne. The Mycobacteria: a Sourcebook. New York: Dekker, 1984. Print.

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External links
Overview (http://www.rpi.edu/dept/bcbp/molbiochem/MBWeb/mb2/part1/fasynthesis.htm) at Rensselaer Polytechnic Institute Overview (http://web.indstate.edu/thcme/mwking/lipid-synthesis.html#synthesis) at Indiana State University

Lipogenesis
Lipogenesis is the process by which acetyl-CoA is converted to fats. The former is an intermediate stage in metabolism of simple sugars, such as glucose, a source of energy of living organisms. Through lipogenesis, the energy can be efficiently stored in the form of fats. Lipogenesis encompasses the processes of fatty acid synthesis and subsequent triglyceride synthesis (when fatty acids are esterified with glycerol to form fats).[1] The products are secreted from the liver in the form of very-low-density lipoproteins (VLDL).

Fatty acid synthesis

Fatty acids synthesis starts with acetyl-CoA and builds up by the addition of two carbon units. The synthesis occurs in the cytoplasm in contrast to the degradation (oxidation), which occurs in the mitochondria. Many of the enzymes for the fatty acid synthesis are organized into a multienzyme complex called fatty acid synthetase.[2]

Control and regulation

Insulin is an indicator of the blood sugar level of the body, as its concentration increases proportionally with blood sugar levels. Thus, a large insulin level is associated with the fed state. As one might expect, therefore, it increases the rate of storage pathways, such as lipogenesis. Insulin stimulates lipogenesis in two main ways: The enzymes pyruvate dehydrogenase (PDH), which forms acetyl-CoA, and acetyl-CoA carboxylase (ACC), which forms malonyl-CoA, are obvious control points. These are activated by insulin. So a high insulin level leads to an overall increase in the levels of malonyl-CoA, which is the substrate required for fatty acids synthesis. PDH dephosphorylation Pyruvate dehydrogenase dephosphorylation is increased with the release of insulin. The dephosphorylated form is more active. As insulin binds to cellular surface transmembrane receptors that intracellularly activate the adenylate cyclase enzyme that catalyze cAMP (cyclic AMP) production from ATP. The increased intracellular cAMP, acts as a second messenger, in response to the insulin binding. cAMP activates protein kinase enzyme that in turn activates phosporylase enzyme that phosphorylates and in doing so activates a number of different intracellular enzymes such as the pyruvate dehydrogenase that dehydrates pyruvate to form AcCoa. So, an extracellular hormone, insulin, can in multistep activation (cascade) activate an enzyme in the cellular matrix. This mechanism leads to the increased rate of catalysis of this enzyme, so increases the levels of acetyl-CoA. Increased levels of acetyl-CoA will increase the flux through not only the fat synthesis pathway but also the citric acid cycle.

Lipogenesis Acetyl-CoA carboxylase Insulin affects ACC in a similar way to PDH. It leads to its dephosphorylation which activates the enzyme. Glucagon has an antagonistic effect and increases phosphorylation, deactivation, thereby inhibiting ACC and slowing fat synthesis. Affecting ACC affects the rate of acetyl-CoA conversion to malonyl-CoA. Increased malonyl-CoA level pushes the equilibrium over to increase production of fatty acids through biosynthesis. Long chain fatty acids are negative allosteric regulators of ACC and so when the cell has sufficient long chain fatty acids, they will eventually inhibit ACC activity and stop fatty acid synthesis. AMP and ATP concentrations of the cell act as a measure of the ATP needs of a cell and as ATP levels get low it activates the ATP synthetase which in turn phosphorylates ACC. When ATP is depleted, there is a rise in 5'AMP. This rise activates AMP-activated protein kinase, which phosphorylates ACC, thereby inhibits fat synthesis. This is a useful way to ensure that glucose is not diverted down a storage pathway in times when energy levels are low. ACC is also activated by citrate. This means that, when there is abundant acetyl-CoA in the cell cytoplasm for fat synthesis, it proceeds at an appropriate rate. Note: Research now shows that glucose metabolism (exact metabolite to be determined), aside from insulin's influence on lipogenic enzyme genes, can induce the gene products for liver's pyruvate kinase, acetyl-CoA carboxylase, and fatty acid synthase. These genes are induced by the transcription factors ChREBP/Mlx via high blood glucose levels[3] and presently unknown signaling events. Insulin induction is due to SREBP-1c, which is also involved in cholesterol metabolism.

291

Fatty acid esterification

Experiments were conducted to study in vivo the over-all fatty acid specificity of the mechanisms involved in chylomicron cholesterol ester and triglyceride formation during fat absorption in the rat. Mixtures containing similar amounts of two, three, or four C14-labeled fatty acids (palmitic, stearic, oleic, and linoleic acids), but with varying ratios of unlabeled fatty acids, were given by gastric intubation to rats with cannulated thoracic ducts. The chyle or chylomicron lipid so obtained was chromatographed on silicic acid columns to separate cholesterol esters and glycerides (the latter being 98.2% triglycerides). After assaying each lipid class for total radioactivity, gas-liquid chromatography was employed to measure the total mass and the distribution of mass and of radioactivity in the individual fatty acid components of each lipid fraction. The specific radioactivity of each fatty acid in each fraction could then be calculated. The data provided quantitative information on the relative specificity of incorporation of each fatty acid into each chylomicron lipid class and on the relative extent to which each fatty acid in each lipid fraction was diluted with endogenous fatty acid. With the exception of a slight discrimination against stearic acid, the processes of fatty acid absorption and chylomicron triglyceride formation displayed no specificity for one fatty acid relative to another. In contrast, chylomicron cholesterol ester formation showed marked specificity for oleic acid, relative to the other three fatty acids. This specificity was not significantly altered by varying the composition of the test meal, by including cholesterol in the test meal, or by feeding the animal a high-cholesterol diet for several weeks preceding the study. Considerable dilution of the dietary fatty acids with endogenous fatty acids was observed. In one experiment, 43% of the chylomicron triglyceride fatty acids was of endogenous origin. Relatively more (54%) of the cholesterol ester fatty acids was of endogenous origin. About 100,000 metric tons of the natural fatty acids are consumed in the preparation of various fatty acid esters. The simple esters with lower chain alcohols (methyl-, ethyl-, n-propyl-, isopropyl- and butyl esters) are used as emollients in cosmetics and other personal care products and as lubricants. Esters of fatty acids with more complex alcohols, such as sorbitol, ethylene glycol, diethylene glycol and polyethylene glycol are consumed in foods, personal care, paper, water treatment, metal working fluids, rolling oils and synthetic lubricants.

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References
[1] Kersten S (April 2001). "Mechanisms of nutritional and hormonal regulation of lipogenesis". EMBO Rep. 2 (4): 2826. doi:10.1093/embo-reports/kve071. PMC1083868. PMID11306547. [2] Elmhurst College. "Lipogenesis" (http:/ / www. elmhurst. edu/ ~chm/ vchembook/ 623acetylCoAfate. html). . Retrieved 2007-12-22. [3] Work from Howard Towle, Catherine Postic, and K. Uyeda.

Acetyl-CoA carboxylase
Acetyl-CoA carboxylase
Identifiers EC number CAS number 6.4.1.2
[1] [2]

9023-93-2

Databases IntEnz BRENDA ExPASy KEGG MetaCyc PRIAM IntEnz view

[3] [4] [5]

BRENDA entry NiceZyme view KEGG entry

[6]

metabolic pathway profile

[8]

[7]

PDB structures RCSB PDB [9] PDBe [10] PDBsum [11] Gene Ontology AmiGO [12] / EGO [13] Search PMC articles
[14]

PubMed articles [15]

Acetyl-CoA carboxylase alpha

Identifiers Symbol Alt. symbols Entrez HUGO OMIM RefSeq UniProt ACACA ACAC, ACC1, ACCA 31 84
[16] [17] [18] [19]

601557

NM_198839 Q13085
[20]

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293
Other data EC number Locus 6.4.1.2
[21] [22]

Chr. 17 q21

Acetyl-CoA carboxylase beta

Identifiers Symbol Alt. symbols Entrez HUGO OMIM RefSeq UniProt ACACB ACC2, ACCB 32 85
[23] [24] [25] [26]

200350

NM_001093 O00763 Other data

[27]

EC number Locus

6.4.1.2

[21] [28]

Chr. 12 q24.1

Acetyl-CoA carboxylase (ACC) is a biotin-dependent enzyme that catalyzes the irreversible carboxylation of acetyl-CoA to produce malonyl-CoA through its two catalytic activities, biotin carboxylase (BC) and carboxyltransferase (CT). ACC is a multi-subunit enzyme in most prokaryotes and in the chloroplasts of most plants and algae, whereas it is a large, multi-domain enzyme in the endoplasmic reticulum of most eukaryotes. The most important function of ACC is to provide the malonyl-CoA substrate for the biosynthesis of fatty acids.[29] The activity of ACC can be controlled at the transcriptional level as well as by small molecule modulators and covalent modification. The human genome contains the genes for two different ACCs[30] ACACA[31] and ACACB.[32]

Structure
Prokaryotes and plants have multi-subunit ACCs composed of several polypeptides encoded by distinct genes. Biotin carboxylase (BC) activity, biotin carboxyl carrier protein (BCCP), and carboxyl transferase (CT) activity are each contained on a different subunit. The stoichiometry of these subunits in the ACC holoenzyme differs amongst organisms.[29] Humans and most eukaryotes have evolved an ACC with CT and BC catalytic domains and biotin carboxyl carrier domains on a single polypeptide. ACC functional regions, starting from the N-terminus to C-terminus are the biotin carboxylase (BC), biotin binding (BB), carboxyltransferase (CT), and ATP-binding (AB). AB lies within BC. Biotin is covalently attached through an amide bond to the long side chain of a lysine reside in BB. As BB is between BC and CT regions, biotin can be easily translocate to both of the active sites where it is required. In mammals where two isoforms of ACC are expressed, the main structural difference between these isoforms is the extended ACC2 N-terminus containing a mitochondria targeting sequence.[29]

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294

Crystallographic structures of E. Coli Acetyl CoA Carboxylase

Cartoon diagram of Biotin Carboxylase of E.Coli Acetyl CoA Carboxylase

Acetyl-CoA carboxylase

295

Biotin Carboxyl Carrier Protein of E. Coli Acetyl CoA Carboxylase

Carboxyltransferase Subunit of E. Coli Acetyl CoA Carboxylase

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296

Mechanism
The overall reaction of ACAC(A,B) proceeds by a two-step mechanism.[33] The first reaction is carried out by BC and involves the ATP-dependent carboxylation of biotin with bicarbonate serving as the source of CO2. The carboxyl group is transferred from biotin to acetyl CoA to form malonyl CoA in the second reaction, which is catalyzed by CT.

The reaction mechanism of ACAC(A,B).The color scheme is as follows: enzyme, coenzymes, substrate names, metal ions, phosphate, and carbonate

In the context of the active site, the reaction proceeds with extensive interaction of the residues Glu296 and positively charged Arg338 and Arg292 with the substrates.[34] Two Mg2+ are coordinated by the phosphate groups on the ATP, and are required for ATP binding to the enzyme. Bicarbonate is deprotonated by Glu296, although in solution, this proton transfer is unlikely as the pKa of bicarbonate is 10.3. The enzyme apparently manipulates pKas to facilitate the deprotonation of bicarbonate. The pKa of bicarbonate is decreased by its interaction with positively charged side chains of Arg338 and Arg292. Furthermore, Glu296 interacts with the side chain of Glu211, an interaction that has been shown to cause an increase in the apparent pKa. Following deprotonation of bicarbonate, the oxygen of the bicarbonate acts as a nucleophile and attacks the gamma phosphate on ATP. The carboxyphosphate intermediate quickly decomposes to CO2 and PO43-. The PO43- deprotonates biotin, creating an enolate, stabilized by Arg338, that subsequently attacks CO₂ resulting in the production of carboxybiotin.[34] The carboxybiotin translocates to the carboxytransferase (CT) active site, where the carboxyl group is transferred to acetyl-CoA. In contrast to the BC domain, little is known about the reaction mechanism of CT. A proposed mechanism is the release of carbon dioxide from biotin, which subsequently abstracts a proton from the methyl group from acetyl CoA carboxylase. The resulting enolate attacks CO2 to form malonyl CoA. In a competing mechanism, proton abstraction is concerted with the attack of acetyl CoA.

Function
The function of ACAC is to regulate the metabolism of fatty acids. When the enzyme is active, the product, malonyl-CoA is produced which is a building block for new fatty acids and can inhibit the transfer of the fatty acyl group from acyl CoA to carnitine with carnitine acyltransferase, which inhibits the beta-oxidation of fatty acids in the mitochondria. In mammals, two main isoforms of ACC are expressed, ACC1 and ACC2, which differ in both tissue distribution and function. ACC1 is found in the cytoplasm of all cells but is enriched in lipogenic tissue, such as adipose tissue and lactating mammary glands, where fatty acid synthesis is important.[35] In oxidative tissues, such as the skeletal muscle and the heart, the ratio of ACC2 expressed is higher. ACC1 and ACC2 are both highly expressed in the liver where both fatty acid oxidation and synthesis is important.[36] The differences in tissue distribution indicate that

Acetyl-CoA carboxylase ACC1 maintains regulation of fatty acid synthesis whereas ACC2 mainly regulates fatty acid oxidation.

297

Regulation
The regulation of mammalian ACC is complex, in order to control two distinct pools of malonyl CoA that direct either the inhibition of beta oxidation or the activation of lipid biosynthesis.

Mammalian ACC1 and ACC2 are regulated transcriptionally by multiple promoters which mediate ACC abundance in response to the cells nutritional status. Activation of gene expression through different promoters results in alternative splicing; however, the physiological significance of specific ACC isozymes remains unclear.[36] The sensitivity to nutritional status results from the control of these promoters by transcription factors such as SREBP1c, controlled by insulin at the transcriptional level, and ChREBP, which increases in expression with high carbohydrates diets.[37] [38] Through a feedforward loop, citrate allosterically activates ACC.[39] Citrate may increase ACC polymerization to increases enzymatic activity; however, it is unclear if polymerization is citrate's main mechanism of increasing ACC activity or if polymerization is an artifact of in vitro experiments. Other allosteric activators include glutamate and other dicarboxylic acids.[40] Long and short chain fatty acyl CoAs are negative feedback inhibitors of ACC.[41] Phosphorylation can result when the hormones glucagon or epinephrine bind to cell surface receptors, but the main cause of phosphorylation is due to a rise in AMP levels when the energy status of the cell is low, leading to the activation of the AMP-activated protein kinase (AMPK). AMPK is the main kinase regulator of ACC, able to phosphorylate a number of serine residues on both isoforms of ACC.[42] On ACC1, AMPK phosphorylates Ser79, Ser1200, and Ser1215. On ACC2, AMPK phosphorylates Ser218.[43] Protein kinase A also has the ability to phosphorylate ACC, with a much greater ability to phosphorylate ACC2 than ACC1. However, the physiological significance of protein kinase A in the regulation of ACC is currently unknown. Researchers hypothesize there are other ACC kinases important to its regulation as there are many other possible phosphorylation sites on ACC.[44] When insulin binds to its receptors on the cellular membrane, it activates a phosphatase to dephosphorylate the enzyme; thereby removing the inhibitory effect.

Control of Acetyl CoA Carboxylase. The AMP regulated kinase triggers the phosphorylation of the enzyme (thus inactivating it) and the phosphatase enzyme removes the phosphate group.

Clinical implications
At the juncture of lipid synthesis and oxidation pathways, ACC presents many clinical possibilities for the production of novel antibiotics and the development of new therapies for diabetes, obesity, and other manifestations of metabolic syndrome.[45] Researchers aim to take advantage of structural differences between bacterial and human ACCs to create antibiotics specific to the bacterial ACC, in efforts to minimize side effects to patients. Promising results for the usefulness of an ACC inhibitor include the finding that ACC2 -/- mice (mice with no expression of ACC2) have continuous fatty acid oxidation, reduced body fat mass, and reduced body weight despite an increase in food consumption. ACC2 -/- mice are also protected from diabetes.[46] It should be noted that mutant mice lacking ACC1 are embryonically lethal. However, it is unknown whether drugs targeting ACCs in humans must be specific for ACC2.[47]

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References
[1] [2] [3] [4] [5] [6] [7] [8] [9] http:/ / www. chem. qmul. ac. uk/ iubmb/ enzyme/ EC6/ 4/ 1/ 2. html http:/ / toolserver. org/ ~magnus/ cas. php?language=en& cas=9023-93-2& title= http:/ / www. ebi. ac. uk/ intenz/ query?cmd=SearchEC& ec=6. 4. 1. 2 http:/ / www. brenda-enzymes. org/ php/ result_flat. php4?ecno=6. 4. 1. 2 http:/ / www. expasy. org/ enzyme/ 6. 4. 1. 2 http:/ / www. genome. ad. jp/ dbget-bin/ www_bget?enzyme+ 6. 4. 1. 2 http:/ / biocyc. org/ META/ substring-search?type=NIL& object=6. 4. 1. 2 http:/ / priam. prabi. fr/ cgi-bin/ PRIAM_profiles_CurrentRelease. pl?EC=6. 4. 1. 2 http:/ / www. rcsb. org/ pdb/ search/ smartSubquery. do?smartSearchSubtype=EnzymeClassificationQuery& Enzyme_Classification=6. 4. 1. 2 [10] http:/ / www. ebi. ac. uk/ pdbe-srv/ view/ search?EC_number=6. 4. 1. 2& id_type=EC_number& id_value=6. 4. 1. 2& search_type=advanced [11] http:/ / www. ebi. ac. uk/ thornton-srv/ databases/ cgi-bin/ enzymes/ GetPage. pl?ec_number=6. 4. 1. 2 [12] http:/ / amigo. geneontology. org/ cgi-bin/ amigo/ go. cgi?query=GO:0003989& view=details [13] http:/ / www. ebi. ac. uk/ ego/ DisplayGoTerm?id=GO:0003989& format=normal [14] http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=pubmed& term=6. 4. 1. 2%5BEC/ RN%20Number%5D%20AND%20pubmed%20pmc%20local%5Bsb%5D [15] http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=pubmed& term=6. 4. 1. 2%5BEC/ RN%20Number%5D [16] http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=gene& amp;cmd=retrieve& amp;dopt=default& amp;list_uids=31& rn=1 [17] http:/ / www. genenames. org/ data/ hgnc_data. php?hgnc_id=84 [18] http:/ / www. ncbi. nlm. nih. gov/ omim/ 601557 [19] http:/ / genome. ucsc. edu/ cgi-bin/ hgTracks?Submit=Submit& position=NM_198839& rn=1 [20] http:/ / www. uniprot. org/ uniprot/ Q13085 [21] http:/ / www. genome. jp/ dbget-bin/ www_bget?enzyme+ 6. 4. 1. 2 [22] http:/ / omim. org/ search?index=geneMap& search=17q21 [23] http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?db=gene& amp;cmd=retrieve& amp;dopt=default& amp;list_uids=32& rn=1 [24] http:/ / www. genenames. org/ data/ hgnc_data. php?hgnc_id=85 [25] http:/ / www. ncbi. nlm. nih. gov/ omim/ 200350 [26] http:/ / genome. ucsc. edu/ cgi-bin/ hgTracks?Submit=Submit& position=NM_001093& rn=1 [27] http:/ / www. uniprot. org/ uniprot/ O00763 [28] http:/ / omim. org/ search?index=geneMap& search=12q24. 1 [29] Tong L (August 2005). "Acetyl-coenzyme A carboxylase: crucial metabolic enzyme and attractive target for drug discovery". Cell. Mol. Life Sci. 62 (16): 1784803. doi:10.1007/s00018-005-5121-4. PMID15968460. [30] Brownsey RW, Zhande R, Boone AN (November 1997). "Isoforms of acetyl-CoA carboxylase: structures, regulatory properties and metabolic functions". Biochem. Soc. Trans. 25 (4): 12328. PMID9449982. [31] Abu-Elheiga L, Jayakumar A, Baldini A, Chirala SS, Wakil SJ (April 1995). "Human acetyl-CoA carboxylase: characterization, molecular cloning, and evidence for two isoforms". Proc. Natl. Acad. Sci. U.S.A. 92 (9): 40115. doi:10.1073/pnas.92.9.4011. PMC42092. PMID7732023. [32] Widmer J, Fassihi KS, Schlichter SC, Wheeler KS, Crute BE, King N, Nutile-McMenemy N, Noll WW, Daniel S, Ha J, Kim KH, Witters LA (June 1996). "Identification of a second human acetyl-CoA carboxylase gene" (http:/ / www. biochemj. org/ bj/ 316/ 0915/ bj3160915. htm). Biochem. J.. 316 ( Pt 3): 91522. PMC1217437. PMID8670171. . [33] Lee CK, Cheong HK, Ryu KS, Lee JI, Lee W, Jeon YH, Cheong C (August 2008). "Biotinoyl domain of human acetyl-CoA carboxylase: Structural insights into the carboxyl transfer mechanism". Proteins 72 (2): 61324. doi:10.1002/prot.21952. PMID18247344. [34] Chou CY, Yu LP, Tong L (April 2009). "Crystal structure of biotin carboxylase in complex with substrates and implications for its catalytic mechanism". J. Biol. Chem. 284 (17): 116907. doi:10.1074/jbc.M805783200. PMC2670172. PMID19213731. [35] Kim TS, Leahy P, Freake HC (August 1996). "Promoter usage determines tissue specific responsiveness of the rat acetyl-CoA carboxylase gene". Biochem. Biophys. Res. Commun. 225 (2): 64753. doi:10.1006/bbrc.1996.1224. PMID8753813. [36] Barber MC, Price NT, Travers MT (March 2005). "Structure and regulation of acetyl-CoA carboxylase genes of metazoa". Biochim. Biophys. Acta 1733 (1): 128. doi:10.1016/j.bbalip.2004.12.001. PMID15749055. [37] Field F. J., Born E., Murthy S. and Mathur S. N. (December 2002). "Polyunsaturated fatty acids decrease the expression of sterol regulatory element binding protein-1 in CaCo-2 cells: effect on fatty acid synthesis and triacylglycerol transport.". Biochem. J. 386 (Pt 3): 85564. doi:10.1042/BJ20020731. PMC1223029. PMID12213084. [38] Ishii S, Iizuka K, Miller BC, Uyeda K (October 2004). "Carbohydrate response element binding protein directly promotes lipogenic enzyme gene transcription". Proc Natl Acad Sci USA 101 (44): 15597602. doi:10.1073/pnas.0405238101. PMC524841. PMID15496471. [39] Martin DB, Vagelos PR (June 1962). "The Mechanism of Tricarboxylic Acid Cycle Regulation of Fatty Acid Synthesis". J Biol Chem 237: 178792. PMID14470343.

Acetyl-CoA carboxylase
[40] Boone AN, Chan A, Kulpa JE, Brownsey RW (April 2000). "Bimodal Activation of Acetyl-CoA Carboxylase by Glutamate". J Biol Chem 275 (15): 1081925. doi:10.1074/jbc.275.15.10819. PMID10753875. [41] Faergeman NJ, Knudsen J (April 1997). "Role of long chain fatty acyl-CoA esters in the regulation of metabolism and in cell signalling". Biochem J. 323 (Pt 1): 112. PMC1218279. PMID9173866. [42] Park SH, Gammon SR, Knippers JD, Paulsen SR, Rubink DS, Winder WW (June 2002). "Phosphorylation-activity relationships of AMPK and acetyl-CoA carboxylase in muscle". J. Appl. Physiol. 92 (6): 247582. doi:10.1152/japplphysiol.00071.2002. PMID12015362. [43] Hardie DG (February 1992). "Regulation of fatty acid and cholesterol metabolism by the AMP-activated protein kinase". Biochim. Biophys. Acta 1123 (3): 2318. PMID1536860. [44] Brownsey RW, Boone AN, Elliott JE, Kulpa JE, Lee WM (April 2006). "Regulation of acetyl-CoA carboxylase". Biochem. Soc. Trans. 34 (Pt 2): 2237. doi:10.1042/BST20060223. PMID16545081. [45] Corbett JW, Harwood JH (November 2007). "Inhibitors of mammalian acetyl-CoA carboxylase". Recent Patents Cardiovasc Drug Discov 2 (3): 16280. doi:10.2174/157489007782418928. PMID18221116. [46] L Abu-Elheiga, M M Matzuk, K A Abo-Hashema, S J Wakil (March 2001). "Continuous Fatty Acid Oxidation and Reduced Fat Storage in Mice Lacking Acetyl-CoA Carboxylase 2". Science 291 (5513): 26136. doi:10.1126/science.1056843. PMID11283375. [47] Lutfi Abu-Elheiga, Martin M Matzuk, Parichher Kordari, WonKeun Oh, Tattym Shaikenov, Ziwei Gu, Salih J Wakil (August 2005). "Mutant Mice Lacking Acetyl CoA Carboxylase are Embryonically Lethal". Proc Natl Acad Sci 102 (34): 12116. doi:10.1073/pnas.0505714102. PMC1189351. PMID16103361.

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Further reading
Voet, Donald; Voet, Judith G. (2004). Biochemistry (3rd ed.). Wiley. ISBN0-471-19350-x. edited by (2000). Buchanan, Bob B.; Gruissem, Wilhelm; Jones, Russell L.. eds. Biochemistry and molecular biology of plants. American Society of Plant Physiologists. ISBN0-943088-37-2. Levert K, Waldrop G, Stephens J (2002). "A biotin analog inhibits acetyl-CoA carboxylase activity and adipogenesis". J. Biol. Chem. 277 (19): 1634750. doi:10.1074/jbc.C200113200. PMID11907024.

Fatty acid degradation

Fatty acid degradation is the process in which fatty acids are broken down into their metabolites, in the end generating acetyl-CoA, the entry molecule for the citric acid cycle, the main energy supply of animals. It includes three major steps: Lipolysis of and release from adipose tissue Activation and transport into mitochondria -oxidation

Lipolysis and release

Initially in the process of degradation, fatty acids are stored in fat cells (adipocytes). The breakdown of this fat is known as lipolysis. The products of lipolysis, free fatty acids, are released into the bloodstream and circulate throughout the body.

Activation and transport into mitochondria

Fatty acids must be activated before they can be carried into the mitochondria, where fatty acid oxidation occurs. This process occurs in two steps catalyzed by the enzyme fatty acyl-CoA synthetase.

Fatty acid degradation

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Formation of an activated thioester bond

The enzyme first catalyzes nucleophilic attack on the -phosphate of ATP to form pyrophosphate and an acyl chain linked to AMP. The next step is formation of an activated thioester bond between the fatty acyl chain and Coenzyme A.

The formula for the above is: RCOO- + CoA + ATP + H2O RCO-CoA + AMP + PPi + 2H+ This two-step reaction is freely reversible and its equilibrium lies near 1. To drive the reaction forward, the reaction is coupled to a strongly exergonic hydrolysis reaction: the enzyme inorganic pyrophosphatase cleaves the pyrophosphate liberated from ATP to two phosphate ions. Thus the net reaction becomes: RCOO- + CoA + ATP + H2O RCO-CoA + AMP + 2Pi + 2H+

Transport into the mitochondrial matrix

The inner mitochondrial membrane is impermeable to fatty acids and a specialized carnitine carrier system operates to transport activated fatty acids from cytosol to mitochondria. Once activated, the acyl CoA is transported into the mitochondrial matrix. This occurs via a series of similar steps: 1. Acyl CoA is conjugated to carnitine by carnitine acyltransferase (palmitoyltransferase) I located on the outer mitochondrial membrane 2. Acyl carnitine is shuttled inside by a translocase 3. Acyl carnitine (such as Palmitoylcarnitine) is converted to acyl CoA by carnitine acyltransferase (palmitoyltransferase) II located on the inner mitochondrial membrane. The liberated carnitine returns to the cytosol. It is important to note that carnitine acyltransferase I undergoes allosteric inhibition as a result of malonyl-CoA, an intermediate in fatty acid biosynthesis, in order to prevent futile cycling between beta-oxidation and fatty acid synthesis.

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-oxidation
Once inside the mitochondria, the -oxidation of fatty acids occurs via four recurring steps: 1. 2. 3. 4. 5. Oxidation by FAD, Hydration, Oxidation by NAD+, Thiolysis, The final product is acetyl-CoA, the entry molecule for the citric acid cycle.

Beta oxidation
Beta oxidation is the process by which fatty acids, in the form of Acyl-CoA molecules, are broken down in mitochondria and/or in peroxisomes to generate Acetyl-CoA, the entry molecule for the Citric Acid cycle. The beta oxidation of fatty acids involve three stages: 1. Activation of fatty acids in the cytosol 2. Transport of fatty acids into mitochondria (carnitine shuttle) 3. Beta oxidation proper in the mitochondrial matrix
Schematic demonstrating mitochondrial fatty acid beta-oxidation and effects of long-chain 3-hydroxyacyl-coenzyme A dehydrogenase deficiency, LCHAD deficiency

Fatty acids are oxidized by most of the tissues in the body. However, the brain can hardly utilize fatty acids for energy requirements, while erythrocytes and adrenal medulla cannot use them at all.

Activation of fatty acids

Free fatty acids can penetrate the plasma membrane due to their poor water solubility and high fat solubility. Once in the cytosol, activation of the fatty acid is catalyzed by long fatty acyl CoA synthetase. A fatty acid reacts with ATP to give a fatty acyl adenylate, plus inorganic pyrophosphate, which then reacts with free coenzyme A to give a fatty acyl-CoA ester plus AMP. The fatty acyl-CoA is then reacted with carnitine to form acylcarnitine, which is transported across the inner mitochondrial membrane by a monosodium glutamate.

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Four recurring steps

Once inside the mitochondria, each cycle of -oxidation, liberating a two carbon unit-acetyl CoA, occurs in a sequence of four reactions:
Description Dehydrogenation by FAD: The first step is the oxidation of the fatty acid by Acyl-CoA-Dehydrogenase. The enzyme catalyzes the formation of a double bond between the C-2 and C-3. Hydration: The next step is the hydration of the bond between C-2 and C-3. The reaction is stereospecific, forming only the L isomer. Diagram Enzyme acyl CoA dehydrogenase End product trans-2-enoyl-CoA

enoyl CoA hydratase

L--hydroxyacyl CoA

Oxidation by NAD+: The third step is the oxidation of L--hydroxyacyl CoA by NAD+. This converts the hydroxyl group into a keto group.

L--hydroxyacyl CoA dehydrogenase

-ketoacyl CoA

Thiolysis: The final step is the cleavage of -ketoacyl CoA by the thiol group of another molecule of CoA. The thiol is inserted between C-2 and C-3.

-ketothiolase

An acetyl CoA molecule, and an acyl CoA molecule that is two carbons shorter

This process continues until the entire chain is cleaved into acetyl CoA units. The final cycle produces two separate acetyl CoAs, instead of one acyl CoA and one acetyl CoA. For every cycle, the Acyl CoA unit is shortened by two carbon atoms. Concomitantly, one molecule of FADH2, NADH and acetyl CoA are formed.

-Oxidation of unsaturated fatty acids

-Oxidation of unsaturated fatty acids poses a problem since the location of a cis bond can prevent the formation of a trans-2 bond. These situations are handled by an additional two enzymes. Whatever the conformation of the hydrocarbon chain, -oxidation occurs normally until the acyl CoA (because of the presence of a double bond) is not an appropriate substrate for acyl CoA dehydrogenase, or enoyl CoA hydratase: If the acyl CoA contains a cis-3 bond, then cis-3-Enoyl CoA isomerase will convert the bond to a trans-2 bond, which is a regular substrate. If the acyl CoA contains a cis-4 double bond, then its dehydrogenation yields a 2,4-dienoyl intermediate, which is not a substrate for enoyl CoA hydratase. However, the enzyme 2,4 Dienoyl CoA reductase reduces the intermediate, using NADPH, into trans-3-enoyl CoA. As in the above case, this compound is converted into a suitable intermediate by 3,2-Enoyl CoA isomerase. To summarize: Odd-numbered double bonds are handled by the isomerase. Even-numbered double bonds by the reductase (which creates an odd-numbered double bond)

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-Oxidation of odd-numbered chains

In general, fatty acids with an odd number of carbon are found in the lipids of plants and some marine organisms. Many ruminant animals form large amount of 3-carbon propionate during fermentation of carbohydrate in rumen.[1] Chains with an odd-number of carbons are oxidized in the same manner as even-numbered chains, but the final products are propionyl-CoA and acetyl-CoA. Propionyl-CoA is first carboxylated using a bicarbonate ion into D-stereoisomer of methylmalonyl-CoA, in a reaction that involves a biotin co-factor, ATP, and the enzyme propionyl-CoA carboxylase. The bicarbonate ion's carbon is added to the middle carbon of propionyl-CoA, forming a D-methylmalonyl-CoA. However, the D conformation is enzymatically converted into the L conformation by methylmalonyl-CoA epimerase, then it undergoes intramolecular rearrangement, which is catalyzed by methylmalonyl-CoA mutase (requiring B12 as a coenzyme) to form succinyl-CoA. The succinyl-CoA formed can then enter the citric acid cycle. Because it cannot be completely metabolized in the citric acid cycle, the products of its partial reaction must be removed in a process called cataplerosis. This allows regeneration of the citric acid cycle intermediates, possibly an important process in certain metabolic diseases.

Oxidation in peroxisomes
Fatty acid oxidation also occurs in peroxisomes, when the fatty acid chains are too long to be handled by the mitochondria. However, the oxidation ceases at octanyl CoA. It is believed that very long chain (greater than C-22) fatty acids undergo initial oxidation in peroxisomes which is followed by mitochondrial oxidation. One significant difference is that oxidation in peroxisomes is not coupled to ATP synthesis. Instead, the high-potential electrons are transferred to O2, which yields H2O2. The enzyme catalase, found exclusively in peroxisomes, converts the hydrogen peroxide into water and oxygen. Peroxisomal -oxidation also requires enzymes specific to the peroxisome and to very long fatty acids. There are three key differences between the enzymes used for mitochondrial and peroxisomal -oxidation: 1. -oxidation in the peroxisome requires the use of a peroxisomal carnitine acyltransferase (instead of carnitine acyltransferase I and II used by the mitochondria) for transport of the activated acyl group into the peroxisome. 2. The first oxidation step in the peroxisome is catalyzed by the enzyme acyl CoA oxidase. 3. The -ketothiolase used in peroxisomal -oxidation has an altered substrate specificity, different from the mitochondrial -ketothiolase. Peroxisomal oxidation is induced by high-fat diet and administration of hypolipidemic drugs like clofibrate.

Energy yield
The ATP yield for every oxidation cycle is 14 ATP (according to the P/O ratio), broken down as follows:

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Source 1 FADH2 1 NADH

ATP

Total

x 1.5 ATP = 1.5 ATP (some sources say 2 ATP) x 2.5 ATP = 2.5 ATP (some sources say 3 ATP) = 10 ATP (some sources say 12 ATP) = 14 ATP

1 acetyl CoA x 10 ATP TOTAL

For an even-numbered saturated fat (C2n), n - 1 oxidations are necessary, and the final process yields an additional acetyl CoA. In addition, two equivalents of ATP are lost during the activation of the fatty acid. Therefore, the total ATP yield can be stated as: (n - 1) * 14 + 10 - 2 = total ATP For instance, the ATP yield of palmitate (C16, n = 8) is: (8 - 1) * 14 + 10 - 2 = 106 ATP Represented in table form:
Source 7 FADH2 7 NADH ATP Total

x 1.5 ATP = 10.5 ATP x 2.5 ATP = 17.5 ATP = 80 ATP = -2 ATP = 106 ATP

8 acetyl CoA x 10 ATP Activation NET

For sources that use the larger ATP production numbers described above, the total would be 129 ATP ={(8-1)*17+12-2} equivalents per palmitate. Beta-oxidation of unsaturated fatty acids changes the ATP yield due to the requirement of two possible additional enzymes.

External links
The chemical logic behind fatty acid metabolism [2] at ufp.pt JEREMY M. BERG,JOHN L. TYMOCZKO and LUBERT STRYER Biochemistry, 2002 [3] Animations [4] at brookscole.com

References
[1] Nelson, D. L. & Cox, M. M. (2005). Lehninger Principles of Biochemistry, 4th Edition. New York: W. H. Freeman and Company, pp. 648-649. ISBN 0-7167-4339-6. [2] http:/ / www2. ufp. pt/ ~pedros/ bq/ fatty. htm [3] http:/ / www. ncbi. nlm. nih. gov/ entrez/ query. fcgi?cmd=Search& db=books& doptcmdl=GenBookHL& term=oxidation+ yield+ AND+ stryer%5Bbook%5D+ AND+ 216592%5Buid%5D& rid=stryer. section. 3050#3060 [4] http:/ / www. brookscole. com/ chemistry_d/ templates/ student_resources/ shared_resources/ animations/ carnitine/ carnitine1. html

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Nitrogen metabolism
Nitrogen fixation
Nitrogen fixation is the natural process, either biological or abiotic, by which nitrogen (N2) in the atmosphere is converted into ammonia (NH3).[1] This process is essential for life because fixed nitrogen is required to biosynthesize the basic building blocks of life, e.g., nucleotides for DNA and RNA and amino acids for proteins. Nitrogen fixation also refers to other biological conversions of nitrogen, such as its conversion to nitrogen dioxide. Microorganisms that fix nitrogen are bacteria called diazotrophs. Some higher plants, and some animals (termites), have formed associations (symbioses) with diazotrophs. Nitrogen fixation also occurs as a result of non-biological processes. These include lightning, industrially through the Haber-Bosch Process, and combustion.[2] Biological nitrogen fixation was discovered by the German agronomist Hermann Hellriegel and Dutch microbiologist Martinus Beijerinck.

Biological nitrogen fixation

Biological nitrogen fixation (BNF) occurs when atmospheric nitrogen is converted to ammonia by an enzyme called nitrogenase.[1] The reaction for BNF is: N2 + 8 H+ + 8 e 2 NH3 + H2 The process is coupled to the hydrolysis of 16 equivalents of ATP and is accompanied by the co-formation of one molecule of H2. In free-living diazotrophs, the nitrogenase-generated ammonium is assimilated into glutamate through the glutamine synthetase/glutamate synthase pathway. Enzymes responsible for nitrogenase Schematic representation of the nitrogen cycle. Abiotic nitrogen fixation has been action are very susceptible to omitted. destruction by oxygen. (In fact, many bacteria cease production of the enzyme in the presence of oxygen).[1] Many nitrogen-fixing organisms exist only in anaerobic conditions, respiring to draw down oxygen levels, or binding the oxygen with a protein such as Leghemoglobin.[1]

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Microorganisms that fix nitrogen (diazotrophs)

Cyanobacteria Azotobacteraceae Rhizobia Frankia

Nitrogen fixation by rhizobia and frankia Rhizobia are Gram-negative soil bacteria with the ability to establish a N2-fixing symbiosis on legume roots and on the stems of some aquatic legumes. During this interaction bacteroids, as rhizobia are called in the symbiotic state, are contained in intracellular compartments within a specialized organ, the nodule, where they fix N2. Similarly, Frankia, Gram-positive soil bacteria induce the formation of nitrogen-fixing nodules in actinorhizal plants.[3] Nitrogen fixation by cyanobacteria Cyanobacteria inhabit nearly all illuminated environments on Earth and play key roles in the carbon and nitrogen cycle of the biosphere. In general, cyanobacteria are able to utilize a variety of inorganic and organic sources of combined nitrogen, like nitrate, nitrite, ammonium, urea, or some amino acids. Several cyanobacterial strains are also capable of diazotrophic growth. Genome sequencing has provided a large amount of information on the genetic basis of nitrogen metabolism and its control in different cyanobacteria. Comparative genomics, together with functional studies, has led to a significant advance in this field over the past years. 2-Oxoglutarate has turned out to be the central signalling molecule reflecting the carbon/nitrogen balance of cyanobacteria. Central players of nitrogen control are the global transcriptional factor NtcA, which controls the expression of many genes involved in nitrogen metabolism, as well as the PII signalling protein, which fine-tunes cellular activities in response to changing C/N conditions. These two proteins are sensors of the cellular 2-oxoglutarate level and have been conserved in all cyanobacteria. In contrast, the adaptation to nitrogen starvation involves heterogeneous responses in different strains.[4] Nitrogen fixation by cyanobacteria in coral reefs can fix twice the amount of nitrogen than on landaround 1.8kg of nitrogen is fixed per hectare per day.

Root nodule symbioses

Legume family Plants that contribute to nitrogen fixation include the legume family Fabaceae with taxa such as clovers, soybeans, alfalfa, lupines, peanuts, and rooibos. They contain symbiotic bacteria called Rhizobia within nodules in their root systems, producing nitrogen compounds that help the plant to grow and compete with other plants. When the plant dies, the fixed nitrogen is released, making it available to other plants and this helps to fertilize the soil[1] [5] The great majority of legumes have this association, but a few genera (e.g., Styphnolobium) do not. In many traditional and organic farming practices, fields are rotated through various types of crops, which usually includes one consisting mainly or entirely of clover or buckwheat (family Polygonaceae), which are often referred to as "green manure." Inga alley farming relies on the leguminous genus, Inga a small tropical, tough-leaved, nitrogen-fixing tree.[6]

Nitrogen fixation Non-leguminous Although by far the majority plants able to form nitrogen-fixing root nodules are in the legume family Fabaceae, there are a few exceptions: Parasponia, a tropical Celtidaceae also able to interact with rhizobia and form nitrogen-fixing nodules[7] Actinorhizal plants such as alder and bayberry, that can also forms nitrogen-fixing nodules, thanks to a symbiotic association with Frankia bacteria. These plants belong to 25 genera[8] distributed among 8 plant families. The ability to fix nitrogen is far from universally present in these families. For instance, of 122 genus in the Rosaceae, only 4 genera are capable of fixing nitrogen. All these families belong to the orders Cucurbitales, Fagales, and Rosales, which together with the Fabales form a clade of eurosids. In this clade, Fabales were the first lineage to branch off; thus, the ability to fix nitrogen may be plesiomorphic and subsequently lost in most descendants of the original nitrogen-fixing plant; however, it may be that the basic genetic and physiological requirements were present in an incipient state in the last common ancestors of all these plants, but only evolved to full function in some of them:

307

A sectioned alder tree root nodule.

A whole alder tree root nodule.

Family: Genera Betulaceae: Alnus (alders) Cannabaceae: Trema Casuarinaceae: Allocasuarina Casuarina Ceuthostoma Gymnostoma

...... Coriariaceae: Coriaria Datiscaceae: Datisca Elaeagnaceae: Elaeagnus (silverberries) Hippophae (sea-buckthorns) Shepherdia (buffaloberries)

...... Myricaceae: Comptonia (sweetfern) Morella Myrica (bayberries)

...... Rhamnaceae: Ceanothus Colletia Discaria Kentrothamnus Retanilla Talguenea Trevoa

...... Rosaceae: Cercocarpus (mountain mahoganies) Chamaebatia (mountain miseries) Dryas Purshia/Cowania (bitterbrushes/cliffroses)

There are also several nitrogen-fixing symbiotic associations that involve cyanobacteria (such as Nostoc): Some lichens such as Lobaria and Peltigera Mosquito fern (Azolla species) Cycads Gunnera

Nitrogen fixation

308

Chemical nitrogen fixation

Haber process
Nitrogen can also be artificially fixed as ammonia for use in fertilizers, explosives, or in other products. The most common method is the Haber process. Artificial fertilizer production is now the largest source of human-produced fixed nitrogen in the Earth's ecosystem.[9] The Haber process requires high pressures (around 200 atm) and high temperatures (at least 400 C), routine conditions for industrial catalysis. This highly efficient process uses natural gas as a hydrogen source and air as a nitrogen source.

Dinitrogen complexes
Much research has been conducted on the discovery of catalysts for nitrogen fixation, often with the goal of reducing the energy required for this conversion. However, such research has thus far failed to even approach the efficiency and ease of the Haber process. Many compounds react with atmospheric nitrogen under ambient conditions. For example, lithium metal converts to lithium nitride under an atmosphere of nitrogen. Treatment of the resulting nitride gives ammonia. The first dinitrogen complex was reported in 1965 based on ammonia coordinated to ruthenium ([Ru(NH3)5(N2)]2+).[10] Research in chemical fixation from then on focused on transition metal complexes. Since then, a large number of transition metal compounds that contain dinitrogen as a ligand have been discovered. The dinitrogen ligand can either be bound to a single metal or bridge two (or more) metals. The coordination chemistry of dinitrogen is complex and currently under intense investigation. This research may lead to new ways of using dinitrogen in synthesis and on an industrial scale.

Ambient nitrogen reduction

Catalytic chemical nitrogen fixation at temperatures considerably lower than the Haber process is an ongoing scientific endeavor. Nitrogen was successfully converted to ammonia and hydrazine by Alexander E. Shilov in 1970.[11] [12] The first example of homolytic cleavage of dinitrogen under mild conditions was published in 1995. Two equivalents of a molybdenum complex reacted with one equivalent of dinitrogen, creating a triple bonded MoN complex.[13] Since then, this triple bounded complex has been used to make nitriles.[14] The first catalytic system converting nitrogen to ammonia at room temperature and pressure was discovered in 2003 and is based on another molybdenum compound, a proton source, and a strong reducing agent.[15] [16] [17] [18] However, this catalytic reduction fixates only a few nitrogen molecules.

Nitrogen fixation

309

In 2011 Arashiba et al. reported another system with a catalyst again based on molybdenum but with a diphosphorus pincer ligand.[19]

References
[1] [2] [3] [4] Postgate, J (1998). Nitrogen Fixation, 3rd Edition. Cambridge University Press, Cambridge UK. http:/ / helios. bto. ed. ac. uk/ bto/ microbes/ nitrogen. htm Moir, JWB (editor) (2011). Nitrogen cycling in bacteria: Molecular analysis. Caister Academic Press. ISBN978-1-904455-86-8. Herrero A and Flores E (editor). (2008). The Cyanobacteria: Molecular Biology, Genomics and Evolution (http:/ / www. horizonpress. com/ cyan) (1st ed.). Caister Academic Press. ISBN978-1-904455-15-8. . . [5] Smil, V (2000). Cycles of Life. Scientific American Library. [6] Elkan, Daniel. Slash-and-burn farming has become a major threat to the world's rainforest The Guardian 21 April 2004 [7] Op den Camp, Rik; et al.. "LysM-Type Mycorrhizal Receptor Recruited for Rhizobium Symbiosis in Nonlegume Parasponia". Science 331 (6019): 909912. doi:10.1126/science.1198181. [8] Dawson, J. O. (2008). "Ecology of actinorhizal plants". Nitrogen-fixing Actinorhizal Symbioses. 6. Springer. pp.199234. doi:10.1007/978-1-4020-3547-0_8. [9] http:/ / www. epa. gov/ watertrain/ nitroabstr. html US Enivronmental Protection Agency: Human Alteration of the Global Nitrogen Cycle: Causes and Consequences by Peter M. Vitousek, Chair, John Aber, Robert W. Howarth, Gene E. Likens, Pamela A. Matson, David W. Schindler, William H. Schlesinger, and G. David Tilman [10] A. D. Allen, C. V. Senoff (1965). "Nitrogenopentammineruthenium(II) complexes". Journal of the Chemical Society, Chemical Communications (24): 621. doi:10.1039/C19650000621. [11] Catalytic reduction of molecular nitrogen in solutions A.E. Shilov Russian Chemical Bulletin Volume 52, Number 12, 2555-2562, doi:10.1023/B:RUCB.0000019873.81002.60 [12] Reduction of dinitrogen Richard R. Schrock PNAS November 14, 2006 vol. 103 no. 46 17087 doi:10.1073/pnas.0603633103 [13] Dinitrogen Cleavage by a Three-Coordinate Molybdenum(III) Complex Catalina E. Laplaza and Christopher C. Cummins Science 12 May 1995: 861-863.10.1126/science.268.5212.861 [14] A Cycle for Organic Nitrile Synthesis via Dinitrogen Cleavage John J. Curley, Emma L. Sceats, and Christopher C. Cummins J. Am. Chem. Soc., 2006, 128 (43), pp 1403614037 doi:10.1021/ja066090a

Nitrogen fixation
[15] Synthesis and Reactions of Molybdenum Triamidoamine Complexes Containing Hexaisopropylterphenyl Substituents Dmitry V. Yandulov, Richard R. Schrock, Arnold L. Rheingold, Christopher Ceccarelli, and William M. Davis Inorg. Chem.; 2003; 42(3) pp 796813; (Article) doi:10.1021/ic020505l [16] Catalytic Reduction of Dinitrogen to Ammonia at a Single Molybdenum Center Dmitry V. Yandulov and Richard R. Schrock Science 4 July 2003: Vol. 301. no. 5629, pp. 7678 doi:10.1126/science.1085326 [17] The catalyst is based on molybdenum(V) chloride and tris(2-aminoethyl)amine substituted with three very bulky hexa-isopropylterphenyl (HIPT) groups. Nitrogen adds end-on to the molybdenum atom, and the bulky HIPT substituents prevent the formation of the stable and nonreactive Mo-N=N-Mo dimer, and the nitrogen is reduced in an isolated pocket. The proton donor is a pyridinium cation, which is accompanied by a tetraborate counter ion. The reducing agent is decamethylchromocene. All ammonia formed is collected as the HCl salt by trapping the distillate with a HCl solution [18] Note also that, although the dinitrogen complex is shown in brackets, this species can be isolated and characterized. Here the brackets do not indicate that the intermediate is not observed. [19] A molybdenum complex bearing PNP-type pincer ligands leads to the catalytic reduction of dinitrogen into ammonia Kazuya Arashiba, Yoshihiro Miyake Yoshiaki Nishibayashi Nature Chemistry Volume: 3, Pages: 120125 Year published:(2011 doi:10.1038/nchem.906

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External links
NITROGEN FIXATION (http://lupins-bk.blogspot.com/2006/07/nitrogen-fixation.html)

Amino acid synthesis

For the non-biological synthesis of amino acids see: Strecker amino acid synthesis Amino acid synthesis is the set of biochemical processes (metabolic pathways) by which the various amino acids are produced from other compounds. The substrates for these processes are various compounds in the organism's diet or growth media. Not all organisms are able to synthesise all amino acids. For example, humans are able to synthesise only 12 of the 20 standard amino acids. A fundamental problem for biological systems is to obtain nitrogen in an easily usable form. This problem is solved by certain microorganisms capable of reducing the inert NN molecule (nitrogen gas) to two molecules of ammonia in one of the most remarkable reactions in biochemistry. Ammonia is the source of nitrogen for all the amino acids. The carbon backbones come from the glycolytic pathway, the pentose phosphate pathway, or the citric acid cycle. In amino acid production, one encounters an important problem in biosynthesis, namely stereochemical control. Because all amino acids except glycine are chiral, biosynthetic pathways must generate the correct isomer with high fidelity. In each of the 19 pathways for the generation of chiral amino acids, the stereochemistry at the -carbon atom is established by a transamination reaction that involves pyridoxal phosphate. Almost all the transaminases that catalyze these reactions descend from a common ancestor, illustrating once again that effective solutions to biochemical problems are retained throughout evolution. Biosynthetic pathways are often highly regulated such that building-blocks are synthesized only when supplies are low. Very often, a high concentration of the final product of a pathway inhibits the activity of enzymes that function early in the pathway. Often present are allosteric enzymes capable of sensing and responding to concentrations of regulatory species. These enzymes are similar in functional properties to aspartate transcarbamoylase and its regulators. Feedback and allosteric mechanisms ensure that all twenty amino acids are maintained in sufficient amounts for protein synthesis and other processes.

Amino acid synthesis

311

Amino acid synthesis

Amino acids are synthesized from -ketoacids, and later transaminated from another aminoacid, usually Glutamate. The enzyme involved in this reaction is an aminotransferase. -ketoacid + glutamate amino acid + -ketoglutarate Glutamate itself is formed by amination of -ketoglutarate: -ketoglutarate + NH glutamate

Nitrogen fixation: Microorganisms use ATP and a powerful reductant to reduce atmospheric nitrogen to ammonia
Microorganisms use ATP and reduced ferredoxin, a powerful reductant, to reduce N2 to NH3. An iron-molybdenum cluster in nitrogenase deftly catalyzes the fixation of N2, a very inert molecule. Higher organisms consume the fixed nitrogen to synthesize amino acids, nucleotides, and other nitrogen-containing biomolecules. The major points of entry of NH4+ into metabolism are glutamine or glutamate.

Amino acids are made from intermediates of the citric acid cycle and other major pathways
Of the basic set of 20 amino acids (not counting selenocysteine), there are 8 that human beings cannot synthesize. In addition, the amino acids arginine, cysteine, glycine, glutamine, histidine, proline, serine, and tyrosine are considered conditionally essential, meaning they are not normally required in the diet, but must be supplied exogenously to specific populations that do not synthesize it in adequate amounts.[1] [2] For example, enough arginine is synthesized by the urea cycle to meet the needs of an adult but perhaps not those of a growing child. Amino acids that must be obtained from the diet are called essential amino acids. Nonessential amino acids are produced in the body. The pathways for the synthesis of nonessential amino acids are quite simple. Glutamate dehydrogenase catalyzes the reductive amination of -ketoglutarate to glutamate. A transamination reaction takes place in the synthesis of most amino acids. At this step, the chirality of the amino acid is established. Alanine and aspartate are synthesized by the transamination of pyruvate and oxaloacetate, respectively. Glutamine is synthesized from NH4+ and glutamate, and asparagine is synthesized similarly. Proline and arginine are derived from glutamate. Serine, formed from 3-phosphoglycerate, is the precursor of glycine and cysteine. Tyrosine is synthesized by the hydroxylation of phenylalanine, an essential amino acid. The pathways for the biosynthesis of essential amino acids are much more complex than those for the nonessential ones. Tetrahydrofolate, a carrier of activated one-carbon units, plays an important role in the metabolism of amino acids and nucleotides. This coenzyme carries one-carbon units at three oxidation states, which are interconvertible: most reducedmethyl; intermediatemethylene; and most oxidizedformyl, formimino, and methenyl. The major donor of activated methyl groups is S-adenosylmethionine, which is synthesized by the transfer of an adenosyl group from ATP to the sulfur atom of methionine. S-Adenosylhomocysteine is formed when the activated methyl group is transferred to an acceptor. It is hydrolyzed to adenosine and homocysteine, the latter of which is then methylated to methionine to complete the activated methyl cycle. Cortisol inhibits protein synthesis.[3]

Amino acid synthesis

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Amino acid biosynthesis is regulated by feedback inhibition

Most of the pathways of amino acid biosynthesis are regulated by feedback inhibition, in which the committed step is allosterically inhibited by the final product. Branched pathways require extensive interaction among the branches that includes both negative and positive regulation. The regulation of glutamine synthetase from E. coli is a striking demonstration of cumulative feedback inhibition and of control by a cascade of reversible covalent modifications.

Amino acids are precursors of many biomolecules

Amino acids are precursors of a variety of biomolecules. Glutathione (-Glu-Cys-Gly) serves as a sulfhydryl buffer and detoxifying agent. Glutathione peroxidase, a selenoenzyme, catalyzes the reduction of hydrogen peroxide and organic peroxides by glutathione. Nitric oxide, a short-lived messenger, is formed from arginine. Porphyrins are synthesized from glycine and succinyl CoA, which condense to give -aminolevulinate. Two molecules of this intermediate become linked to form porphobilinogen. Four molecules of porphobilinogen combine to form a linear tetrapyrrole, which cyclizes to uroporphyrinogen III. Oxidation and side-chain modifications lead to the synthesis of protoporphyrin IX, which acquires an iron atom to form heme. [4]

References
[1] Frst P, Stehle P (1 June 2004). "What are the essential elements needed for the determination of amino acid requirements in humans?" (http:/ / jn. nutrition. org/ cgi/ content/ full/ 134/ 6/ 1558S). J. Nutr. 134 (6 Suppl): 1558S1565S. PMID15173430. . [2] Reeds PJ (1 July 2000). "Dispensable and indispensable amino acids for humans" (http:/ / jn. nutrition. org/ cgi/ content/ full/ 130/ 7/ 1835S). J. Nutr. 130 (7): 1835S40S. PMID10867060. . [3] Manchester, K.L., Sites of Hormonal Regulation of Protein Metabolism. p. 229, Mammalian Protein [Munro, H.N., Ed.]. Academic Press, New York. On p273. [4] Biochemistry. Berg, Jeremy M.; Tymoczko, John L.; and Stryer, Lubert. New York: W. H. Freeman and Co. ; c2002

External links
NCBI Bookshelf Free Textbook Access (http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stryer.TOC& depth=2)

Nucleotide

313

Nucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling (cGMP and cAMP), and are incorporated into important cofactors of enzymatic reactions (coenzyme A, FAD, FMN, and NADP+). Nucleotide derivatives such as the nucleoside triphosphates play central roles in metabolism, in which capacity they serve as sources of chemical energy (ATP and GTP).[1]

Structural elements of the most common nucleotides

Nucleotide structure
A nucleotide is composed of a nucleobase (nitrogenous base), a five-carbon sugar (either ribose or 2'-deoxyribose), and one phosphate group.[2] Together, the nucleobase and sugar compose a nucleoside. The phosphate groups form bonds with either the 2, 3, or 5-carbon of the sugar, with the 5-carbon site most common. Cyclic nucleotides form when the phosphate group is bound to two of the sugar's hydroxyl groups.[1] Ribonucleotides are nucleotides where the sugar is ribose, and deoxyribonucleotides contain the sugar deoxyribose. Nucleotides can contain either a purine or a pyrimidine base. Nucleic acids are polymeric macromolecules made from nucleotide Ribose structure indicating numbering of carbon monomers. In DNA, the purine bases are adenine and guanine, while atoms the pyrimidines are thymine and cytosine. RNA uses uracil in place of thymine. Adenine always pairs with thymine by 2 hydrogen bonds, while guanine pairs with cytosine through 3 hydrogen bonds, each due to their unique structures.

Synthesis
Nucleotides can be synthesized by a variety of means both in vitro and in vivo. In vivo, nucleotides can be synthesized de novo or recycled through salvage pathways.[3] The components used in de novo nucleotide synthesis are derived from biosynthetic precursors of carbohydrate and amino acid metabolism, and from ammonia and carbon dioxide. The liver is the major organ of de novo synthesis of all four nucleotides. De novo synthesis of pyrimidines and purines follows two different pathways. Pyrimidines are synthesized first from aspartate and carbamoyl-phosphate in the cytoplasm to the common precursor ring structure orotic acid, onto which a phosphorylated ribosyl unit is covalently linked. Purines, however, are first synthesized from the sugar template onto which the ring synthesis occurs. For reference, the syntheses of the purine and pyrimidine nucleotides are carried out by several enzymes in the cytoplasm of the cell, not within a specific organelle. Nucleotides undergo breakdown such that useful parts can be reused in synthesis reactions to create new nucleotides.

Nucleotide In vitro, protecting groups may be used during laboratory production of nucleotides. A purified nucleoside is protected to create a phosphoramidite, which can then be used to obtain analogues not found in nature and/or to synthesize an oligonucleotide.

314

Pyrimidine ribonucleotide synthesis

The synthesis of the pyrimidines CTP and UTP occurs in the cytoplasm and starts with the formation of carbamoyl phosphate from glutamine and CO2. Next, aspartate undergoes a condensation reaction with carbamoyl-phosphate to form orotic acid. In a subsequent cyclization reaction, the enzyme Aspartate carbamoyltransferase forms N-carbamoyl-aspartate which is converted into dihydroorotic acid by Dihydroorotase. The latter is converted to orotate by Dihydroorotate oxidase. The net reaction is: (S)-Dihydroorotate + O2 = Orotate + H2O2 Orotate is covalently linked with a phosphorylated ribosyl unit. The The synthesis of Uridine monophosphateUMP.The color scheme is as follows: enzymes, covalent linkage between the ribose coenzymes, substrate names, inorganic molecules and pyrimidine occurs at position C1 of the ribose unit, which contains a pyrophosphate, and N1 of the pyrimidine ring. Orotate phosphoribosyltransferase (aka "PRPP transferase") catalyzes the net reaction yielding orotidine monophosphate (OMP): Orotate + 5-Phospho--D-ribose 1-diphosphate (aka. "PRPP") = Orotidine 5'-phosphate + Pyrophosphate Orotidine-5-phosphate is decarboxylated by Orotidine-5'-phosphate decarboxylase to form uridine monophosphate (UMP). PRPP transferase catalyzes both the ribosylation and decarboxylation reactions, forming UMP from orotic acid in the presence of PRPP. It is from UMP that other pyrimidine nucleotides are derived. UMP is phosphorylated by two kinases to uridine triphosphate (UTP) via two sequential reactions with ATP. First the diphosphate form UDP is produced, which in turn is phosphorylated to UTP. Both steps are fueled by ATP hydrolysis: ATP + UMP = ADP + UDP UDP + ATP = UTP + ADP CTP is subsequently formed by amination of UTP by the catalytic activity of CTP synthetase. Glutamine is the NH3 donor and the reaction is fueled by ATP hydrolysis, too: UTP + Glutamine + ATP + H2O = CTP + ADP + Pi Cytidine monophosphate (CMP) is derived from cytidine triphosphate (CTP) with subsequent loss of two phosphates.[4] [5]

Nucleotide

315

Purine ribonucleotide synthesis

The atoms which are used to build the purine nucleotides come from a variety of sources:

The synthesis of IMP. The color scheme is as follows: enzymes, coenzymes, substrate names, metal ions, inorganic molecules

The biosynthetic origins of purine ring atoms N1 arises from the amine group of Asp C2 and C8 originate from formate N3 and N9 are contributed by the amide group of Gln C4, C5 and N7 are derived from Gly C6 comes from HCO3- (CO2)

The de novo synthesis of purine nucleotides by which these precursors are incorporated into the purine ring proceeds by a 10-step pathway to the branch-point intermediate IMP, the nucleotide of the base hypoxanthine. AMP and GMP are subsequently synthesized from this intermediate via separate, two-step pathways. Thus, purine moieties are initially formed as part of the ribonucleotides rather than as free bases. Six enzymes take part in IMP synthesis. Three of them are multifunctional: GART (reactions 2, 3, and 5) PAICS (reactions 6, and 7) ATIC (reactions 9, and 10) The pathway starts with the formation of PRPP. PRPS1 is the enzyme that activates R5P, which is formed primarily by the pentose phosphate pathway, to PRPP by reacting it with ATP. The reaction is unusual in that a pyrophosphoryl group is directly transferred from ATP to C1 of R5P and that the product has the configuration about C1. This reaction is also shared with the pathways for the synthesis of Trp, His, and the pyrimidine nucleotides. Being on a major metabolic crossroad and requiring much energy, this reaction is highly regulated.

Nucleotide In the first reaction unique to purine nucleotide biosynthesis, PPAT catalyzes the displacement of PRPP's pyrophosphate group (PPi) by an amide nitrogen donated from either glutamine (N), glycine (N&C), aspartate (N), folic acid (C1), or CO2. This is the committed step in purine synthesis. The reaction occurs with the inversion of configuration about ribose C1, thereby forming -5-phosphorybosylamine (5-PRA) and establishing the anomeric form of the future nucleotide. Next, a glycine is incorporated fueled by ATP hydrolysis and the carboxyl group forms an amine bond to the NH2 previously introduced. A one-carbon unit from folic acid coenzyme N10-formyl-THF is then added to the amino group of the substituted glycine followed by the closure of the imidazole ring. Next, a second NH2 group is transferred from a glutamine to the first carbon of the glycine unit. A carboxylation of the second carbon of the glycin unit is concomittantly added. This new carbon is modified by the additional of a third NH2 unit, this time transferred from an aspartate residue. Finally, a second one-carbon unit from formyl-THF is added to the nitrogen group and the ring covalently closed to form the common purine precursor inosine monophosphate (IMP). Inosine monophosphate is converted to adenosine monophosphate in two steps. First, GTP hydrolysis fuels the addition of aspartate to IMP by adenylosuccinate synthase, substituting the carbonyl oxygen for a nitrogen and forming the intermediate adenylosuccinate. Fumarate is then cleaved off forming adenosine monophosphate. This step is catalyzed by adenylosuccinate lyase. Inosine monophosphate is converted to guanosine monophosphate by the oxidation of IMP forming xanthylate, followed by the insertion of an amino group at C2. NAD+ is the electron acceptor in the oxidation reaction. The amide group transfer from glutamine is fueled by ATP hydrolysis.

316

Pyramidine and purine degradation

In humans, pyrimidine rings (C, T, U) can be degraded completely to CO2 and NH3 (urea excretion). That having been said, purine rings (G, A) cannot. Instead they are degraded to the metabolically inert uric acid which is then excreted from the body. Uric acid is formed when GMP is split into the base guanine and ribose. Guanine is deaminated to xanthine which in turn is oxidized to uric acid. This last reaction is irreversible. Similarly, uric acid can be formed when AMP is deaminated to IMP from which the ribose unit is removed to form hypoxanthine. Hypoxanthine is oxidized to xanthine and finally to uric acid. Instead of uric acid secretion, guanine and IMP can be used for recycling purposes and nucleic acid synthesis in the presence of PRPP and aspartate (NH3 donor).

Length unit
Nucleotide (abbreviated nt) is a common length unit for single-stranded RNA, similar to how base pair is a length unit for double-stranded DNA.

Abbreviation codes for degenerate bases

The IUPAC has designated the symbols for nucleotides.[6] Apart from the five (A, G, C, T/U) bases, often degenerate bases are used especially for designing PCR primers. These nucleotide codes are listed here.

Nucleotide

317

IUPAC nucleotide code A C G T (or U) R Y S W K M B D H V N . or Adenine Cytosine Guanine

Base

Thymine (or Uracil) A or G [puRine] C or T (U) [pYrimidine] G or C A or T (U) G or T (U) A or C C or G or T (U) A or G or T (U) A or C or T (U) A or C or G any base gap

References
[1] Alberts B, Johnson A, Lewis J, Raff M, Roberts K & Wlater P (2002). Molecular Biology of the Cell (4th ed.). Garland Science. ISBN 0-8153-3218-1. pp. 120-121. [2] Coghill, Anne M.; Garson, Lorrin R., ed (2006). The ACS style guide: effective communication of scientific information (3rd ed.). Washington, D.C.: American Chemical Society. p.244. ISBN9780841239999. [3] Zaharevitz, DW; Anerson, LW; Manlinowski, NM; Hyman, R; Strong, JM; Cysyk, RL.. Contribution of de-novo and salvage synthesis to the uracil nucleotide pool in mouse tissues and tumors in vivo. [4] Jones, ME (1980). "Pyrimidine nucleotide biosynthesis in animals: Genes, enzymes, and regulation of UMP biosynthesis". Ann. Rev. Biochem 49 (1): 25379. doi:10.1146/annurev.bi.49.070180.001345. PMID6105839. [5] McMurry, JE; Begley, TP (2005). The organic chemistry of biological pathways. Roberts & Company. ISBN9780974707716. [6] IUPAC nucleotide code (http:/ / users. ox. ac. uk/ ~linc1775/ blueprint. htm)

External links
Abbreviations and Symbols for Nucleic Acids, Polynucleotides and their Constituents (http://www.chem.qmul. ac.uk/iupac/misc/naabb.html) (IUPAC) Provisional Recommendations 2004 (http://www.iupac.org/reports/provisional/abstract04/BB-prs310305/ Chapter10.pdf) (IUPAC) Chemistry explanation of nucleotide structure (http://dl.clackamas.cc.or.us/ch106-09/nucleoti.htm)

Urea cycle

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Urea cycle
The urea cycle (also known as the ornithine cycle) is a cycle of biochemical reactions occurring in many animals that produces urea ((NH2)2CO) from ammonia (NH3). This cycle was the first metabolic cycle discovered (Hans Krebs and Kurt Henseleit, 1932), five years before the discovery of the TCA cycle. In mammals, the urea cycle takes place primarily in the liver, and to a lesser extent in the kidney.

Function
Organisms that cannot easily and quickly remove ammonia usually have to convert it to some other substance, like urea or uric acid, which are much less toxic. Insufficiency of the urea cycle occurs in some genetic disorders (inborn errors of metabolism), and in liver failure. The result of liver failure is accumulation of nitrogenous waste, mainly ammonia, which leads to hepatic encephalopathy.

Reactions
The urea cycle consists of five reactions: two mitochondrial and three cytosolic. The cycle converts two amino groups, one from NH4+ and one from Asp, and a carbon atom from HCO3, to the relatively nontoxic excretion product urea at the cost of four "high-energy" phosphate bonds (3 ATP hydrolyzed to 2 ADP and one AMP). Ornithine is the carrier of these carbon and nitrogen atoms.

Reactions of the urea cycle

Step 1 2 3 4 5 Reactants NH4+ + HCO3 + 2ATP Products Catalyzed by Location mitochondria mitochondria cytosol cytosol cytosol

carbamoyl phosphate + 2ADP + Pi CPS1 OTC ASS ASL ARG1

carbamoyl phosphate + ornithine citrulline + Pi citrulline + aspartate + ATP argininosuccinate Arg + H2O argininosuccinate + AMP + PPi Arg + fumarate ornithine + urea

The reactions of the urea cycle

Urea cycle

319 1 L-ornithine 2 carbamoyl phosphate 3 L-citrulline 4 argininosuccinate 5 fumarate 6 L-arginine 7 urea L-Asp L-aspartate CPS-1 carbamoyl phosphate synthetase I OTC Ornithine transcarbamoylase ASS argininosuccinate synthetase ASL argininosuccinate lyase ARG1 arginase 1 In the first reaction, NH4+ + HCO3 is equivalent to NH3 + CO2 + H2O. Thus, the overall equation of the urea cycle is: NH3 + CO2 + aspartate + 3 ATP + 2 H2O urea + fumarate + 2 ADP + 2 Pi + AMP + PPi Since fumarate is obtained by removing NH3 from aspartate (by means of reactions 3 and 4), and PPi + H2O 2 Pi, the equation can be simplified as follows: 2 NH3 + CO2 + 3 ATP + H2O urea + 2 ADP + 4 Pi

+ AMP Note that reactions related to the urea cycle also cause the oxidation of 2 NADH, so the urea cycle releases slightly more energy than it consumes. These NADH are produced in two ways: One NADH molecule is reduced by the enzyme glutamate dehydrogenase in the conversion of glutamate to ammonium and -ketoglutarate. Glutamate is the non-toxic carrier of amine groups. This provides the ammonium ion used in the initial synthesis of carbamoyl phosphate. The fumarate released in the cytosol is converted to malate by cytosolic fumarase. This malate is then converted to oxaloacetate by cytosolic malate dehydrogenase, generating a reduced NADH in the cytosol. Oxaloacetate is one of the keto acids preferred by transaminases, and so will be recycled to aspartate, maintained the flow of nitrogen into the urea cycle. The two NADH produced can provide energy for the formation of 5 ATP, a net production of one high-energy phosphate bond for the urea cycle. However, if gluconeogenesis is underway in the cytosol, the latter reducing equivalent is used to drive the reversal of the GAPDH step instead of generating ATP. The fate of oxaloacetate is either to produce aspartate via transamination or to be converted to phosphoenol pyruvate, which is a substrate to glucose.

Urea cycle

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Regulation
N-Acetylglutamic acid
The synthesis of carbamoyl phosphate and the urea cycle are dependent on the presence of NAcGlu, which allosterically activates CPS1. Synthesis of NAcGlu by NAGS, is stimulated by both Arg, allosteric stimulator of NAGS, and Glu, a product in the transamination reactions and one of NAGS's substrates, both of which are elevated when free amino acids are elevated. So, Arg is not only a substrate for the urea cycle reactions but also serves as an activator for the urea cycle.

Substrate concentrations
The remaining enzymes of the cycle are controlled by the concentrations of their substrates. Thus, inherited deficiencies in the cycle enzymes other than ARG1 do not result in significant decrease in urea production (the total lack of any cycle enzyme results in death shortly after birth). Rather, the deficient enzyme's substrate builds up, increasing the rate of the deficient reaction to normal. The anomalous substrate buildup is not without cost, however. The substrate concentrations become elevated all the way back up the cycle to NH4+, resulting in hyperammonemia (elevated [NH4+]P). Although the root cause of NH4+ toxicity is not completely understood, a high [NH4+] puts an enormous strain on the NH4+-clearing system, especially in the brain (symptoms of urea cycle enzyme deficiencies include mental retardation and lethargy). This clearing system involves GLUD1 and GLUL, which decrease the 2-oxoglutarate (2OG) and Glu pools. The brain is most sensitive to the depletion of these pools. Depletion of 2OG decreases the rate of TCAC, whereas Glu is both a neurotransmitter and a precursor to GABA, another neurotransmitter. [1](p.734)

Pathology
Anomalies of the urea cycle cause urea cycle disorders: ornithine transcarbamoylase deficiency Carbamoyl phosphate synthetase deficiency (Ornithine translocase deficiency) Argininosuccinic aciduria Argininemia Hyperornithinemia, hyperammonemia, homocitrullinuria syndrome (HHH syndrome) Lysinuric protein intolerance Citrullinemia N-Acetylglutamate synthase deficiency

Most of them are associated with hyperammonemia.

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Additional images

Urea cycle.

Urea cycle colored.

External links
The chemical logic behind the urea cycle [2] Basic Neurochemistry [3] - amino acid disorders

References
[1] http:/ / www. wiley. com/ college/ math/ chem/ cg/ sales/ voet. html [2] http:/ / homepage. ufp. pt/ pedros/ bq/ urea. htm [3] http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=bnchm. figgrp. 3102

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Integration of metabolism
Hormone
A hormone (from Greek "impetus") is a chemical released by a cell or a gland in one part of the body that sends out messages that affect cells in other parts of the organism. Only a small amount of hormone is required to alter cell metabolism. In essence, it is a chemical messenger that transports a signal from one cell to another. All multicellular organisms produce hormones; plant hormones are also called phytohormones. Hormones in animals are often transported in the blood. Cells respond to a hormone when they express a specific receptor for that hormone. The hormone binds to the receptor protein, resulting in the activation of a signal transduction mechanism that ultimately leads to cell type-specific responses.

Epinephrine (adrenaline), a catecholamine-type hormone

Endocrine hormone molecules are secreted (released) directly into the bloodstream, whereas exocrine hormones (or ectohormones) are secreted directly into a duct, and, from the duct, they flow either into the bloodstream or from cell to cell by diffusion in a process known as paracrine signalling. Recently it has been found that a variety of exogenous modern chemical compounds have hormone-like effects on both humans and wildlife. Their interference with the synthesis, secretion, transport, binding, action, or elimination of natural hormones in the body can change the homeostasis, reproduction, development, and/or behavior the same as endogenous produced hormones."[1]

Hormones as signals
Hormonal signaling involves the following: 1. 2. 3. 4. 5. Biosynthesis of a particular hormone in a particular tissue Storage and secretion of the hormone Transport of the hormone to the target cell(s) Recognition of the hormone by an associated cell membrane or intracellular receptor protein Relay and amplification of the received hormonal signal via a signal transduction process: This then leads to a cellular response. The reaction of the target cells may then be recognized by the original hormone-producing cells, leading to a down-regulation in hormone production. This is an example of a homeostatic negative feedback loop. 6. Degradation of the hormone. Hormone cells are typically of a specialized cell type, residing within a particular endocrine gland, such as thyroid gland, ovaries, and testes. Hormones exit their cell of origin via exocytosis or another means of membrane transport. The hierarchical model is an oversimplification of the hormonal signaling process. Cellular recipients of a particular hormonal signal may be one of several cell types that reside within a number of different tissues, as is the case for insulin, which triggers a diverse range of systemic physiological effects. Different tissue types may also respond differently to the same hormonal signal. Because of this, hormonal signaling is elaborate and hard to dissect.

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Interactions with receptors

Most hormones initiate a cellular response by initially combining with either a specific intracellular or cell membrane associated receptor protein. A cell may have several different receptors that recognize the same hormone and activate different signal transduction pathways, or a cell may have several different receptors that recognize different hormones and activate the same biochemical pathway. For many hormones, including most protein hormones, the receptor is membrane-associated and embedded in the plasma membrane at the surface of the cell. The interaction of hormone and receptor typically triggers a cascade of secondary effects within the cytoplasm of the cell, often involving phosphorylation or dephosphorylation of various other cytoplasmic proteins, changes in ion channel permeability, or increased concentrations of intracellular molecules that may act as secondary messengers (e.g., cyclic AMP). Some protein hormones also interact with intracellular receptors located in the cytoplasm or nucleus by an intracrine mechanism. For hormones such as steroid or thyroid hormones, their receptors are located intracellularly within the cytoplasm of their target cell. To bind their receptors, these hormones must cross the cell membrane. They can do so because they are lipid-soluble. The combined hormone-receptor complex then moves across the nuclear membrane into the nucleus of the cell, where it binds to specific DNA sequences, effectively amplifying or suppressing the action of certain genes, and affecting protein synthesis.[2] However, it has been shown that not all steroid receptors are located intracellularly. Some are associated with the plasma membrane.[3] An important consideration, dictating the level at which cellular signal transduction pathways are activated in response to a hormonal signal, is the effective concentration of hormone-receptor complexes that are formed. Hormone-receptor complex concentrations are effectively determined by three factors: 1. The number of hormone molecules available for complex formation 2. The number of receptor molecules available for complex formation 3. The binding affinity between hormone and receptor. The number of hormone molecules available for complex formation is usually the key factor in determining the level at which signal transduction pathways are activated, the number of hormone molecules available being determined by the concentration of circulating hormone, which is in turn influenced by the level and rate at which they are secreted by biosynthetic cells. The number of receptors at the cell surface of the receiving cell can also be varied, as can the affinity between the hormone and its receptor.

Physiology of hormones
Most cells are capable of producing one or more molecules, which act as signaling molecules to other cells, altering their growth, function, or metabolism. The classical hormones produced by cells in the endocrine glands mentioned so far in this article are cellular products, specialized to serve as regulators at the overall organism level. However, they may also exert their effects solely within the tissue in which they are produced and originally released. The rate of hormone biosynthesis and secretion is often regulated by a homeostatic negative feedback control mechanism. Such a mechanism depends on factors that influence the metabolism and excretion of hormones. Thus, higher hormone concentration alone cannot trigger the negative feedback mechanism. Negative feedback must be triggered by overproduction of an "effect" of the hormone. Hormone secretion can be stimulated and inhibited by: Other hormones (stimulating- or releasing -hormones) Plasma concentrations of ions or nutrients, as well as binding globulins Neurons and mental activity Environmental changes, e.g., of light or temperature

Hormone One special group of hormones is the tropic hormones that stimulate the hormone production of other endocrine glands. For example, thyroid-stimulating hormone (TSH) causes growth and increased activity of another endocrine gland, the thyroid, which increases output of thyroid hormones. A recently identified class of hormones is that of the "hunger hormones" - ghrelin, orexin, and PYY 3-36 - and "satiety hormones" - e.g., cholecystokinin, leptin, nesfatin-1, obestatin. To release active hormones quickly into the circulation, hormone biosynthetic cells may produce and store biologically inactive hormones in the form of pre- or prohormones. These can then be quickly converted into their active hormone form in response to a particular stimulus.

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Effects of hormones
Hormones have the following effects on the body: stimulation or inhibition of growth mood swings induction or suppression of apoptosis (programmed cell death) activation or inhibition of the immune system regulation of metabolism preparation of the body for mating, fighting, fleeing, and other activity preparation of the body for a new phase of life, such as puberty, parenting, and menopause control of the reproductive cycle hunger cravings Sexual arousal

A hormone may also regulate the production and release of other hormones. Hormone signals control the internal environment of the body through homeostasis.

Chemical classes of hormones

Vertebrate hormones fall into three chemical classes: Peptide hormones consist of chains of amino acids. Examples of small peptide hormones are TRH and vasopressin. Peptides composed of scores or hundreds of amino acids are referred to as proteins. Examples of protein hormones include insulin and growth hormone. More complex protein hormones bear carbohydrate side-chains and are called glycoprotein hormones. Luteinizing hormone, follicle-stimulating hormone and thyroid-stimulating hormone are glycoprotein hormones. There is also another type of hydrophilic hormone called nonpeptide hormones. Although they don't have peptide connections, they are assimilated as peptide hormones. Lipid and phospholipid-derived hormones derive from lipids such as linoleic acid and arachidonic acid and phospholipids. The main classes are the steroid hormones that derive from cholesterol and the eicosanoids. Examples of steroid hormones are testosterone and cortisol. Sterol hormones such as calcitriol are a homologous system. The adrenal cortex and the gonads are primary sources of steroid hormones. Examples of eicosanoids are the widely studied prostaglandins. Monoamines derived from aromatic amino acids like phenylalanine, tyrosine, tryptophan by the action of aromatic amino acid decarboxylase enzymes.

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Pharmacology
Many hormones and their analogues are used as medication. The most commonly prescribed hormones are estrogens and progestagens (as methods of hormonal contraception and as HRT), thyroxine (as levothyroxine, for hypothyroidism) and steroids (for autoimmune diseases and several respiratory disorders). Insulin is used by many diabetics. Local preparations for use in otolaryngology often contain pharmacologic equivalents of adrenaline, while steroid and vitamin D creams are used extensively in dermatological practice. A "pharmacologic dose" of a hormone is a medical usage referring to an amount of a hormone far greater than naturally occurs in a healthy body. The effects of pharmacologic doses of hormones may be different from responses to naturally occurring amounts and may be therapeutically useful. An example is the ability of pharmacologic doses of glucocorticoid to suppress inflammation.

Important human hormones

See: List of human hormones

References
[1] Crisp TM, Clegg ED, Cooper RL, Wood WP, Anderson DG, Baetcke KP, Hoffmann JL, Morrow MS, Rodier DJ, Schaeffer JE, Touart LW, Zeeman MG, Patel YM (1998). "Environmental endocrine disruption: An effects assessment and analysis". Environ. Health Perspect. 106 (Suppl 1): 1156. PMC1533291. PMID9539004. [2] Beato M, Chavez S and Truss M (1996). "Transcriptional regulation by steroid hormones". Steroids 61 (4): 240251. doi:10.1016/0039-128X(96)00030-X. PMID8733009. [3] Hammes SR (2003). "The further redefining of steroid-mediated signaling". Proc Natl Acad Sci USA 100 (5): 216802170. doi:10.1073/pnas.0530224100. PMC151311. PMID12606724.

External links
The Hormone Foundation (http://www.hormone.org) Article on hormones and their receptors (http://www.biomedcentral.com/1471-2164/10/307/) HMRbase: A database of hormones and their receptors (http://crdd.osdd.net/raghava/hmrbase/) MeSH Hormones (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=&term=Hormones)

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Signal transduction
Signal transduction occurs when an extracellular signaling molecule activates a cell surface receptor. In turn, this receptor alters intracellular molecules creating a response.[1] There are two stages in this process: 1. A signaling molecule activates a specific receptor on the cell membrane 2. Causing a second messenger to continue the signal into the cell and elicit a physiological response. In either step, the signal can be amplified. Thus, one signalling molecule can cause many responses.[2]
An overview of major signal transduction pathways.

History
In 1970, Martin Rodbell examined the effects of glucagon on a rat's liver cell membrane receptor. He noted that guanosine triphosphate disassociated glucagon from this receptor and stimulated the G-protein, which strongly influenced the cell's metabolism. Thus he deduced that the G-protein was a transducer that accepted glucagon molecules and affected the cell.[3] For this he shared the 1994 Nobel Prize in Physiology or Medicine with Alfred G. Gilman. The current understanding of signal transduction processes reflects contributions made by Rodbell and many other research groups.

Occurrence of the term signal transduction in papers since 1977. These figures were extracted through an analysis of the papers contained within the MEDLINE database.

The earliest scientific paper recorded in the MEDLINE database as containing the specific term signal transduction was published in 1972.[4] Some articles published before 1977 used the term signal transmission or sensory transduction for signal transduction:[5] [6] a total of 48,377 scientific papers related to signal transduction were published in 1977, of which 11,211 were reviews of other papers. That year the actual term signal transduction was included in abstracts until in 1979 it appeared in a paper's title.[7] [8] One source attributes the widespread use of this term to a 1980 review article by Rodbell:[9] [3] research papers directly addressing signal transduction processes began to appear in large numbers in the late 1980s and early 1990s.
[10]

==Signaling molecules==Sordner (talk) 00:05, 7 December 2011 (UTC)

Signal transduction involves the binding of extracellular signalling molecules and ligands to cell-surface receptors that trigger events inside the cell. The combination of messenger with receptor causes a change in the conformation of the receptor, known as receptor activation. This activation is always the initial step (the cause) leading to the cell's ultimate responses (effect) to the messenger. Despite the myriad of these ultimate responses, they are all directly due to changes in particular cell proteins. Intracellular signaling cascades can be started through cell-substratum

Signal transduction interactions; examples are the integrin that binds ligands in the extracellular matrix and steroids.[11] Most steroid hormones have receptors within the cytoplasm and act by stimulating the binding of their receptors to the promoter region of steroid-responsive genes.[12] Examples of signaling molecules include the hormone melatonin,[13] the neurotransmitter acetylcholine[14] and the cytokine interferon .[15] The classifications of signalling molecules do not take into account the molecular nature of each class member; neurotransmitters range in size from small molecules such as dopamine[16] to neuropeptides such as endorphins.[17] Some molecules may fit into more than one class; for example, epinephrine is a neurotransmitter when secreted by the central nervous system and a hormone when secreted by the adrenal medulla.

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Environmental stimuli
With single-celled organisms, the variety of signal transduction processes influence its reaction to its environment. With multicellular organisms, numerous processes are required for coordinating individual cells to support the organism as a whole; the complexity of these processes tend to increase with the complexity of the organism. Sensing of environments at the cellular level relies on signal transduction; many disease processes, such as diabetes and heart disease arise from defects in these pathways, highlighting the importance of this process in biology and medicine. Various environmental stimuli exist that initiate signal transmission processes in multicellular organisms; examples include photons hitting cells in the retina of the eye,[18] and odorants binding to odorant receptors in the nasal epithelium.[19] Certain microbial molecules, such as viral nucleotides and protein antigens, can elicit an immune system response against invading pathogens mediated by signal transduction processes. This may occur independent of signal transduction stimulation by other molecules, as is the case for the toll-like receptor. It may occur with help from stimulatory molecules located at the cell surface of other cells, as with T-cell receptor signaling. Single-celled organisms may respond to environmental stimuli through the activation of signal transduction pathways. For example, slime molds secrete cyclic adenosine monophosphate upon starvation, stimulating individual cells in the immediate environment to aggregate,[20] and yeast cells use mating factors to determine the mating types of other cells and to participate in sexual reproduction.[21]

Receptors
Receptors can be roughly divided into two major classes: intracellular receptors and extracellular receptors.

Extracellular
Extracellular receptors are integral transmembrane proteins and make up most receptors. They span the plasma membrane of the cell, with one part of the receptor on the outside of the cell and the other on the inside. Signal transduction occurs as a result of a ligand binding to the outside; the molecule does not pass through the membrane. This binding stimulates a series of events inside the cell; different types of receptor stimulate different responses and receptors typically respond to only the binding of a specific ligand. Upon binding, the ligand induces a change in the conformation of the inside part of the receptor.[22] These result in either the activation of an enzyme in the receptor or the exposure of a binding site for other intracellular signaling proteins within the cell, eventually propagating the signal through the cytoplasm. In eukaryotic cells, most intracellular proteins activated by a ligand/receptor interaction possess an enzymatic activity; examples include tyrosine kinase and phosphatases. Some of them create second messengers such as cyclic AMP and IP3, the latter controlling the release of intracellular calcium stores into the cytoplasm. Other activated proteins interact with adapter proteins that facilitate signalling protein interactions and coordination of signalling complexes necessary to respond to a particular stimulus. Enzymes and adapter proteins are both responsive to various second messenger molecules.

Signal transduction Many adapter proteins and enzymes activated as part of signal transduction possess specialized protein domains that bind to specific secondary messenger molecules. For example, calcium ions bind to the EF hand domains of calmodulin, allowing it to bind and activate calmodulin-dependent kinase. PIP3 and other phosphoinositides do the same thing to the Pleckstrin homology domains of proteins such as the kinase protein AKT. G protein-coupled G protein-coupled receptors (GPCRs) are a family of integral transmembrane proteins that possess seven transmembrane domains and are linked to a heterotrimeric G protein. Many receptors are in this family, including adrenergic receptors and chemokine receptors. Signal transduction by a GPCR begins with an inactive G protein coupled to the receptor; it exists as a heterotrimer consisting of G, G, and G. Once the GPCR recognizes a ligand, the conformation of the receptor changes to activate the G protein, causing G to bind a molecule of GTP and dissociate from the other two G-protein subunits. The dissociation exposes sites on the subunits that can interact with other molecules.[23] The activated G protein subunits detach from the receptor and initiate signalling from many downstream effector proteins such as phospholipases and ion channels, the latter permitting the release of second messenger molecules.[24] The total strength of signal amplification by a GPCR is determined by the lifetimes of the ligand-receptor complex and receptor-effector protein complex and the deactivation time of the activated receptor and effectors through intrinsic enzymatic activity. A study was conducted where a point mutation was inserted into the gene encoding the chemokine receptor CXCR2; mutated cells underwent a malignant transformation due to the expression of CXCR2 in an active conformation despite the absence of chemokine-binding. This meant that chemokine receptors participate in cancer development.[25] Tyrosine and histidine kinase Receptor tyrosine kinases (RTKs) are transmembrane proteins with an intracellular kinase domain and an extracellular domain that binds ligands; examples include growth factor receptors such as the insulin receptor.[26] To perform signal transduction, RTKs need to form dimers in the plasma membrane;[27] the dimer is stabilized by ligands binding to the receptor. The interaction between the cytoplasmic domains stimulates the autophosphorylation of tyrosines within the domains of the RTKs, causing conformational changes. The receptors' kinase domains are subsequently activated, initiating phosphorylation signalling cascades of downstream cytoplasmic molecules that facilitate various cellular processes such as cell differentiation and metabolism.[26] As is the case with GPCRs, proteins that bind GTP play a major role in signal transduction from the activated RTK into the cell. In this case, the G proteins are members of the Ras, Rho, and Raf families, referred to collectively as small G proteins. They act as molecular switches usually tethered to membranes by isoprenyl groups linked to their carboxyl ends. Upon activation, they assign proteins to specific membrane subdomains where they participate in signaling. Activated RTKs in turn activate small G proteins that activate guanine nucleotide exchange factors such as SOS1. Once activated, these exchange factors can activate more small G proteins, thus amplifying the receptor's initial signal. The mutation of certain RTK genes, as with that of GPCRs, can result in the expression of receptors that exist in a constitutively-activate state; such mutated genes may act as oncogenes.[28] Histidine-specific protein kinases are structurally distinct from other protein kinases and are found in prokaryotes, fungi, and plants as part of a two-component signal transduction mechanism: a phosphate group from ATP is first added to a histidine residue within the kinase, then transferred to an aspartate residue on a receiver domain on a different protein or the kinase itself, thus activating the aspartate residue.[29]

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Signal transduction Integrin Integrins are produced by a wide variety of cells; they play a role in cell attachment to other cells and the extracellular matrix and in the transduction of signals from extracellular matrix components such as fibronectin and collagen. Ligand binding to the extracellular domain of integrins changes the protein's conformation, clustering it at the cell membrane to initiate signal transduction. Integrins lack kinase activity; hence integrin-mediated signal transduction is achieved through a variety of intracellular protein kinases and adaptor molecules, the main coordinator being integrin-linked kinase.[30] As shown in the picture to the right, cooperative integrin-RTK signalling determines the timing of cellular survival, apoptosis, proliferation, and differentiation.

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An overview of integrin-mediated signal transduction, adapted from Hehlgens et [30] al. (2007).

Important differences exist between integrin signalling in circulating blood cells and non-circulating cells such as epithelial cells; integrins of circulating cells are normally inactive. For example, cell membrane integrins on circulating leukocytes are maintained in an inactive state to avoid epithelial cell attachment; they are only activated in response to stimuli such as those received at the site of an inflammatory response. In a similar manner, integrins at the cell membrane of circulating platelets are normally kept inactive to avoid thrombosis. Epithelial cells (which are non-circulating) normally have active integrins at their cell membrane, helping maintain their stable adhesion to underlying stromal cells that provide signals to maintain normal functioning.[31] Toll gate When activated, toll-like receptors (TLRs) take adapter molecules within the cytoplasm of cells in order to propagate a signal. Four adaptor molecules are known to be involved in signaling, which are Myd88, TIRAP, TRIF, and TRAM.[32] [33] [34] These adapters activate other intracellular molecules such as IRAK1, IRAK4, TBK1, and IKKi that amplify the signal, eventually leading to the induction or suppression of genes that cause certain responses. Thousands of genes are activated by TLR signaling, implying that this method constitutes an important gateway for gene modulation. Ligand-gated ion channel A ligand-gated ion channel, upon binding with a ligand, changes conformation to open a channel in the cell membrane through which ions relaying signals can pass. An example of this mechanism is found in the receiving cell of a neural synapse. The influx of ions that occurs in response to these channels opening induces action potentials, such as those that travel along nerves, by depolarizing the membrane of post-synaptic cells, resulting in the opening of voltage-gated ion channels. An example of an ion allowed into the cell during a ligand-gated ion channel opening is Ca2+; it acts as a second messenger initiating signal transduction cascades and altering the physiology of the responding cell. This results in amplification of the synapse response between synaptic cells by remodelling the dendritic spines involved in the synapse.

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Intracellular
Further information: Intracellular receptor Intracellular receptors, such as nuclear receptors and cytoplasmic receptors, are soluble proteins localized within their respective areas. The typical ligands for nuclear receptors are lipophilic hormones like the steroid hormones testosterone and progesterone and derivatives of vitamins A and D. To initiate signal transduction, the ligand must pass through the plasma membrane by passive diffusion. On binding with the receptor, the ligands pass through the nuclear membrane into the nucleus, enabling gene transcription and protein production. Activated nuclear receptors attach to the DNA at receptor-specific hormone-responsive element (HRE) sequences, located in the promoter region of the genes activated by the hormone-receptor complex. Due to them enabling gene transcription, they are alternatively called inductors of gene expression. Activation of gene transcription is slower than signals directly affecting existing proteins; therefore, the effects of hormones that use nucleic receptors are long-term. Signal transduction via these receptors involves little proteins, but the details of gene regulation by this method are not well understood. Nucleic receptors have DNA-binding domains containing zinc fingers and a ligand-binding domain; the zinc fingers stabilize DNA binding by holding its phosphate backbone. DNA sequences that match the receptor are usually hexameric repeats of any kind; the sequences are similar but their orientation and distance differentiate them. The ligand-binding domain is additionally responsible for dimerization of nucleic receptors prior to binding and providing structures for transactivation used for communication with the translational apparatus. Steroid receptors are a subclass of nuclear receptors located primarily within the cytosol; in the absence of steroids, they cling together in an aporeceptor complex containing chaperone or heatshock proteins (HSPs). The HSPs are necessary to activate the receptor by assisting the protein to fold in a way such that the signal sequence enabling its passage into the nucleus is accessible. Steroid receptors, on the other hand, may be repressive on gene expression when their transactivation domain is hidden; activity can be enhanced by phosphorylation of serine residues at their N-terminal as a result of another signal transduction pathway, a process called crosstalk. Retinoic acid receptors are another subset of nuclear receptors. They can be activated by an endocrine-synthesized ligand that entered the cell by diffusion, a ligand synthesised from a precursor like retinol brought to the cell through the bloodstream or a completely intracellularly synthesised ligand like prostaglandin. These receptors are located in the nucleus and are not accompanied by HSPs; they repress their gene by binding to their specific DNA sequence when no ligand binds to them and vice versa. Certain intracellular receptors of the immune system are cytoplasmic receptors; recently identified NOD-like receptors (NLRs) reside in the cytoplasm of some eukaryotic cells and interact with ligands using a leucine-rich repeat (LRR) motif similar to TLRs. Some of these molecules like NOD2 interact with RIP2 kinase that activates NF-B signaling, whereas others like NALP3 interact with inflammatory caspases and initiate processing of particular cytokines like interleukin-1.[35]
[36]

== Second messengers ==

First messengers are the intracellular chemical messengers (hormones, neurotransmitters, and panacrine/autocrine agents) which reach the cell from from the extracellular fluid and bind to their specific receptors. Second messengers are the substances which enter the cytoplasm and act within the cell to trigger a response. Second messengers essentially serve as chemical relays from the plasma membrane to the cytoplasm, thus carrying out intracellular signal transduction. Sordner (talk) 00:22, 7 December 2011 (UTC)

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Calcium
The release of calcium ions from the endoplasmic reticulum into the cytosol results in its binding to signaling proteins that are then activated; it is then sequestered in the smooth endoplasmic reticulum and the mitochondria. Two combined receptor/ion channel proteins control the transport of calcium: the InsP3-receptor that transports calcium upon interaction with inositol triphosphate on its cytosolic side and the ryanodine receptor named after the alkaloid ryanodine, similar to the InsP3 receptor but having a feedback mechanism that releases more calcium upon binding with it. The nature of calcium in the cytosol means that it is active for only a very short time, meaning its free state concentration is very low and is mostly bound to organelle molecules like calreticulin when inactive. Calcium is used in many processes including muscle contraction, neurotransmitter release from nerve endings and cell migration. The three main pathways that lead to its activation are GPCR pathways, RTK pathways and gated ion channels; it regulates proteins either directly or by binding to an enzyme.

Lipophilics
Lipophilic second messenger molecules are derived from lipids residing in cellular membranes; enzymes stimulated by activated receptors activate the lipids by modifying them. Examples include diacylglycerol and ceramide, the former required for the activation of protein kinase C.

Nitric oxide
Nitric oxide (NO) acts as a second messenger because it is a free radical that can diffuse through the plasma membrane and affect nearby cells. It is synthesised from arginine and oxygen by the NO synthase and works through activation of soluble guanylyl cyclase, which when activated produces another second messenger, cGMP. NO can also act through covalent modification of proteins or their metal co-factors; some have a redox mechanism and are reversible. It is toxic in high concentrations and causes damage during stroke, but is the cause of many other functions like relaxation of blood vessels, apoptosis and erections.

Redox Signaling
In addition to nitric oxide, other electronically-activated species are also signal-transducing agents in a process called redox signaling. Examples include superoxide, hydrogen peroxide, carbon monoxide, and hydrogen sulfide. Redox signaling also includes active modulation of electonic flows in semiconductive macromolecules.

Cellular responses
Gene activations[37] and metabolism alterations[38] are examples of cellular responses to extracellular stimulation that require signal transduction. Gene activation leads to further cellular effects, since the products of responding genes include instigators of activation; transcription factors produced as a result of a signal transduction cascade can activate even more genes. Hence, an initial stimulus can trigger the expression of a large number of genes, leading to physiological events like the increased uptake of glucose from the blood stream[38] and the migration of neutrophils to sites of infection. The set of genes and their activation order to certain stimuli is referred to as a genetic program.[39] Mammalian cells require stimulation for cell division and survival; in the absence of growth factor, apoptosis ensues. Such requirements for extracellular stimulation are necessary for controlling cell behavior in unicellular and multicellular organisms; signal transduction pathways are perceived to be so central to biological processes that a large number of diseases are attributed to their disregulation.

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Major pathways
Following are some major signaling pathways, demonstrating how ligands binding to their receptors can affect second messengers and eventually result in altered cellular responses. MAPK/ERK pathway: A pathway that couples intracellular responses to the binding of growth factors to cell surface receptors. This pathway is very complex and includes many protein components.[40] In many cell types, activation of this pathway promotes cell division, and many forms of cancer are associated with aberrations in it.[41] cAMP dependent pathway: In humans, cAMP works by activating protein kinase A (PKA, cAMP-dependent protein kinase) (see picture), and thus, further effects mainly depend on cAMP-dependent protein kinase, which vary based on the type of cell. IP3/DAG pathway: PLC cleaves the phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) yielding diacyl glycerol (DAG) and inositol 1,4,5-triphosphate (IP3). DAG remains bound to the membrane, and IP3 is released as a soluble structure into the cytosol. IP3 then diffuses through the cytosol to bind to IP3 receptors, particular calcium channels in the endoplasmic reticulum (ER). These channels are specific to calcium and only allow the passage of calcium to move through. This causes the cytosolic concentration of Calcium to increase, causing a cascade of intracellular changes and activity.[42] In addition, calcium and DAG together works to activate PKC, which goes on to phosphorylate other molecules, leading to altered cellular activity. End effects include taste, manic depression, tumor promotion, etc.[42]

References
[1] Silverthorn (2007). Human Physiology. 4th ed. [2] Reece, Jane; Campbell, Neil (2002). Biology. San Francisco: Benjamin Cummings. ISBN0-8053-6624-5. [3] Rodbell, M. (1980). "The role of hormone receptors and GTP-regulatory proteins in membrane transduction". Nature 284 (5751): 1722. doi:10.1038/284017a0. PMID6101906. [4] Rensing, L. (1972). "Periodic geophysical and biological signals as Zeitgeber and exogenous inducers in animal organisms". Int. J. Biometeorol. 16: Suppl:113125. PMID4621276. [5] Tonndorf J. (1975). "Davis-1961 revisited. Signal transmission in the cochlear hair cell-nerve junction". Arch. Otolaryngol. 101 (9): 528535. PMID169771. [6] Ashcroft SJ, Crossley JR, Crossley PC. (1976). "The effect of N-acylglucosamines on the biosynthesis and secretion of insulin in the rat". Biochem. J. 154 (3): 701707. PMC1172772. PMID782447. [7] Hildebrand E. (1977). "What does Halobacterium tell us about photoreception?". Biophys. Struct. Mech. 3 (1): 6977. doi:10.1007/BF00536457. PMID857951. [8] Kenny JJ, Martinez-Maza O. et al (1979). "Lipid synthesis: an indicator of antigen-induced signal transduction in antigen-binding cells". J. Immunol. 112 (4): 12781284. PMID376714. [9] Gomperts, BD.; Kramer, IM. Tatham, PER. (2002). Signal transduction. Academic Press. ISBN0-12-289631-9. [10] Vander "et al" (1998). Human Physiology. McGraw-Hill. pp.159. ISBN0-07-067065-X. [11] Beato M, Chavez S and Truss M (1996). "Transcriptional regulation by steroid hormones". Steroids 61 (4): 240251. doi:10.1016/0039-128X(96)00030-X. PMID8733009. [12] Hammes SR (2003). "The further redefining of steroid-mediated signaling". Proc Natl Acad Sci USA 100 (5): 216802170. doi:10.1073/pnas.0530224100. PMC151311. PMID12606724. [13] Sugden D, Davidson K. et al. (2004). "Melatonin, melatonin receptors and melanophores: a moving story". Pigment Cell Res. 17 (5): 454460. doi:10.1111/j.1600-0749.2004.00185.x. PMID15357831. [14] Kistler J, Stroud RM et al. (1982). "Structure and function of an acetylcholine receptor". Biophys. J. 37 (1): 371383. Bibcode1982BpJ....37..371K. doi:10.1016/S0006-3495(82)84685-7. PMC1329155. PMID7055628. [15] Schroder et al. (2004). "Interferon- an overview of signals, mechanisms and functions" (http:/ / www. jleukbio. org/ cgi/ content/ full/ 75/ 2/ 163). Journal of Leukocyte Biology 75 (2): 163189. doi:10.1189/jlb.0603252. PMID14525967. . [16] Missale C, Nash SR. et al (1998). "Dopamine receptors:from structure to function". Physiol. Rev. 78 (1): 189225. PMID9457173. [17] Goldstein, A. (1976). "Opioid peptides endorphins in pituitary and brain". Science 193 (4258): 10811086. doi:10.1126/science.959823. PMID959823. [18] Burns ME and Arshavsky VY. (2005). "Beyond counting photons: trials and trends in vertebrate visual transduction". Neuron 48 (3): 387401. doi:10.1016/j.neuron.2005.10.014. PMID16269358.

Signal transduction
[19] Ronnett GV and Moon C. (2002). "G proteins and olfactory signal transduction". Annu Rev Physiol 64 (1): 189222. doi:10.1146/annurev.physiol.64.082701.102219. PMID11826268. [20] Hanna MH, Nowicki JJ and Fatone MA (1984). "Extracellular cyclic AMP (cAMP) during development of the cellular slime mold Polysphondylium violaceum: comparison of accumulation in the wild type and an aggregation-defective mutant". J Bacteriol 157 (2): 345349. PMID215252. [21] Sprague GF Jr. (1991). "Signal transduction in yeast mating: receptors, transcription factors, and the kinase connection". Trends Genet 7 (1112): 393398. doi:10.1016/0168-9525(91)90218-F. PMID1668192. [22] A molecular model for receptor activation (http:/ / www. bio-balance. com/ JMGM_article. pdf) [23] Jeremy M. Berg, John L. Tymoczko, Lubert Stryer; Web content by Neil D. Clarke (2002). Biochemistry. San Francisco: W.H. Freeman. ISBN0-7167-4954-8. [24] Yang W, Xia S (2006). "Mechanisms of regulation and function of G-protein-coupled receptor kinases". World J Gastroenterol 12 (48): 77537. PMID17203515. [25] Burger M, Burger, JA et al (1999). "Point mutation causing constitutive signaling of CXCR2 leads to transforming activity similar to Kaposi's sarcoma herpesvirus-G protein-coupled receptor". J. Immunol. 163 (4): 20172022. PMID10438939. [26] Li E, Hristova K (2006). "Role of receptor tyrosine kinase transmembrane domains in cell signaling and human pathologies". Biochemistry 45 (20): 624151. doi:10.1021/bi060609y. PMID16700535. [27] Schlessinger, J. (1988). "Signal transduction by allosteric receptor oligomerization". Trends Biochem Sci 13 (11): 4437. doi:10.1016/0968-0004(88)90219-8. PMID3075366. [28] Roskoski, R, Jr. (2004). "The ErbB/HER receptor protein-tyrosine kinases and cancer". Biochem. Biophys. Res. Commun. 319 (1): 111. doi:10.1016/j.bbrc.2004.04.150. PMID15158434. [29] Wolanin PW, Thomason PA, Stock JB (2002). "Histidine protein kinases: key signal transducers outside the animal kingdom" (http:/ / genomebiology. com/ 2002/ 3/ 10/ reviews/ 3013). Genome Biology 3 (10): reviews3013.13013.8. doi:10.1186/gb-2002-3-10-reviews3013. PMC244915. PMID12372152. . [30] Hehlgans, S. Haase, M. and Cordes, N. (2007). "Signaling via integrins: Implications for cell survival and anticancer strategies". Biochim. Biophys. Acta. 1775 (1): 163180. doi:10.1016/j.bbcan.2006.09.001. PMID17084981. [31] Gilcrease MZ. (2006). "Integrin signaling in epithelial cells". Cancer Lett. 247 (1): 125. doi:10.1016/j.canlet.2006.03.031. PMID16725254. [32] Yamamoto M, Sato S, Hemmi H, Hoshino K, Kaisho T, Sanjo H, Takeuchi O, Sugiyama M, Okabe M, Takeda K, Akira S (2003). "Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway". Science 301 (5633): 6403. doi:10.1126/science.1087262. PMID12855817. [33] Yamamoto M, Sato S, Hemmi H, Uematsu S, Hoshino K, Kaisho T, Takeuchi O, Takeda K, Akira S (2003). "TRAM is specifically involved in the Toll-like receptor 4-mediated MyD88-independent signaling pathway". Nat Immunol 4 (11): 114450. doi:10.1038/ni986. PMID14556004. [34] Yamamoto M, Sato S, Hemmi H, Sanjo H, Uematsu S, Kaisho T, Hoshino K, Takeuchi O, Kobayashi M, Fujita T, Takeda K, Akira S (2002). "Essential role for TIRAP in activation of the signalling cascade shared by TLR2 and TLR4". Nature 420 (6913): 3249. doi:10.1038/nature01182. PMID12447441. [35] Delbridge L, O'Riordan M (2007). "Innate recognition of intracellular bacteria". Curr Opin Immunol 19 (1): 106. doi:10.1016/j.coi.2006.11.005. PMID17126540. [36] Vander "et al" (1998). Human Physiology. McGraw-Hill. pp.160. ISBN0-07-067065-X. [37] Lalli E and Sassone-Corsi P (1994). "Signal transduction and gene regulation: the nuclear response to cAMP". J Biol Chem 269 (26): 1735917362. PMID8021233. [38] Rosen O (1987). "After insulin binds". Science 237 (4821): 14521458. doi:10.1126/science.2442814. PMID2442814. [39] Massague J and Gomis RR (2006). "The logic of TGFbeta signaling". FEBS Lett 580 (12): 28112820. doi:10.1016/j.febslet.2006.04.033. PMID16678165. [40] Orton RJ, Sturm OE, Vyshemirsky V, Calder M, Gilbert DR, Kolch W (Dec 2005). "Computational modelling of the receptor-tyrosine-kinase-activated MAPK pathway". The Biochemical journal 392 (Pt 2): 24961. doi:10.1042/BJ20050908. PMC1316260. PMID16293107. [41] Vogelstein, B.; Kinzler, K. W. (2004). "Cancer genes and the pathways they control". Nature Medicine 10 (8): 789799. doi:10.1038/nm1087. PMID15286780. [42] Alberts B, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular biology of the cell (4th ed.). New York: Garland Science. ISBN0-8153-3218-1.

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External links
Netpath - A curated resource of signal transduction pathways in humans (http://www.netpath.org/) Signal Transduction - The Virtual Library of Biochemistry and Cell Biology (http://www.biochemweb.org/ signaling.shtml) TRANSPATH(R) (http://www.gene-regulation.com/cgi-bin/pub/databases/transpath/search.cgi) - A database about signal transduction pathways Redox Signaling Molecules (http://www.energeticforum.com/health-fitness-nutrition/ 8819-redox-signaling-molecules.html) - A public discussion. Redox Signaling Molecules (http://www.redoxsignalingmoleculesbook.com) - PDf download Science's STKE - Signal Transduction Knowledge Environment (http://stke.sciencemag.org/), from the journal Science, published by AAAS. MeSH Signal+Transduction (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=&term=Signal+ Transduction) UCSD-Nature Signaling Gateway (http://www.signaling-gateway.org/), from Nature Publishing Group LitInspector (http://www.litinspector.org) - Signal transduction pathway mining in PubMed abstracts Huaxian Chen, et al. A Cell Based Immunocytochemical Assay For Monitoring Kinase Signaling Pathways And Drug Efficacy (PDF) (http://www.licor.com./bio/PDF/AppNote_AnalBiochem.pdf) Analytical Biochemistry 338 (2005) 136-142 www.Redoxsignaling.com (http://www.redoxsignaling.com) Signaling PAthway Database (http://www.grt.kyushu-u.ac.jp/spad/) - Kyushu University Cell cycle - Homo sapiens (human) (http://www.genome.jp/kegg/pathway/hsa/hsa04110.html) - KEGG PATHWAY (http://www.genome.jp/kegg/pathway.html) Pathway Interaction Database (http://pid.nci.nih.gov/) - NCI

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Diabetes mellitus
Diabetes mellitus
Classification and external resources

Universal blue circle symbol for diabetes. ICD-10 ICD-9 MedlinePlus eMedicine MeSH E10 250 [2] [4] [5] [6] E14 [3]

[1]

001214

med/546

emerg/134 [8]

[7]

C18.452.394.750

Diabetes mellitus, often simply referred to as diabetes, is a group of metabolic diseases in which a person has high blood sugar, either because the body does not produce enough insulin, or because cells do not respond to the insulin that is produced. This high blood sugar produces the classical symptoms of polyuria (frequent urination), polydipsia (increased thirst) and polyphagia (increased hunger). There are three main types of diabetes: Type1 diabetes: results from the body's failure to produce insulin, and presently requires the person to inject insulin. (Also referred to as insulin-dependent diabetes mellitus, IDDM for short, and juvenile diabetes.) Type2 diabetes: results from insulin resistance, a condition in which cells fail to use insulin properly, sometimes combined with an absolute insulin deficiency. (Formerly referred to as non-insulin-dependent diabetes mellitus, NIDDM for short, and adult-onset diabetes.) Gestational diabetes: is when pregnant women, who have never had diabetes before, have a high blood glucose level during pregnancy. It may precede development of type2 DM. Other forms of diabetes mellitus include congenital diabetes, which is due to genetic defects of insulin secretion, cystic fibrosis-related diabetes, steroid diabetes induced by high doses of glucocorticoids, and several forms of monogenic diabetes. All forms of diabetes have been treatable since insulin became available in 1921, and type 2 diabetes may be controlled with medications. Both type 1 and 2 are chronic conditions that usually cannot be cured. Pancreas transplants have been tried with limited success in type1 DM; gastric bypass surgery has been successful in many

Diabetes mellitus with morbid obesity and type2 DM. Gestational diabetes usually resolves after delivery. Diabetes without proper treatments can cause many complications. Acute complications include hypoglycemia, diabetic ketoacidosis, or nonketotic hyperosmolar coma. Serious long-term complications include cardiovascular disease, chronic renal failure, retinal damage. Adequate treatment of diabetes is thus important, as well as blood pressure control and lifestyle factors such as smoking cessation and maintaining a healthy body weight. As of 2000 at least 171 million people worldwide have diabetes, or 2.8% of the population.[9] Type 2 diabetes is by far the most common, affecting 90 to 95% of the U.S. diabetes population.[10]

336

Classification
Most cases of diabetes mellitus fall into three broad categories: type1, type2, and gestational diabetes. A few other types are described. The term diabetes, without qualification, usually refers to diabetes mellitus. The rare disease diabetes insipidus has similar symptoms as diabetes mellitus, but without disturbances in the sugar metabolism (insipidus meaning "without taste" in Latin).
Comparison of type 1 and 2 diabetes Feature Onset Age at onset Type 1 diabetes Sudden [11] Type 2 diabetes Gradual [11]

Any age [11] (mostly young) Thin [11] or normal [11] [11] [12]

Mostly in adults

Body habitus Ketoacidosis Autoantibodies Endogenous insulin

Often obese [11] Rare Absent [11]

[11]

Common

Usually present Low or absent

[11]

Normal, decreased [11] or increased 90% [11]

Concordance in identical twins Prevalence

50%

[11]

Less prevalent

More prevalent - 90 to 95% of [10] U.S. diabetics

The term "type1 diabetes" has replaced several former terms, including childhood-onset diabetes, juvenile diabetes, and insulin-dependent diabetes mellitus (IDDM). Likewise, the term "type2 diabetes" has replaced several former terms, including adult-onset diabetes, obesity-related diabetes, and non-insulin-dependent diabetes mellitus (NIDDM). Beyond these two types, there is no agreed-upon standard nomenclature. Various sources have defined "type3 diabetes" as: gestational diabetes,[13] insulin-resistant type1 diabetes (or "double diabetes"), type2 diabetes which has progressed to require injected insulin, and latent autoimmune diabetes of adults (or LADA or "type1.5" diabetes).[14]

Type 1 diabetes
Type 1 diabetes mellitus is characterized by loss of the insulin-producing beta cells of the islets of Langerhans in the pancreas leading to insulin deficiency. This type of diabetes can be further classified as immune-mediated or idiopathic. The majority of type1 diabetes is of the immune-mediated nature, where beta cell loss is a T-cell mediated autoimmune attack.[15] There is no known preventive measure against type1 diabetes, which causes approximately 10% of diabetes mellitus cases in North America and Europe. Most affected people are otherwise

Diabetes mellitus healthy and of a healthy weight when onset occurs. Sensitivity and responsiveness to insulin are usually normal, especially in the early stages. Type1 diabetes can affect children or adults but was traditionally termed "juvenile diabetes" because it represents a majority of the diabetes cases in children. "Brittle" diabetes, also known as unstable diabetes or labile diabetes, is a term that was traditionally used to describe to dramatic and recurrent swings in glucose levels, often occurring for no apparent reason in insulin-dependent diabetes. This term, however, has no biologic basis and should not be used.[16] There are many different reasons for type 1 diabetes to be accompanied by irregular and unpredictable hyperglycemias, frequently with ketosis, and sometimes serious hypoglycemias, including an impaired counterregulatory response to hypoglycemia, occult infection, gastroparesis (which leads to erratic absorption of dietary carbohydrates), and endocrinopathies (eg, Addison's disease).[17] These phenomena are believed to occur no more frequently than in 1% to 2% of persons with type 1 diabetes.[18]

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Type 2 diabetes
Type2 diabetes mellitus is characterized by insulin resistance which may be combined with relatively reduced insulin secretion. The defective responsiveness of body tissues to insulin is believed to involve the insulin receptor. However, the specific defects are not known. Diabetes mellitus due to a known defect are classified separately. Type2 diabetes is the most common type. In the early stage of type2 diabetes, the predominant abnormality is reduced insulin sensitivity. At this stage hyperglycemia can be reversed by a variety of measures and medications that improve insulin sensitivity or reduce glucose production by the liver.

Gestational diabetes
Gestational diabetes mellitus (GDM) resembles type2 diabetes in several respects, involving a combination of relatively inadequate insulin secretion and responsiveness. It occurs in about 2%5% of all pregnancies and may improve or disappear after delivery. Gestational diabetes is fully treatable but requires careful medical supervision throughout the pregnancy. About 20%50% of affected women develop type2 diabetes later in life. Even though it may be transient, untreated gestational diabetes can damage the health of the fetus or mother. Risks to the baby include macrosomia (high birth weight), congenital cardiac and central nervous system anomalies, and skeletal muscle malformations. Increased fetal insulin may inhibit fetal surfactant production and cause respiratory distress syndrome. Hyperbilirubinemia may result from red blood cell destruction. In severe cases, perinatal death may occur, most commonly as a result of poor placental perfusion due to vascular impairment. Labor induction may be indicated with decreased placental function. A cesarean section may be performed if there is marked fetal distress or an increased risk of injury associated with macrosomia, such as shoulder dystocia. A 2008 study completed in the U.S. found that the number of American women entering pregnancy with preexisting diabetes is increasing. In fact the rate of diabetes in expectant mothers has more than doubled in the past 6 years.[19] This is particularly problematic as diabetes raises the risk of complications during pregnancy, as well as increasing the potential that the children of diabetic mothers will also become diabetic in the future.

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Other types
Pre-diabetes indicates a condition that occurs when a person's blood glucose levels are higher than normal but not high enough for a diagnosis of type2 diabetes. Many people destined to develop type 2 diabetes spend many years in a state of pre-diabetes which has been termed "America's largest healthcare epidemic."[20] :1011 Latent autoimmune diabetes of adults is a condition in which Type 1 diabetes develops in adults. Adults with LADA are frequently initially misdiagnosed as having Type 2 diabetes, based on age rather than etiology. Some cases of diabetes are caused by the body's tissue receptors not responding to insulin (even when insulin levels are normal, which is what separates it from type2 diabetes); this form is very uncommon. Genetic mutations (autosomal or mitochondrial) can lead to defects in beta cell function. Abnormal insulin action may also have been genetically determined in some cases. Any disease that causes extensive damage to the pancreas may lead to diabetes (for example, chronic pancreatitis and cystic fibrosis). Diseases associated with excessive secretion of insulin-antagonistic hormones can cause diabetes (which is typically resolved once the hormone excess is removed). Many drugs impair insulin secretion and some toxins damage pancreatic beta cells. The ICD-10 (1992) diagnostic entity, malnutrition-related diabetes mellitus (MRDM or MMDM, ICD-10 code E12), was deprecated by the World Health Organization when the current taxonomy was introduced in 1999.[21]

Signs and symptoms

Hyperglycemia and osmosis
The classical symptoms of diabetes are polyuria (frequent urination), polydipsia (increased thirst) and polyphagia (increased hunger).[22] Symptoms may develop rapidly (weeks or months) in type1 diabetes while in type2 diabetes they usually develop much more slowly and may be subtle or absent. Prolonged high blood glucose causes glucose absorption, which leads to changes in the shape of the lenses of the eyes, resulting in vision changes; sustained sensible glucose control usually returns the lens to its original shape. Blurred vision is a common complaint leading to a diabetes diagnosis; type1 should always be suspected in cases of rapid vision change, whereas with type2 change is generally more gradual, but should still be suspected .

Overview of the most significant symptoms of diabetes.

Diabetic emergencies
People (usually with type1 diabetes) may also present with diabetic ketoacidosis, a state of metabolic dysregulation characterized by the smell of acetone; a rapid, deep breathing known as Kussmaul breathing; nausea; vomiting and abdominal pain; and altered states of consciousness. A rarer but equally severe possibility is hyperosmolar nonketotic state, which is more common in type2 diabetes and is mainly the result of dehydration. Often, the patient has been drinking extreme amounts of sugar-containing drinks, leading to a vicious circle in regard to the water loss.

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Complications
All forms of diabetes increase the risk of long-term complications. These typically develop after many years (1020), but may be the first symptom in those who have otherwise not received a diagnosis before that time. The major long-term complications relate to damage to blood vessels. Diabetes doubles the risk of cardiovascular disease.[23] The main "macrovascular" diseases (related to atherosclerosis of larger arteries) are ischemic heart disease (angina and myocardial infarction), stroke and peripheral vascular disease. Diabetes also causes "microvascular" complicationsdamage to the small blood vessels.[24] Diabetic retinopathy, which affects blood vessel formation in the retina of the eye, can lead to visual symptoms, reduced vision, and potentially blindness. Diabetic nephropathy, the impact of diabetes on the kidneys, can lead to scarring changes in the kidney tissue, loss of small or progressively larger amounts of protein in the urine, and eventually chronic kidney disease requiring dialysis. Diabetic neuropathy is the impact of diabetes on the nervous system, most commonly causing numbness, tingling and pain in the feet and also increasing the risk of skin damage due to altered sensation. Together with vascular disease in the legs, neuropathy contributes to the risk of diabetes-related foot problems (such as diabetic foot ulcers) that can be difficult to treat and occasionally require amputation.

Other problems
A number of skin rashes can occur in diabetes that are collectively known as diabetic dermadromes.

Causes
The cause of diabetes depends on the type. Type1 diabetes is partly inherited and then triggered by certain infections, with some evidence pointing at Coxsackie B4 virus. There is a genetic element in individual susceptibility to some of these triggers which has been traced to particular HLA genotypes (i.e., the genetic "self" identifiers relied upon by the immune system). However, even in those who have inherited the susceptibility, type1 diabetes mellitus seems to require an environmental trigger. Type2 diabetes is due primarily to lifestyle factors and genetics.[25] Following is a comprehensive list of other causes of diabetes:[26]
Genetic defects of -cell Function Maturity onset diabetes of the young (MODY) Mitochondrial DNA mutations Genetic defects in insulin processing or insulin action Defects in proinsulin conversion Insulin gene mutations Insulin receptor mutations Exocrine Pancreatic Defects Chronic pancreatitis Pancreatectomy Pancreatic neoplasia Cystic fibrosis Hemochromatosis Fibrocalculous pancreatopathy Endocrinopathies Growth hormone excess (acromegaly) Cushing syndrome Hyperthyroidism Pheochromocytoma Glucagonoma Infections Cytomegalovirus infection Coxsackievirus B Drugs Glucocorticoids Thyroid hormone -adrenergic agonists

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Pathophysiology
Insulin is the principal hormone that regulates uptake of glucose from the blood into most cells (primarily muscle and fat cells, but not central nervous system cells). Therefore deficiency of insulin or the insensitivity of its receptors plays a central role in all forms of diabetes mellitus. Humans are capable of digesting some carbohydrates, in particular those most common in food; starch, and some disaccharides such as sucrose, are converted within a few hours to simpler forms most notably the monosaccharide glucose, the principal carbohydrate energy source used by the body. The rest are passed on for processing by gut flora largely in the colon. Insulin is released into the blood by beta cells (-cells), found in the Islets of Langerhans in the pancreas, in response to rising levels of blood glucose, typically after eating. Insulin is used by about two-thirds of the body's cells to absorb glucose from the blood for use as fuel, for conversion to other needed molecules, or for storage. Insulin is also the principal control signal for conversion of glucose to glycogen for internal storage in liver and muscle cells. Lowered glucose levels result both in the reduced release of insulin from the beta cells and in the reverse conversion of glycogen to glucose when glucose levels fall. This is mainly controlled by the hormone glucagon which acts in the opposite manner to insulin. Glucose thus forcibly produced from internal liver cell stores (as glycogen) re-enters the bloodstream; muscle cells lack the necessary export mechanism. Normally liver cells do this when the level of insulin is low (which normally correlates with low levels of blood glucose).

The fluctuation of blood sugar (red) and the sugar-lowering hormone insulin (blue) in humans during the course of a day with three meals. One of the effects of a sugar-rich vs a starch-rich meal is highlighted.

Mechanism of insulin release in normal pancreatic beta cells. Insulin production is more or less constant within the beta cells. Its release is triggered by food, chiefly food containing absorbable glucose.

Higher insulin levels increase some anabolic ("building up") processes such as cell growth and duplication, protein synthesis, and fat storage. Insulin (or its lack) is the principal signal in converting many of the bidirectional processes of metabolism from a catabolic to an anabolic direction, and vice versa. In particular, a low insulin level is the trigger for entering or leaving ketosis (the fat burning metabolic phase). If the amount of insulin available is insufficient, if cells respond poorly to the effects of insulin (insulin insensitivity or resistance), or if the insulin itself is defective, then glucose will not have its usual effect so that glucose will not be absorbed properly by those body cells that require it nor will it be stored appropriately in the liver and muscles. The net effect is persistent high levels of blood glucose, poor protein synthesis, and other metabolic derangements, such as acidosis. When the glucose concentration in the blood is raised beyond its renal threshold (about 10mmol/L, although this may be altered in certain conditions, such as pregnancy), reabsorption of glucose in the proximal renal tubuli is incomplete, and part of the glucose remains in the urine (glycosuria). This increases the osmotic pressure of the urine and inhibits reabsorption of water by the kidney, resulting in increased urine production (polyuria) and increased fluid loss. Lost blood volume will be replaced osmotically from water held in body cells and other body compartments, causing dehydration and increased thirst.

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Diagnosis
2006 WHO Diabetes criteria[27]
Condition 2 hour glucose mmol/l(mg/dl) Normal Impaired fasting glycaemia Impaired glucose tolerance Diabetes mellitus <7.8 (<140) <7.8 (<140) 7.8 (140) 11.1 (200) Fasting glucose mmol/l(mg/dl) <6.1 (<110) 6.1(110) & <7.0(<126) <7.0 (<126) 7.0 (126)

Diabetes mellitus is characterized by recurrent or persistent hyperglycemia, and is diagnosed by demonstrating any one of the following:[21] Fasting plasma glucose level 7.0mmol/L (126mg/dL). Plasma glucose 11.1mmol/L (200mg/dL) two hours after a 75g oral glucose load as in a glucose tolerance test. Symptoms of hyperglycemia and casual plasma glucose 11.1mmol/L (200mg/dL). Glycated hemoglobin (Hb A1C) 6.5%.[28] A positive result, in the absence of unequivocal hyperglycemia, should be confirmed by a repeat of any of the above-listed methods on a different day. It is preferable to measure a fasting glucose level because of the ease of measurement and the considerable time commitment of formal glucose tolerance testing, which takes two hours to complete and offers no prognostic advantage over the fasting test.[29] According to the current definition, two fasting glucose measurements above 126mg/dL (7.0mmol/L) is considered diagnostic for diabetes mellitus. People with fasting glucose levels from 100 to 125mg/dL (5.6 to 6.9mmol/L) are considered to have impaired fasting glucose. Patients with plasma glucose at or above 140mg/dL (7.8mmol/L), but not over 200mg/dL (11.1mmol/L), two hours after a 75 g oral glucose load are considered to have impaired glucose tolerance. Of these two pre-diabetic states, the latter in particular is a major risk factor for progression to full-blown diabetes mellitus as well as cardiovascular disease.[30] Glycated hemoglobin is better than fasting glucose for determining risks of cardiovascular disease and death from any cause.[31]

Management
Diabetes mellitus is a chronic disease which cannot be cured except in very specific situations. Management concentrates on keeping blood sugar levels as close to normal ("euglycemia") as possible, without causing hypoglycemia. This can usually be accomplished with diet, exercise, and use of appropriate medications (insulin in the case of type 1 diabetes, oral medications as well as possibly insulin in type 2 diabetes). Patient education, understanding, and participation is vital since the complications of diabetes are far less common and less severe in people who have well-managed blood sugar levels.[32] [33] The goal of treatment is an HbA1C level of 6.5%, but should not be lower than that, and may be set higher.[34] Attention is also paid to other health problems that may accelerate the deleterious effects of diabetes. These include smoking, elevated cholesterol levels, obesity, high blood pressure, and lack of regular exercise.[34]

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Lifestyle
There are roles for patient education, dietetic support, sensible exercise, with the goal of keeping both short-term and long-term blood glucose levels within acceptable bounds. In addition, given the associated higher risks of cardiovascular disease, lifestyle modifications are recommended to control blood pressure.[35]

Medications
Oral medications Metformin is generally recommended as a first line treatment for type 2 diabetes as there is good evidence that it decreases mortality.[36] Routine use of aspirin however has not been found to improve outcomes in uncomplicated diabetes.[37] Insulin Type1 diabetes is typically treated with a combinations of regular and NPH insulin, or synthetic insulin analogs. When insulin is used in type2 diabetes, a long-acting formulation is usually added initially, while continuing oral medications.[36] Doses of insulin are then increased to effect.[36]

Support
In countries using a general practitioner system, such as the United Kingdom, care may take place mainly outside hospitals, with hospital-based specialist care used only in case of complications, difficult blood sugar control, or research projects. In other circumstances, general practitioners and specialists share care of a patient in a team approach. Optometrists, podiatrists/chiropodists, dietitians, physiotherapists, nursing specialists (e.g., DSNs (Diabetic Specialist Nurse)), nurse practitioners, or certified diabetes educators, may jointly provide multidisciplinary expertise.

Epidemiology
In 2000, according to the World Health Organization, at least 171 million people Prevalence of diabetes worldwide in 2000 (per 1000 inhabitants). World average was 2.8%. no worldwide suffer data7.57.5151522.522.5303037.537.5454552.552.5606067.567.5757582.582.5 from diabetes, or 2.8% of the population.[9] Its incidence is increasing rapidly, and it is estimated that by Disability-adjusted life year for diabetes mellitus per 100,000inhabitants in 2004. no 2030, this data<100100-200200-300300-400400-500500-600600-700700-800800-900900-10001000-1500>1500 number will almost double.[9] Diabetes mellitus occurs throughout the world, but is more common (especially type2) in the more developed countries. The greatest increase in prevalence is, however, expected to occur in Asia and Africa, where most patients

Diabetes mellitus will probably be found by 2030.[9] The increase in incidence of diabetes in developing countries follows the trend of urbanization and lifestyle changes, perhaps most importantly a "Western-style" diet. This has suggested an environmental (i.e., dietary) effect, but there is little understanding of the mechanism(s) at present, though there is much speculation, some of it most compellingly presented.[9] Prevalence in the United States For at least 20 years, diabetes rates in North America have been increasing substantially. In 2010 nearly 26 million people have diabetes in the United States alone, from those 7 million people remain undiagnosed. Another 57 million people are estimated to have pre-diabetes.[38] The Centers for Disease Control has termed the change an epidemic.[39] The National Diabetes Information Clearinghouse estimates that diabetes costs $132 billion in the United States alone every year. About 5%10% of diabetes cases in North America are type1, with the rest being type2. The fraction of type1 in other parts of the world differs. Most of this difference is not currently understood. The American Diabetes Association cite the 2003 assessment of the National Center for Chronic Disease Prevention and Health Promotion (Centers for Disease Control and Prevention) that 1 in 3 Americans born after 2000 will develop diabetes in their lifetime.[40] [41] According to the American Diabetes Association, approximately 18.3% (8.6 million) of Americans age 60 and older have diabetes.[42] Diabetes mellitus prevalence increases with age, and the numbers of older persons with diabetes are expected to grow as the elderly population increases in number. The National Health and Nutrition Examination Survey (NHANES III) demonstrated that, in the population over 65 years old, 18% to 20% have diabetes, with 40% having either diabetes or its precursor form of impaired glucose tolerance.[43] Prevalence in Australia Indigenous populations in first world countries have a higher prevalence and increasing incidence of diabetes than their corresponding non-indigenous populations. In Australia the age-standardised prevalence of self-reported diabetes in Indigenous Australians is almost 4 times that of non-indigenous Australians.[44] Preventative community health programs such as Sugar Man (diabetes education) are showing some success in tackling this problem.

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Etymology
The word diabetes ( /da.bitiz/ or /da.bits/) comes from Latin diabts, which in turn comes from Ancient Greek (diabts) which literally means a passer through; a siphon.[45] Ancient Greek physician Aretaeus of Cappadocia (fl. 1st century CE) used that word, with the intended meaning excessive discharge of urine, as the name for the disease.[46] [47] Ultimately, the word comes from Greek (diabainein), meaning to pass through,[45] which is composed of - (dia-), meaning through and (bainein), meaning to go.[46] The word diabetes is first recorded in English, in the form diabete, in a medical text written around 1425. The word mellitus (/mlats/ or /mlts/) comes from the classical Latin word melltus, meaning mellite[48] (i.e. sweetened with honey;[48] honey-sweet[49] ). The Latin word comes from mell-, which comes from mel, meaning honey;[48] [49] sweetness;[49] pleasant thing,[49] and the suffix -tus,[48] whose meaning is the same as that of the English suffix -ite.[50] It was Thomas Willis who in 1675 added mellitus to the word diabetes as a designation for the disease, when he noticed that the urine of a diabetic had a sweet taste (glycosuria).[47] This sweet taste had been noticed in urine by the ancient Greeks, Chinese, Egyptians, Indians, and Persians.

History
Diabetes is one of the oldest known diseases.[47] An Egyptian manuscript from c. 1550 BCE mentions the phrase the passing of too much urine.[47] The great Indian physician Sushruta (fl. 6th century BCE)[47] identified the disease and classified it as Medhumeha.[51] He further identified it with obesity and sedentary lifestyle, advising exercises to help "cure" it.[51] The ancient Indians tested for diabetes by observing whether ants were attracted to a person's urine, and called the ailment "sweet urine disease" (Madhumeha).

Diabetes mellitus Concerning the sweetness of urine, it is to be noted that the Chinese, Japanese and Korean words for diabetes are based on the same ideographs () which mean "sugar urine disease". It was in 1776 that Matthew Dobson confirmed that the sweet taste comes from an excess of a kind of sugar in the urine and blood.[52] The first complete clinical description of diabetes was given by the Ancient Greek physician Aretaeus of Cappadocia (fl. 1st century CE), who noted the excessive amount of urine which passed through the kidneys and gave the disease the name diabetes.[47] Diabetes mellitus appears to have been a death sentence in the ancient era. Hippocrates makes no mention of it, which may indicate that he felt the disease was incurable. Aretaeus did attempt to treat it but could not give a good prognosis; he commented that "life (with diabetes) is short, disgusting and painful."[53] In medieval Persia, Avicenna (9801037) provided a detailed account on diabetes mellitus in The Canon of Medicine, "describing the abnormal appetite and the collapse of sexual functions," and he documented the sweet taste of diabetic urine. Like Aretaeus before him, Avicenna recognized a primary and secondary diabetes. He also described diabetic gangrene, and treated diabetes using a mixture of lupine, trigonella (fenugreek), and zedoary seed, which produces a considerable reduction in the excretion of sugar, a treatment which is still prescribed in modern times. Avicenna also "described diabetes insipidus very precisely for the first time", though it was later Johann Peter Frank (17451821) who first differentiated between diabetes mellitus and diabetes insipidus.[54] Although diabetes has been recognized since antiquity, and treatments of various efficacy have been known in various regions since the Middle Ages, and in legend for much longer, pathogenesis of diabetes has only been understood experimentally since about 1900.[55] The discovery of a role for the pancreas in diabetes is generally ascribed to Joseph von Mering and Oskar Minkowski, who in 1889 found that dogs whose pancreas was removed developed all the signs and symptoms of diabetes and died shortly afterwards.[56] In 1910, Sir Edward Albert Sharpey-Schafer suggested that people with diabetes were deficient in a single chemical that was normally produced by the pancreashe proposed calling this substance insulin, from the Latin insula, meaning island, in reference to the insulin-producing islets of Langerhans in the pancreas. The endocrine role of the pancreas in metabolism, and indeed the existence of insulin, was not further clarified until 1921, when Sir Frederick Grant Banting and Charles Herbert Best repeated the work of Von Mering and Minkowski, and went further to demonstrate they could reverse induced diabetes in dogs by giving them an extract from the pancreatic islets of Langerhans of healthy dogs.[57] Banting, Best, and colleagues (especially the chemist Collip) went on to purify the hormone insulin from bovine pancreases at the University of Toronto. This led to the availability of an effective treatmentinsulin injectionsand the first patient was treated in 1922. For this, Banting and laboratory director MacLeod received the Nobel Prize in Physiology or Medicine in 1923; both shared their Prize money with others in the team who were not recognized, in particular Best and Collip. Banting and Best made the patent available without charge and did not attempt to control commercial production. Insulin production and therapy rapidly spread around the world, largely as a result of this decision. Banting is honored by World Diabetes Day which is held on his birthday, November 14. The distinction between what is now known as type1 diabetes and type2 diabetes was first clearly made by Sir Harold Percival (Harry) Himsworth, and published in January 1936.[58] Despite the availability of treatment, diabetes has remained a major cause of death. For instance, statistics reveal that the cause-specific mortality rate during 1927 amounted to about 47.7 per 100,000 population in Malta.[59] Other landmark discoveries include:[55] Identification of the first of the sulfonylureas in 1942 Reintroduction of the use of biguanides for Type2 diabetes in the late 1950s. The initial phenformin was withdrawn worldwide (in the U.S. in 1977) due to its potential for sometimes fatal lactic acidosis and metformin was first marketed in France in 1979, but not until 1994 in the US.

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Diabetes mellitus The determination of the amino acid sequence of insulin (by Sir Frederick Sanger, for which he received a Nobel Prize) The radioimmunoassay for insulin, as discovered by Rosalyn Yalow and Solomon Berson (gaining Yalow the 1977 Nobel Prize in Physiology or Medicine)[60] The three-dimensional structure of insulin (PDB 2INS [61]) Dr Gerald Reaven's identification of the constellation of symptoms now called metabolic syndrome in 1988 Demonstration that intensive glycemic control in type1 diabetes reduces chronic side effects more as glucose levels approach 'normal' in a large longitudinal study,[62] and also in type2 diabetics in other large studies Identification of the first thiazolidinedione as an effective insulin sensitizer during the 1990s In 1980, U.S. biotech company Genentech developed biosynthetic human insulin. The insulin was isolated from genetically altered bacteria (the bacteria contain the human gene for synthesizing synthetic human insulin), which produce large quantities of insulin. The purified insulin is distributed to pharmacies for use by diabetes patients. Initially, this development was not regarded by the medical profession as a clinically meaningful development. However, by 1996, the advent of insulin analogues which had vastly improved absorption, distribution, metabolism, and excretion (ADME) characteristics which were clinically meaningful based on this early biotechnology development.

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Society and culture

The 1990 "St. Vincent Declaration"[63] [64] was the result of international efforts to improve the care accorded to those with diabetes. Doing so is important both in terms of quality of life and life expectancy but also economicallyexpenses due to diabetes have been shown to be a major drain on health-and productivity-related resources for healthcare systems and governments. Several countries established more and less successful national diabetes programmes to improve treatment of the disease.[65] A study shows that diabetic patients with neuropathic symptoms such as numbness or tingling in feet or hands are twice as likely to be unemployed as those without the symptoms.[66]

In other animals
In animals, diabetes is most commonly encountered in dogs and cats. Middle-aged animals are most commonly affected. Female dogs are twice as likely to be affected as males, while according to some sources male cats are also more prone than females. In both species, all breeds may be affected, but some small dog breeds are particularly likely to develop diabetes, such as Miniature Poodles.[67] The symptoms may relate to fluid loss and polyuria, but the course may also be insidious. Diabetic animals are more prone to infections. The long-term complications recognised in humans are much rarer in animals. The principles of treatment (weight loss, oral antidiabetics, subcutaneous insulin) and management of emergencies (e.g. ketoacidosis) are similar to those in humans.[67]

Diabetes mellitus

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References
[1] [2] [3] [4] [5] [6] [7] [8] [9] "Diabetes Blue Circle Symbol" (http:/ / www. diabetesbluecircle. org). International Diabetes Federation. 17 March 2006. . http:/ / apps. who. int/ classifications/ icd10/ browse/ 2010/ en#/ E10 http:/ / apps. who. int/ classifications/ icd10/ browse/ 2010/ en#/ E14 http:/ / www. icd9data. com/ getICD9Code. ashx?icd9=250 http:/ / www. nlm. nih. gov/ medlineplus/ ency/ article/ 001214. htm http:/ / www. emedicine. com/ med/ topic546. htm http:/ / www. emedicine. com/ emerg/ topic134. htm# http:/ / www. nlm. nih. gov/ cgi/ mesh/ 2011/ MB_cgi?mode=& term=Diabetes& field=entry#TreeC18. 452. 394. 750 Wild S, Roglic G, Green A, Sicree R, King H (May 2004). "Global prevalence of diabetes: estimates for 2000 and projections for 2030". Diabetes Care 27 (5): 104753. doi:10.2337/diacare.27.5.1047. PMID15111519. [10] "Type 2 Diabetes Overview" (http:/ / diabetes. webmd. com/ guide/ type-2-diabetes). Web MD. . [11] Elizabeth D Agabegi; Agabegi, Steven S. (2008). Step-Up to Medicine (Step-Up Series). Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-7153-6. [12] Lambert, P. (2002). "What is Type 1 Diabetes?". Medicine 30: 15. doi:10.1383/medc.30.1.1.28264. [13] "Other "types" of diabetes" (http:/ / www. diabetes. org/ other-types. jsp). American Diabetes Association. August 25, 2005. . [14] "Diseases: Johns Hopkins Autoimmune Disease Research Center" (http:/ / autoimmune. pathology. jhmi. edu/ diseases. cfm?systemID=3& DiseaseID=23). . Retrieved 2007-09-23. [15] Rother KI (April 2007). "Diabetes treatmentbridging the divide". The New England Journal of Medicine 356 (15): 1499501. doi:10.1056/NEJMp078030. PMID17429082. [16] "Diabetes Mellitus (DM): Diabetes Mellitus and Disorders of Carbohydrate Metabolism: Merck Manual Professional" (http:/ / www. merck. com/ mmpe/ sec12/ ch158/ ch158b. html#sec12-ch158-ch158b-1206). Merck.com. . Retrieved 2010-07-30. [17] "Diabetes Mellitus (DM): Diabetes Mellitus and Disorders of Carbohydrate Metabolism: Merck Manual Professional" (http:/ / www. merck. com/ mmpe/ sec12/ ch158/ ch158b. html#sec12-ch158-ch158b-1206). Merck.com. . Retrieved 2010-07-30. [18] Dorner M, Pinget M, Brogard JM (May 1977). "Essential labile diabetes" (in German). MMW Munch Med Wochenschr 119 (19): 6714. PMID406527. [19] Lawrence JM, Contreras R, Chen W, Sacks DA (May 2008). "Trends in the prevalence of preexisting diabetes and gestational diabetes mellitus among a racially/ethnically diverse population of pregnant women, 19992005". Diabetes Care 31 (5): 899904. doi:10.2337/dc07-2345. PMID18223030. [20] Handelsman Yehuda, MD. "A Doctor's Diagnosis: Prediabetes". Power of Prevention 1 (2): 2009. [21] World Health Organisation Department of Noncommunicable Disease Surveillance (1999). "Definition, Diagnosis and Classification of Diabetes Mellitus and its Complications" (http:/ / whqlibdoc. who. int/ hq/ 1999/ WHO_NCD_NCS_99. 2. pdf) (PDF). . [22] Cooke DW, Plotnick L (November 2008). "Type 1 diabetes mellitus in pediatrics". Pediatr Rev 29 (11): 37484; quiz 385. doi:10.1542/pir.29-11-374. PMID18977856. [23] "Diabetes mellitus, fasting blood glucose concentration, and risk of vascular disease: a collaborative meta-analysis of 102 prospective studies : The Lancet" (http:/ / www. thelancet. com/ journals/ lancet/ article/ PIIS0140-6736(10)60484-9/ fulltext). . [24] Boussageon R, Bejan-Angoulvant T, Saadatian-Elahi M, et al. (2011). "Effect of intensive glucose lowering treatment on all cause mortality, cardiovascular death, and microvascular events in type 2 diabetes: meta-analysis of randomised controlled trials". BMJ 343: d4169. doi:10.1136/bmj.d4169. PMC3144314. PMID21791495. [25] Risrus U, Willett WC, Hu FB (January 2009). "Dietary fats and prevention of type 2 diabetes". Progress in Lipid Research 48 (1): 4451. doi:10.1016/j.plipres.2008.10.002. PMC2654180. PMID19032965. [26] Unless otherwise specified, reference is: Table 20-5 in Mitchell, Richard Sheppard; Kumar, Vinay; Abbas, Abul K.; Fausto, Nelson. Robbins Basic Pathology. Philadelphia: Saunders. ISBN1-4160-2973-7. 8th edition. [27] "Definition and Diagnosis of Diabetes Mellitus and Intermediate Hyperglycemia" (http:/ / www. who. int/ diabetes/ publications/ Definition and diagnosis of diabetes_new. pdf) (pdf). World Health Organization. www.who.int. 2006. . Retrieved 2011-02-20. [28] ""Diabetes Care" January 2010" (http:/ / care. diabetesjournals. org/ content/ 33/ Supplement_1/ S3. full). American Diabetes Association. . Retrieved 2010-01-29. [29] Saydah SH, Miret M, Sung J, Varas C, Gause D, Brancati FL (August 2001). "Postchallenge hyperglycemia and mortality in a national sample of U.S. adults". Diabetes Care 24 (8): 1397402. doi:10.2337/diacare.24.8.1397. PMID11473076. [30] Santaguida PL, Balion C, Hunt D, Morrison K, Gerstein H, Raina P, Booker L, Yazdi H. "Diagnosis, Prognosis, and Treatment of Impaired Glucose Tolerance and Impaired Fasting Glucose" (http:/ / www. ahrq. gov/ clinic/ epcsums/ impglusum. htm). Summary of Evidence Report/Technology Assessment, No. 128. Agency for Healthcare Research and Quality. . Retrieved 2008-07-20. [31] Selvin E, Steffes MW, Zhu H et al (2010). "Glycated hemoglobin, diabetes, and cardiovascular risk in nondiabetic adults". N. Engl. J. Med. 362 (9): 80011. doi:10.1056/NEJMoa0908359. PMC2872990. PMID20200384. [32] Nathan DM, Cleary PA, Backlund JY, et al. (December 2005). "Intensive diabetes treatment and cardiovascular disease in patients with type 1 diabetes". The New England Journal of Medicine 353 (25): 264353. doi:10.1056/NEJMoa052187. PMC2637991. PMID16371630. [33] "The effect of intensive diabetes therapy on the development and progression of neuropathy. The Diabetes Control and Complications Trial Research Group" (http:/ / www. annals. org/ cgi/ pmidlookup?view=long& pmid=7887548). Annals of Internal Medicine 122 (8): 5618.

Diabetes mellitus
April 1995. doi:10.1059/0003-4819-122-8-199504150-00001 (inactive 2009-10-31). PMID7887548. . [34] National Institute for Health and Clinical Excellence. Clinical guideline 66: Type 2 diabetes (http:/ / guidance. nice. org. uk/ CG66). London, 2008. [35] Adler AI, Stratton IM, Neil HA, et al. (August 2000). "Association of systolic blood pressure with macrovascular and microvascular complications of type 2 diabetes (UKPDS 36): prospective observational study". BMJ 321 (7258): 4129. doi:10.1136/bmj.321.7258.412. PMC27455. PMID10938049. [36] Ripsin, CM; Kang, H, Urban, RJ (2009-01-01). "Management of blood glucose in type 2 diabetes mellitus". American family physician 79 (1): 2936. PMID19145963. [37] Pignone M, Alberts MJ, Colwell JA, et al. (June 2010). "Aspirin for primary prevention of cardiovascular events in people with diabetes: a position statement of the American Diabetes Association, a scientific statement of the American Heart Association, and an expert consensus document of the American College of Cardiology Foundation". Diabetes Care 33 (6): 1395402. doi:10.2337/dc10-0555. PMC2875463. PMID20508233. [38] CDC.gov (http:/ / www. cdc. gov/ media/ releases/ 2011/ p0126_diabetes. html) [39] "CDC's Diabetes Program-News and Information-Press Releases-October 26, 2000" (http:/ / www. cdc. gov/ Diabetes/ news/ docs/ 010126. htm). . Retrieved 2008-06-23. [40] Narayan KM, Boyle JP, Thompson TJ, Sorensen SW, Williamson DF (October 2003). "Lifetime risk for diabetes mellitus in the United States". JAMA 290 (14): 188490. doi:10.1001/jama.290.14.1884. PMID14532317. [41] American Diabetes Association (2005). "Total Prevalence of Diabetes & Pre-diabetes" (http:/ / web. archive. org/ web/ 20060208032127/ http:/ / www. diabetes. org/ diabetes-statistics/ prevalence. jsp). Archived from the original (http:/ / www. diabetes. org/ diabetes-statistics/ prevalence. jsp) on 2006-02-08. . Retrieved 2006-03-17. [42] "Seniors and Diabetes" (http:/ / www. dlife. com/ dLife/ do/ ShowContent/ daily_living/ seniors/ ). Elderly And Diabetes-Diabetes and Seniors. LifeMed Media. 2006. . Retrieved 2007-05-14. [43] Harris MI, Flegal KM, Cowie CC, et al. (April 1998). "Prevalence of diabetes, impaired fasting glucose, and impaired glucose tolerance in U.S. adults. The Third National Health and Nutrition Examination Survey, 19881994". Diabetes Care 21 (4): 51824. doi:10.2337/diacare.21.4.518. PMID9571335. [44] Australian Institute for Health and Welfare. "Diabetes, an overview" (http:/ / web. archive. org/ web/ 20080617222036/ http:/ / www. aihw. gov. au/ indigenous/ health/ diabetes. cfm). Archived from the original (http:/ / www. aihw. gov. au/ indigenous/ health/ diabetes. cfm) on 2008-06-17. . Retrieved 2008-06-23. [45] Oxford English Dictionary. diabetes. Retrieved 2011-06-10. [46] Harper, Douglas (20012010). "Online Etymology Dictionary. diabetes." (http:/ / www. etymonline. com/ index. php?search=diabetes& searchmode=none). . Retrieved 2011-06-10 [47] Dallas, John (2011). "Royal College of Physicians of Edinburgh. Diabetes, Doctors and Dogs: An exhibition on Diabetes and Endocrinology by the College Library for the 43rd St. Andrew's Day Festival Symposium" (http:/ / www. rcpe. ac. uk/ library/ exhibitions/ diabetes/ ). [48] Oxford English Dictionary. mellite. Retrieved 2011-06-10. [49] "MyEtimology. mellitus." (http:/ / www. myetymology. com/ latin/ mellitus. html). . Retrieved 2011-06-10 [50] Oxford English Dictionary. -ite. Retrieved 2011-06-10. [51] Dwivedi, Girish & Dwivedi, Shridhar (2007). History of Medicine: Sushruta the Clinician Teacher par Excellence (http:/ / medind. nic. in/ iae/ t07/ i4/ iaet07i4p243. pdf). National Informatics Centre (Government of India). [52] Dobson, M. (1776). "Nature of the urine in diabetes". Medical Observations and Inquiries 5: 298310. [53] Medvei, Victor Cornelius (1993). The history of clinical endocrinology. Carnforth, Lancs., U.K: Parthenon Pub. Group. pp.2334. ISBN1-85070-427-9. [54] Nabipour, I. (2003). "Clinical Endocrinology in the Islamic Civilization in Iran". International Journal of Endocrinology and Metabolism 1: 4345 [445]. [55] Patlak M (December 2002). "New weapons to combat an ancient disease: treating diabetes" (http:/ / www. fasebj. org/ content/ 16/ 14/ 1853. 2). The FASEB Journal 16 (14): 1853. PMID12468446. . [56] Von Mehring J, Minkowski O. (1890). "Diabetes mellitus nach pankreasexstirpation". Arch Exp Pathol Pharmakol 26 (56): 371387. doi:10.1007/BF01831214. [57] Banting FG, Best CH, Collip JB, Campbell WR, Fletcher AA (November 1991). "Pancreatic extracts in the treatment of diabetes mellitus: preliminary report. 1922". CMAJ 145 (10): 12816. PMC1335942. PMID1933711. [58] Himsworth (1936). "Diabetes mellitus: its differentiation into insulin-sensitive and insulin-insensitive types". Lancet i (5864): 12730. doi:10.1016/S0140-6736(01)36134-2. [59] Department of Health (Malta), 18971972:Annual Reports. [60] Yalow RS, Berson SA (July 1960). "Immunoassay of endogenous plasma insulin in man". The Journal of Clinical Investigation 39 (7): 115775. doi:10.1172/JCI104130. PMC441860. PMID13846364. [61] http:/ / www. rcsb. org/ pdb/ explore/ explore. do?structureId=2INS [62] The Diabetes Control And Complications Trial Research Group (September 1993). "The effect of intensive treatment of diabetes on the development and progression of long-term complications in insulin-dependent diabetes mellitus. The Diabetes Control and Complications Trial Research Group". The New England Journal of Medicine 329 (14): 97786. doi:10.1056/NEJM199309303291401. PMID8366922.

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Diabetes mellitus
[63] Theodore H. Tulchinsky, Elena A. Varavikova (2008). The New Public Health, Second Edition. New York: Academic Press. p.200. ISBN0-12-370890-7. [64] Piwernetz K, Home PD, Snorgaard O, Antsiferov M, Staehr-Johansen K, Krans M (May 1993). "Monitoring the targets of the St Vincent Declaration and the implementation of quality management in diabetes care: the DIABCARE initiative. The DIABCARE Monitoring Group of the St Vincent Declaration Steering Committee". Diabetic Medicine 10 (4): 3717. doi:10.1111/j.1464-5491.1993.tb00083.x. PMID8508624. [65] Dubois, HFW and Bankauskaite, V (2005). "Type2 diabetes programmes in Europe" (http:/ / www. euro. who. int/ Document/ Obs/ EuroObserver7_3. pdf) (PDF). Euro Observer 7 (2): 56. . [66] Stewart WF, Ricci JA, Chee E, Hirsch AG, Brandenburg NA (June 2007). "Lost productive time and costs due to diabetes and diabetic neuropathic pain in the US workforce". J. Occup. Environ. Med. 49 (6): 6729. doi:10.1097/JOM.0b013e318065b83a. PMID17563611. [67] "Diabetes mellitus" (http:/ / www. merckvetmanual. com/ mvm/ index. jsp?cfile=htm/ bc/ 40302. htm). Merck Veterinary Manual, 9th edition (online version). 2005. . Retrieved 2011-10-23.

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External links
Diabetes (http://www.dmoz.org/Health/Conditions_and_Diseases/Endocrine_Disorders/Pancreas/Diabetes/) at the Open Directory Project American Diabetes Association (http://www.diabetes.org/) IDF Diabetes Atlas (http://www.diabetesatlas.org/) International Diabetes Federation (http://www.idf.org/) National Diabetes Education Program (http://ndep.nih.gov/) Peers for Progress (http://www.peersforprogress.org/) World Diabetes Day (http://www.worlddiabetesday.org/)

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Informational Macromolecules

350

DNA synthesis and repair

DNA replication
DNA replication is a biological process that occurs in all living organisms and copies their DNA; it is the basis for biological inheritance. The process starts with one double-stranded DNA molecule and produces two identical copies of the molecule. Each strand of the original double-stranded DNA molecule serves as template for the production of the complementary strand, a process referred to as semiconservative replication. Cellular proofreading and error toe-checking mechanisms ensure near perfect fidelity for DNA replication.[1] [2] In a cell, DNA replication begins at specific locations in the genome, called "origins".[3] Unwinding of DNA at the origin, and synthesis of new strands, forms a replication fork. In addition to DNA polymerase, the enzyme that synthesizes the new DNA by adding nucleotides matched to the template strand, a number of other proteins are associated with the fork and assist in the initiation and continuation of DNA synthesis. DNA replication can also be performed in vitro (artificially, outside a cell). DNA polymerases, isolated from cells, and artificial DNA primers are used to initiate DNA synthesis at known sequences in a template molecule. The polymerase chain reaction (PCR), a common laboratory technique, employs such artificial synthesis in a cyclic manner to amplify a specific target DNA fragment from a pool of DNA.

DNA replication. The double helix is unwound and each strand acts as a template for the next strand. Bases are matched to synthesize the new partner strands.

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351

DNA structure
DNA usually exists as a double-stranded structure, with both strands coiled together to form the characteristic double-helix. Each single strand of DNA is a chain of four types of nucleotides having the bases: adenine, cytosine, guanine, and thymine. A nucleotide is a mono-, di-, or triphosphate deoxyribonucleoside; that is, a deoxyribose sugar is attached to one, two, or three phosphates. Chemical interaction of these nucleotides forms phosphodiester linkages, creating the phosphate-deoxyribose backbone of the DNA double helix with the bases pointing inward. Nucleotides (bases) are matched between strands through hydrogen bonds to form base pairs. Adenine pairs with thymine, and cytosine pairs with guanine. DNA strands have a directionality, and the different ends of a single strand are called the "3' (three-prime) end" and the "5' (five-prime) end" with the direction of the naming going 5 prime to the 3 prime region.The strands of the helix are anti-parallel with one being 5 prime to 3 then the opposite strand 3 prime to 5 . These terms refer to the carbon atom in deoxyribose to which the next phosphate in the chain attaches. Directionality has consequences in DNA synthesis, because DNA polymerase can synthesize DNA in only one direction by adding nucleotides to the 3' end of a DNA strand. The pairing of bases in DNA through hydrogen bonding means that the information contained within each strand is redundant. The nucleotides on a single strand can be used to reconstruct nucleotides on a newly synthesized partner strand.[4]

The chemical structure of DNA

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352

DNA polymerase
DNA polymerases are a family of enzymes that carry out all forms of DNA replication.[6] However, a DNA polymerase can only extend an existing DNA strand paired with a template strand; it cannot begin the synthesis of a new strand. To begin synthesis, a short fragment of DNA or RNA, called a primer, must be created and paired with the template DNA strand. DNA polymerase then synthesizes a new strand of DNA by extending the 3' end of an existing nucleotide chain, adding new nucleotides matched to the template strand one at a time via the creation of phosphodiester bonds. The energy for this process of DNA polymerization comes from two of the three total phosphates attached to each unincorporated base. (Free bases with their attached phosphate groups are called nucleoside triphosphates.) When a nucleotide is being added to a growing DNA strand, two of the phosphates are removed and the energy produced creates a phosphodiester bond that attaches the remaining phosphate to the growing chain. The energetics of this process also help explain the directionality of synthesis - if DNA were synthesized in the 3' to 5' direction, the energy for the process would come from the 5' end of the growing strand rather than from free nucleotides. In general, DNA polymerases are extremely accurate, making less than one mistake for every 107 nucleotides added.[7] Even so, some DNA polymerases also have proofreading ability; they can remove nucleotides from the end of a strand in order to correct mismatched bases. If the 5' nucleotide needs to be removed during proofreading, the triphosphate end is lost. Hence, the energy source that usually provides energy to add a new nucleotide is also lost.

DNA polymerases adds nucleotides to the 5' end [5] of a strand of DNA. If a mismatch is accidentally incorporated, the polymerase is inhibited from further extension. Proofreading removes the mismatched nucleotide and extension continues.

Replication process
Origins
For a cell to divide, it must first replicate its DNA.[8] This process is initiated at particular points in the DNA, known as "origins", which are targeted by proteins that separate the two strands and initiate DNA synthesis.[3] Origins contain DNA sequences recognized by replication initiator proteins (e.g., dnaA in E. coli' and the Origin Recognition Complex in yeast).[9] These initiator proteins recruit other proteins to separate the two strands and initiate replication forks. Initiator proteins recruit other proteins and form the pre-replication complex, which separate the DNA strands at the origin and forms a bubble. Origins tend to be "AT-rich" (rich in adenine and thymine bases) to assist this process, because A-T base pairs have two hydrogen bonds (rather than the three formed in a C-G pair)in general, strands rich in these nucleotides are easier to separate because a greater number of hydrogen bonds requires more energy to break them.[10] All known DNA replication systems require a free 3' OH group before synthesis can be initiated. Four distinct mechanisms are have been described.

DNA replication 1. All cellular life forms and many DNA viruses, phages and plasmids use a primase to synthesize a short RNA primer with a free 3 OH group which is subsequently elongated by a DNA polymerase. 2. The retroelements (including retroviruses) employ a transfer RNA that primes DNA replication by providing a free 3 OH that is used for elongation by the reverse transcriptase. 3. In the adenoviruses and the 29 family of bacteriophages, the 3' OH group is provided by the side chain of an amino acid of the genome attached protein (the terminal protein) to which nucleotides are added by the DNA polymerase to form a new strand. 4. In the single stranded DNA viruses - a group that includes the circoviruses, the geminiviruses, the parvoviruses and others - and also the many phages and plasmids that use the rolling circle replication (RCR) mechanism, the RCR endonuclease creates a nick the genome strand (single stranded viruses) or one of the DNA strands (plasmids). The 5 end of the nicked strand is transferred to a tyrosine residue on the nuclease and the free 3 OH group is then used by the DNA polymerase for new strand synthesis. The best known of these mechanisms is that used by the cellular organisms. In these once the two strands are separated, RNA primers are created on the template strands. To be more specific, the leading strand receives one RNA primer per active origin of replication while the lagging strand receives several; these several fragments of RNA primers found on the lagging strand of DNA are called Okazaki fragments, named after their discoverer. DNA polymerase extends the leading strand in one continuous motion and the lagging strand in a discontinuous motion (due to the Okazaki fragments). RNase removes the RNA fragments used to initiate replication by DNA polymerase, and another DNA Polymerase enters to fill the gaps. When this is complete, a single nick on the leading strand and several nicks on the lagging strand can be found. Ligase works to fill these nicks in, thus completing the newly replicated DNA molecule. The primase used in this process differs significantly between bacteria and archaea/eukaryotes. Bacteria use a primase belonging to the DnaG protein superfamily which contains a catalytic domain of the TOPRIM fold type. The TOPRIM fold contains an / core with four conserved strands in a Rossmann-like topology. This structure is also found in the catalytic domains of topoisomerase Ia, topoisomerase II, the OLD-family nucleases and DNA repair proteins related to the RecR protein. The primase used by archaea and eukaryotes in contrast contains a highly derived version of the RNA recognition motif (RRM). This primase is structurally similar to many viral RNA dependent RNA polymerases, reverse transcriptases, cyclic nucleotide generating cyclases and DNA polymerases of the A/B/Y families that are involved in DNA replication and repair. All these proteins share a catalytic mechanism of di-metal-ion-mediated nucleotide transfer, whereby two acidic residues located at the end of the first strand and between the second and third strands of the RRM-like unit respectively, chelate two divalent cations. As DNA synthesis continues, the original DNA strands continue to unwind on each side of the bubble, forming a replication fork with two prongs. In bacteria, which have a single origin of replication on their circular chromosome, this process eventually creates a "theta structure" (resembling the Greek letter theta: ). In contrast, eukaryotes have longer linear chromosomes and initiate replication at multiple origins within these.

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Replication fork
The replication fork is a structure that forms within the nucleus during DNA replication. It is created by helicases, which break the hydrogen bonds holding the two DNA strands together. The resulting structure has two branching "prongs", each one made up of a single strand of DNA. These two strands serve as the template for the leading and lagging strands, which will be created as DNA polymerase matches complementary Many enzymes are involved in the DNA replication fork. nucleotides to the templates; The templates may be properly referred to as the leading strand template and the lagging strand template. Leading strand The leading strand is the template strand of the DNA double helix so that the replication fork moves along it in the 3' to 5' direction. This allows the newly synthesized strand complementary to the original strand to be synthesized 5' to 3' in the same direction as the movement of the replication fork. On the leading strand, a polymerase "reads" the DNA and adds nucleotides to it continuously. This polymerase is DNA polymerase III (DNA Pol III) in prokaryotes and presumably Pol [11] [12] in yeasts. In human cells the leading and lagging strands are synthesized by Pol and Pol within the nucleus and Pol in the mitochondria. Pol can substitute for Pol in special circumstances[13] . Lagging strand The lagging strand is the strand of the template DNA double helix that is oriented so that the replication fork moves along it in a 5' to 3' manner. Because of its orientation, opposite to the working orientation of DNA polymerase III, which moves on a template in a 3' to 5' manner, replication of the lagging strand is more complicated than that of the leading strand. On the lagging strand, primase "reads" the DNA and adds RNA to it in short, separated segments. In eukaryotes, primase is intrinsic to Pol .[14] DNA polymerase III or Pol lengthens the primed segments, forming Okazaki fragments. Primer removal in eukaryotes is also performed by Pol .[15] In prokaryotes, DNA polymerase I "reads" the fragments, removes the RNA using its flap endonuclease domain (RNA primers are removed by 5'-3' exonuclease activity of polymerase I [weaver, 2005], and replaces the RNA nucleotides with DNA nucleotides (this is necessary because RNA and DNA use slightly different kinds of nucleotides). DNA ligase joins the fragments together.

DNA replication Dynamics at the replication fork As helicase unwinds DNA at the replication fork, the DNA ahead is forced to rotate. This process results in a build-up of twists in the DNA ahead.[16] This build-up would form a resistance that would eventually halt the progress of the replication fork. DNA Gyrase is an enzyme that temporarily breaks the strands of DNA, relieving the tension caused by unwinding the two strands of the DNA helix; DNA Gyrase achieves this by adding negative supercoils to the DNA helix. [17] Bare single-stranded DNA tends to fold back on itself and form secondary structures; these structures can interfere with the movement of DNA polymerase. To prevent this, single-strand binding proteins bind to the DNA until a second strand is synthesized, preventing secondary structure formation.[18]
The assembled human DNA clamp, a trimer of the Clamp proteins form a sliding clamp around DNA, helping the protein PCNA. DNA polymerase maintain contact with its template, thereby assisting with processivity. The inner face of the clamp enables DNA to be threaded through it. Once the polymerase reaches the end of the template or detects double-stranded DNA, the sliding clamp undergoes a conformational change that releases the DNA polymerase. Clamp-loading proteins are used to initially load the clamp, recognizing the junction between template and RNA primers.

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Regulation
Eukaryotes Within eukaryotes, DNA replication is controlled within the context of the cell cycle. As the cell grows and divides, it progresses through stages in the cell cycle; DNA replication occurs during the S phase (synthesis phase). The progress of the eukaryotic cell through the cycle is controlled by cell cycle checkpoints. Progression through checkpoints is controlled through complex interactions between various proteins, including cyclins and cyclin-dependent kinases.[19] The G1/S checkpoint (or restriction checkpoint) regulates whether eukaryotic cells enter the process of DNA replication and subsequent division. Cells that do not proceed through this checkpoint are remain in the G0 stage and do not replicate their DNA.

The cell cycle of eukaryotic cells.

Replication of chloroplast and mitochondrial genomes occurs independent of the cell cycle, through the process of D-loop replication. Bacteria Most bacteria do not go through a well-defined cell cycle but instead continuously copy their DNA; during rapid growth, this can result in the concurrent occurrences of multiple rounds of replication.[20] In E. coli, the best-characterized bacteria, DNA replication is regulated through several mechanisms, including: the hemimethylation and sequestering of the origin sequence, the ratio of ATP to ADP, and the levels of protein DnaA. All these control the process of initiator proteins binding to the origin sequences. Because E. coli methylates GATC DNA sequences, DNA synthesis results in hemimethylated sequences. This hemimethylated DNA is recognized by the protein SeqA, which binds and sequesters the origin sequence; in

DNA replication addition, dnaA (required for initiation of replication) binds less well to hemimethylated DNA. As a result, newly replicated origins are prevented from immediately initiating another round of DNA replication.[21] ATP builds up when the cell is in a rich medium, triggering DNA replication once the cell has reached a specific size. ATP competes with ADP to bind to DnaA, and the DnaA-ATP complex is able to initiate replication. A certain number of DnaA proteins are also required for DNA replication each time the origin is copied, the number of binding sites for DnaA doubles, requiring the synthesis of more DnaA to enable another initiation of replication.

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Termination
Eukaryotes initiate DNA replication at multiple points in the chromosome, so replication forks meet and terminate at many points in the chromosome; these are not known to be regulated in any particular way. Because eukaryotes have linear chromosomes, DNA replication is unable to reach the very end of the chromosomes, but ends at the telomere region of repetitive DNA close to the end. This shortens the telomere of the daughter DNA strand. This is a normal process in somatic cells. As a result, cells can only divide a certain number of times before the DNA loss prevents further division. (This is known as the Hayflick limit.) Within the germ cell line, which passes DNA to the next generation, telomerase extends the repetitive sequences of the telomere region to prevent degradation. Telomerase can become mistakenly active in somatic cells, sometimes leading to cancer formation. Because bacteria have circular chromosomes, termination of replication occurs when the two replication forks meet each other on the opposite end of the parental chromosome. E coli regulate this process through the use of termination sequences that, when bound by the Tus protein, enable only one direction of replication fork to pass through. As a result, the replication forks are constrained to always meet within the termination region of the chromosome.[22]

Polymerase chain reaction

Researchers commonly replicate DNA in vitro using the polymerase chain reaction (PCR). PCR uses a pair of primers to span a target region in template DNA, and then polymerizes partner strands in each direction from these primers using a thermostable DNA polymerase. Repeating this process through multiple cycles produces amplification of the targeted DNA region. At the start of each cycle, the mixture of template and primers is heated, separating the newly synthesized molecule and template. Then, as the mixture cools, both of these become templates for annealing of new primers, and the polymerase extends from these. As a result, the number of copies of the target region doubles each round, increasing exponentially.[23]

References
[1] hczohzhohzohz Berg JM, Tymoczko JL, Stryer L, Clarke ND (2002). Biochemistry. W.H. Freeman and Company. ISBN0-7167-3051-0. Chapter 27: DNA Replication, Recombination, and Repair (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fc,kgi?rid=stryer. chapter. 3740) [2] Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell. Garland Science. ISBN0-8153-3218-1. Chapter 5: DNA Replication, Repair, and Recombination (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. chapter. 747) [3] Berg JM, Tymoczko JL, Stryer L, Clarke ND (2002). Biochemistry. W.H. Freeman and Company. ISBN0-7167-3051-0. Chapter 27, Section 4: DNA Replication of Both Strands Proceeds Rapidly from Specific Start Sites (http:/ / www. ncbi. nlm. nih. gov/ bofghoks/ bv. fcvbngi?rid=stryer. section. 3794) [4] Alberts, B., et.al., Molecular Biology of the Cell, Garland Science, 4th ed., 2002, pp. 238-240 ISBN 0-8153-3218-1 [5] Allison, Lizabeth A. Fundamental Molecular Biology. Blackwell Publishing. 2007. p.112. [6] Berg JM, Tymoczko JL, Stryer L, Clarke ND (2002). Biochemistry. W.H. Freeman and Company. ISBN0-7167-3051-0. Chapter 27, Section 2: DNA Polymerases Require a Template and a Primer (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=stryer. section. 3769) [7] McCulloch SD, Kunkel TA (January 2008). "The fidelity of DNA synthesis by eukaryotic replicative and translesion synthesis polymerases". Cell Research 18 (1): 14861. doi:10.1038/cr.2008.4. PMID18166979. [8] Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell. Garland Science. ISBN0-8153-3218-1. Chapter 5: DNA Replication Mechanisms (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. section. 754) [9] Weigel C, Schmidt A, Rckert B, Lurz R, Messer W (November 1997). "DnaA protein binding to individual DnaA boxes in the Escherichia coli replication origin, oriC". The EMBO Journal 16 (21): 657483. doi:10.1093/emboj/16.21.6574. PMC1170261. PMID9351837.

DNA replication
[10] Lodish H, Berk A, Zipursky LS, Matsudaira P, Baltimore D, Darnell J (2000). Molecular Cell Biology. W. H. Freeman and Company. ISBN0-7167-3136-3. 12.1. General Features of Chromosomal Replication: Three Common Features of Replication Origins (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mcb. section. 3163#3179) [11] Pursell, Z.F. et al. (2007). "Yeast DNA Polymerase Participates in Leading-Strand DNA Replication". Science 317 (5834): 127130. doi:10.1126/science.1144067. PMC2233713. PMID17615360. [12] Scott D McCulloch; Thomas A Kunkel (01/2008). "The fidelity of DNA synthesis by eukaryotic replicative and translesion synthesis polymerases". Cell Research 19 (1): 148161. doi:10.1038/cr.2008.4. PMID18166979. [13] Hansen, Barbara (2011). Biochemistry and Medical Genetics: Lecture Notes. Kaplan Medical. pp.21. [14] Elizabeth R. Barry; Stephen D. Bell (12/2006). "DNA Replication in the Archaea". Microbiology and Molecular Biology Reviews 70 (4): 876887. doi:10.1128/MMBR.00029-06. PMC1698513. PMID17158702. [15] Distinguishing the pathways of primer removal during Eukaryotic Okazaki fragment maturation (http:/ / hdl. handle. net/ 1802/ 6537) Contributor Author Rossi, Marie Louise. Date Accessioned: 2009-02-23T17:05:09Z. Date Available: 2009-02-23T17:05:09Z. Date Issued: 2009-02-23T17:05:09Z. Identifier Uri: http:/ / hdl. handle. net/ 1802/ 6537. Description: Dr. Robert A. Bambara, Faculty Advisor. Thesis (PhD) - School of Medicine and Dentistry, University of Rochester. UR only until January 2010. UR only until January 2010. [16] Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell. Garland Science. ISBN0-8153-3218-1. DNA Replication Mechanisms: DNA Topoisomerases Prevent DNA Tangling During Replication (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. section. 754#787) [17] http:/ / www. ncbi. nlm. nih. gov/ pubmed/ 1657531 [18] Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell. Garland Science. ISBN0-8153-3218-1. DNA Replication Mechanisms: Special Proteins Help to Open Up the DNA Double Helix in Front of the Replication Fork (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. section. 754#774) [19] Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). Molecular Biology of the Cell. Garland Science. ISBN0-8153-3218-1. Intracellular Control of Cell-Cycle Events: S-Phase Cyclin-Cdk Complexes (S-Cdks) Initiate DNA Replication Once Per Cycle (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=mboc4. section. 3214#3215) [20] Tobiason DM, Seifert HS (2006). "The Obligate Human Pathogen, Neisseria gonorrhoeae, Is Polyploid". PLoS Biology 4 (6): e185. doi:10.1371/journal.pbio.0040185. PMC1470461. PMID16719561. [21] Slater S, Wold S, Lu M, Boye E, Skarstad K, Kleckner N (September 1995). "E. coli SeqA protein binds oriC in two different methyl-modulated reactions appropriate to its roles in DNA replication initiation and origin sequestration". Cell 82 (6): 92736. doi:10.1016/0092-8674(95)90272-4. PMID7553853. [22] TA Brown (2002). Genomes. BIOS Scientific Publishers. ISBN1-85996-228-9. 13.2.3. Termination of replication (http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?rid=genomes. section. 8121#8156) [23] Saiki, RK; Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA (1988). "Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase" (http:/ / sunsite. berkeley. edu/ cgi-bin/ ebind2html/ pcr/ 009). Science 239 (4839): 48791. doi:10.1126/science.2448875. PMID2448875. .

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DNA repair
DNA repair refers to a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome. In human cells, both normal metabolic activities and environmental factors such as UV light and radiation can cause DNA damage, resulting in as many as 1 million individual molecular lesions per cell per day.[1] Many of these lesions cause structural damage to the DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions induce potentially harmful mutations in the cell's genome, which affect the survival of its daughter cells after it undergoes mitosis. As a consequence, the DNA repair process is constantly active as it responds to damage in the DNA structure. When normal repair processes fail, and when cellular apoptosis does not occur, irreparable DNA damage may occur, including double-strand breaks and DNA crosslinkages.[2] [3]
DNA damage resulting in multiple broken The rate of DNA repair is dependent on many factors, including the chromosomes cell type, the age of the cell, and the extracellular environment. A cell that has accumulated a large amount of DNA damage, or one that no longer effectively repairs damage incurred to its DNA, can enter one of three possible states:

1. an irreversible state of dormancy, known as senescence 2. cell suicide, also known as apoptosis or programmed cell death 3. unregulated cell division, which can lead to the formation of a tumor that is cancerous The DNA repair ability of a cell is vital to the integrity of its genome and thus to its normal functioning and that of the organism. Many genes that were initially shown to influence life span have turned out to be involved in DNA damage repair and protection.[4] Failure to correct molecular lesions in cells that form gametes can introduce mutations into the genomes of the offspring and thus influence the rate of evolution.

DNA damage
DNA damage, due to environmental factors and normal metabolic processes inside the cell, occurs at a rate of 1,000 to 1,000,000 molecular lesions per cell per day.[1] While this constitutes only 0.000165% of the human genome's approximately 6 billion bases (3 billion base pairs), unrepaired lesions in critical genes (such as tumor suppressor genes) can impede a cell's ability to carry out its function and appreciably increase the likelihood of tumor formation. The vast majority of DNA damage affects the primary structure of the double helix; that is, the bases themselves are chemically modified. These modifications can in turn disrupt the molecules' regular helical structure by introducing non-native chemical bonds or bulky adducts that do not fit in the standard double helix. Unlike proteins and RNA, DNA usually lacks tertiary structure and therefore damage or disturbance does not occur at that level. DNA is, however, supercoiled and wound around "packaging" proteins called histones (in eukaryotes), and both superstructures are vulnerable to the effects of DNA damage.

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Sources of damage
DNA damage can be subdivided into two main types: 1. endogenous damage such as attack by reactive oxygen species produced from normal metabolic byproducts (spontaneous mutation), especially the process of oxidative deamination 1. also includes replication errors 2. exogenous damage caused by external agents such as 1. 2. 3. 4. 5. 6. 7. ultraviolet [UV 200-300 nm] radiation from the sun other radiation frequencies, including x-rays and gamma rays hydrolysis or thermal disruption certain plant toxins human-made mutagenic chemicals, especially aromatic compounds that act as DNA intercalating agents cancer chemotherapy and radiotherapy viruses [5]

The replication of damaged DNA before cell division can lead to the incorporation of wrong bases opposite damaged ones. Daughter cells that inherit these wrong bases carry mutations from which the original DNA sequence is unrecoverable (except in the rare case of a back mutation, for example, through gene conversion).

Types of damage
There are five main types of damage to DNA due to endogenous cellular processes: 1. oxidation of bases [e.g. 8-oxo-7,8-dihydroguanine (8-oxoG)] and generation of DNA strand interruptions from reactive oxygen species, 2. alkylation of bases (usually methylation), such as formation of 7-methylguanine, 1-methyladenine, 6-O-Methylguanine 3. hydrolysis of bases, such as deamination, depurination, and depyrimidination. 4. "bulky adduct formation" (i.e., benzo[a]pyrene diol epoxide-dG adduct) 5. mismatch of bases, due to errors in DNA replication, in which the wrong DNA base is stitched into place in a newly forming DNA strand, or a DNA base is skipped over or mistakenly inserted. Damage caused by exogenous agents comes in many forms. Some examples are: 1. UV-B light causes crosslinking between adjacent cytosine and thymine bases creating pyrimidine dimers. This is called direct DNA damage. 2. UV-A light creates mostly free radicals. The damage caused by free radicals is called indirect DNA damage. 3. Ionizing radiation such as that created by radioactive decay or in cosmic rays causes breaks in DNA strands. Low-level ionizing radiation may induce irreparable DNA damage (leading to replicational and transcriptional errors needed for neoplasia or may trigger viral interactions) leading to pre-mature aging and cancer.[6] [7] [8] 4. Thermal disruption at elevated temperature increases the rate of depurination (loss of purine bases from the DNA backbone) and single-strand breaks. For example, hydrolytic depurination is seen in the thermophilic bacteria, which grow in hot springs at 40-80 C.[9] [10] The rate of depurination (300 purine residues per genome per generation) is too high in these species to be repaired by normal repair machinery, hence a possibility of an adaptive response cannot be ruled out. 5. Industrial chemicals such as vinyl chloride and hydrogen peroxide, and environmental chemicals such as polycyclic hydrocarbons found in smoke, soot and tar create a huge diversity of DNA adducts- ethenobases, oxidized bases, alkylated phosphotriesters and Crosslinking of DNA just to name a few. UV damage, alkylation/methylation, X-ray damage and oxidative damage are examples of induced damage. Spontaneous damage can include the loss of a base, deamination, sugar ring puckering and tautomeric shift.[11]

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Nuclear versus mitochondrial DNA damage

In human cells, and eukaryotic cells in general, DNA is found in two cellular locations - inside the nucleus and inside the mitochondria. Nuclear DNA (nDNA) exists as chromatin during non-replicative stages of the cell cycle and is condensed into aggregate structures known as chromosomes during cell division. In either state the DNA is highly compacted and wound up around bead-like proteins called histones. Whenever a cell needs to express the genetic information encoded in its nDNA the required chromosomal region is unravelled, genes located therein are expressed, and then the region is condensed back to its resting conformation. Mitochondrial DNA (mtDNA) is located inside mitochondria organelles, exists in multiple copies, and is also tightly associated with a number of proteins to form a complex known as the nucleoid. Inside mitochondria, reactive oxygen species (ROS), or free radicals, byproducts of the constant production of adenosine triphosphate (ATP) via oxidative phosphorylation, create a highly oxidative environment that is known to damage mtDNA. A critical enzyme in counteracting the toxicity of these species is superoxide dismutase, which is present in both the mitochondria and cytoplasm of eukaryotic cells.

Senescence and apoptosis

Senescence, an irreversible state in which the cell no longer divides, is a protective response to the shortening of the chromosome ends. The telomeres are long regions of repetitive noncoding DNA that cap chromosomes and undergo partial degradation each time a cell undergoes division (see Hayflick limit).[12] In contrast, quiescence is a reversible state of cellular dormancy that is unrelated to genome damage (see cell cycle). Senescence in cells may serve as a functional alternative to apoptosis in cases where the physical presence of a cell for spatial reasons is required by the organism,[13] which serves as a "last resort" mechanism to prevent a cell with damaged DNA from replicating inappropriately in the absence of pro-growth cellular signaling. Unregulated cell division can lead to the formation of a tumor (see cancer), which is potentially lethal to an organism. Therefore, the induction of senescence and apoptosis is considered to be part of a strategy of protection against cancer.[14]

DNA damage and mutation

It is important to distinguish between DNA damage and mutation, the two major types of error in DNA. DNA damages and mutation are fundamentally different. Damages are physical abnormalities in the DNA, such as singleand double-strand breaks, 8-hydroxydeoxyguanosine residues, and polycyclic aromatic hydrocarbon adducts. DNA damages can be recognized by enzymes, and, thus, they can be correctly repaired if redundant information, such as the undamaged sequence in the complementary DNA strand or in a homologous chromosome, is available for copying. If a cell retains DNA damage, transcription of a gene can be prevented, and, thus, translation into a protein will also be blocked. Replication may also be blocked and/or the cell may die. In contrast to DNA damage, a mutation is a change in the base sequence of the DNA. A mutation cannot be recognized by enzymes once the base change is present in both DNA strands, and, thus, a mutation cannot be repaired. At the cellular level, mutations can cause alterations in protein function and regulation. Mutations are replicated when the cell replicates. In a population of cells, mutant cells will increase or decrease in frequency according to the effects of the mutation on the ability of the cell to survive and reproduce. Although distinctly different from each other, DNA damages and mutations are related because DNA damages often cause errors of DNA synthesis during replication or repair; these errors are a major source of mutation. Given these properties of DNA damage and mutation, it can be seen that DNA damages are a special problem in non-dividing or slowly dividing cells, where unrepaired damages will tend to accumulate over time. On the other hand, in rapidly dividing cells, unrepaired DNA damages that do not kill the cell by blocking replication will tend to cause replication errors and thus mutation. The great majority of mutations that are not neutral in their effect are deleterious to a cells survival. Thus, in a population of cells comprising a tissue with replicating cells, mutant cells will tend to be lost. However, infrequent mutations that provide a survival advantage will tend to clonally expand at

DNA repair the expense of neighboring cells in the tissue. This advantage to the cell is disadvantageous to the whole organism, because such mutant cells can give rise to cancer. Thus, DNA damages in frequently dividing cells, because they give rise to mutations, are a prominent cause of cancer. In contrast, DNA damages in infrequently dividing cells are likely a prominent cause of aging.[15]

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DNA repair mechanisms

Cells cannot function if DNA damage corrupts the integrity and accessibility of essential information in the genome (but cells remain superficially functional when so-called "non-essential" genes are missing or damaged). Depending on the type of damage inflicted on the DNA's double helical structure, a variety of repair strategies have evolved to restore lost information. If possible, cells use the unmodified complementary strand of the DNA or the sister chromatid as a template to recover the original information. Without access to a template, cells use an error-prone recovery mechanism known as translesion synthesis as a last resort. Damage to DNA alters the spatial configuration of the helix, and such alterations can be detected by the cell. Once damage is localized, specific DNA repair molecules bind at or near the site of damage, inducing other molecules to bind and form a complex that enables the actual repair to take place.

Direct reversal
Cells are known to eliminate three types of damage to their DNA by chemically reversing it. These mechanisms do not require a template, since the types of damage they counteract can occur in only one of the four bases. Such direct reversal mechanisms are specific to the type of damage incurred and do not involve breakage of the phosphodiester backbone. The formation of pyrimidine dimers upon irradiation with UV light results in an abnormal covalent bond between adjacent pyrimidine bases. The photoreactivation process directly reverses this damage by the action of the enzyme photolyase, whose activation is obligately Single-strand and double-strand dependent on energy absorbed from blue/UV light (300500nm wavelength) to DNA damage promote catalysis.[16] Another type of damage, methylation of guanine bases, is directly reversed by the protein methyl guanine methyl transferase (MGMT), the bacterial equivalent of which is called ogt. This is an expensive process because each MGMT molecule can be used only once; that is, the reaction is stoichiometric rather than catalytic.[17] A generalized response to methylating agents in bacteria is known as the adaptive response and confers a level of resistance to alkylating agents upon sustained exposure by upregulation of alkylation repair enzymes.[18] The third type of DNA damage reversed by cells is certain methylation of the bases cytosine and adenine.

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Single-strand damage
When only one of the two strands of a double helix has a defect, the other strand can be used as a template to guide the correction of the damaged strand. In order to repair damage to one of the two paired molecules of DNA, there exist a number of excision repair mechanisms that remove the damaged nucleotide and replace it with an undamaged nucleotide complementary to that found in the undamaged DNA strand.[17] 1. Base excision repair (BER), which repairs damage to a single base caused by oxidation, alkylation, hydrolysis, or deamination. The damaged base is Structure of the base-excision repair enzyme uracil-DNA removed by a DNA glycosylase. The "missing glycosylase. The uracil residue is shown in yellow. tooth" is then recognized by an enzyme called AP endonuclease, which cuts the Phosphodiester bond. The missing part is then resynthesized by a DNA polymerase, and a DNA ligase performs the final nick-sealing step. 2. Nucleotide excision repair (NER), which recognizes bulky, helix-distorting lesions such as pyrimidine dimers and 6,4 photoproducts. A specialized form of NER known as transcription-coupled repair deploys NER enzymes to genes that are being actively transcribed. 3. Mismatch repair (MMR), which corrects errors of DNA replication and recombination that result in mispaired (but undamaged) nucleotides.

Double-strand breaks
Double-strand breaks, in which both strands in the double helix are severed, are particularly hazardous to the cell because they can lead to genome rearrangements. Three mechanisms exist to repair double-strand breaks (DSBs): non-homologous end joining (NHEJ), microhomology-mediated end joining (MMEJ), and homologous recombination.[17] PVN Acharya noted that double-strand breaks and a "cross-linkage joining both strands at the same point is irreparable because neither strand can then serve as a template for repair. The cell will die in the next mitosis or in some rare instances, mutate."[2] [3]

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In NHEJ, DNA Ligase IV, a specialized DNA ligase that forms a complex with the cofactor XRCC4, directly joins the two ends.[19] To guide accurate repair, NHEJ relies on short homologous sequences called microhomologies present on the single-stranded tails of the DNA ends to be joined. If these overhangs are compatible, repair is usually accurate.[20] [21] [22] [23] NHEJ can also introduce mutations during repair. Loss of damaged nucleotides at the break site can lead to deletions, and joining of nonmatching termini forms translocations. NHEJ is especially important before the cell has replicated its DNA, since there is no template available for repair by homologous recombination. There are "backup" NHEJ pathways in higher eukaryotes.[24] Besides its role as a genome caretaker, NHEJ is required for joining hairpin-capped double-strand breaks induced during V(D)J recombination, the process that generates diversity in B-cell and T-cell receptors in the vertebrate immune system.[25] Homologous recombination requires the presence of an an enzyme that joins broken nucleotides together by catalyzing identical or nearly identical sequence to be used as a the formation of an internucleotide ester bond between the phosphate backbone and the deoxyribose nucleotides. template for repair of the break. The enzymatic machinery responsible for this repair process is nearly identical to the machinery responsible for chromosomal crossover during meiosis. This pathway allows a damaged chromosome to be repaired using a sister chromatid (available in G2 after DNA replication) or a homologous chromosome as a template. DSBs caused by the replication machinery attempting to synthesize across a single-strand break or unrepaired lesion cause collapse of the replication fork and are typically repaired by recombination. Topoisomerases introduce both single- and double-strand breaks in the course of changing the DNA's state of supercoiling, which is especially common in regions near an open replication fork. Such breaks are not considered DNA damage because they are a natural intermediate in the topoisomerase biochemical mechanism and are immediately repaired by the enzymes that created them. A team of French researchers bombarded Deinococcus radiodurans to study the mechanism of double-strand break DNA repair in that organism. At least two copies of the genome, with random DNA breaks, can form DNA fragments through annealing. Partially overlapping fragments are then used for synthesis of homologous regions through a moving D-loop that can continue extension until they find complementary partner strands. In the final step there is crossover by means of RecA-dependent homologous recombination.[26]
DNA ligase, shown above repairing chromosomal damage, is

Translesion synthesis
Translesion synthesis (TLS) is a DNA damage tolerance process that allows the DNA replication machinery to replicate past DNA lesions such as thymine dimers or AP sites.[27] It involves switching out regular DNA polymerases for specialized translesion polymerases (i.e. DNA polymerase IV or V, from the Y Polymerase family), often with larger active sites that can facilitate the insertion of bases opposite damaged nucleotides. The polymerase switching is thought to be mediated by, among other factors, the post-translational modification of the replication processivity factor PCNA. Translesion synthesis polymerases often have low fidelity (high propensity to insert wrong bases) on undamaged templates relative to regular polymerases. However, many are extremely efficient at inserting correct bases opposite specific types of damage. For example, Pol mediates error-free bypass of lesions induced by UV irradiation, whereas Pol introduces mutations at these sites. Pol is known to add the first adenine

DNA repair across the T^T photodimer using Watson-Crick base pairing and the second adenine will be added in its syn conformation using Hoogsteen base pairing. From a cellular perspective, risking the introduction of point mutations during translesion synthesis may be preferable to resorting to more drastic mechanisms of DNA repair, which may cause gross chromosomal aberrations or cell death. In short, the process involves specialized polymerases either bypassing or repairing lesions at locations of stalled DNA replication. A bypass platform is provided to these polymerases by Proliferating cell nuclear antigen (PCNA). Under normal circumstances, PCNA bound to polymerases replicates the DNA. At a site of lesion, PCNA is ubiquitinated, or modified, by the RAD6/RAD18 proteins to provide a platform for the specialized polymerases to bypass the lesion and resume DNA replication.[28] [29] After translesion synthesis, extension is required. This extension can be carried out by a replicative polymerase if the TLS is error-free, as in the case of Pol , yet if TLS results in a mismatch, a specialized polymerase is needed to extend it; Pol . Pol is unique in that it can extend terminal mismatches, whereas more processive polymerases cannot. So when a lesion is encountered, the replication fork will stall, PCNA will switch from a processive polymerase to a TLS polymerase such as Pol to fix the lesion, then PCNA may switch to Pol to extend the mismatch, and last PCNA will switch to the processive polymerase to continue replication.

364

Global response to DNA damage

Cells exposed to ionizing radiation, ultraviolet light or chemicals are prone to acquire multiple sites of bulky DNA lesions and double-strand breaks. Moreover, DNA damaging agents can damage other biomolecules such as proteins, carbohydrates, lipids, and RNA. The accumulation of damage, to be specific, double-strand breaks or adducts stalling the replication forks, are among known stimulation signals for a global response to DNA damage.[30] The global response to damage is an act directed toward the cells' own preservation and triggers multiple pathways of macromolecular repair, lesion bypass, tolerance, or apoptosis. The common features of global response are induction of multiple genes, cell cycle arrest, and inhibition of cell division.

DNA damage checkpoints

After DNA damage, cell cycle checkpoints are activated. Checkpoint activation pauses the cell cycle and gives the cell time to repair the damage before continuing to divide. DNA damage checkpoints occur at the G1/S and G2/M boundaries. An intra-S checkpoint also exists. Checkpoint activation is controlled by two master kinases, ATM and ATR. ATM responds to DNA double-strand breaks and disruptions in chromatin structure,[31] whereas ATR primarily responds to stalled replication forks. These kinases phosphorylate downstream targets in a signal transduction cascade, eventually leading to cell cycle arrest. A class of checkpoint mediator proteins including BRCA1, MDC1, and 53BP1 has also been identified.[32] These proteins seem to be required for transmitting the checkpoint activation signal to downstream proteins. P53 is an important downstream target of ATM and ATR, as it is required for inducing apoptosis following DNA damage.[33] At the G1/S checkpoint, p53 functions by deactivating the CDK2/cyclin E complex. Similarly, p21 mediates the G2/M checkpoint by deactivating the CDK1/cyclin B complex.

The prokaryotic SOS response

The SOS response is the term used to describe changes in gene expression in Escherichia coli and other bacteria in response to extensive DNA damage. The prokaryotic SOS system is regulated by two key proteins: LexA and RecA. The LexA homodimer is a transcriptional repressor that binds to operator sequences commonly referred to as SOS boxes. In Escherichia coli it is known that LexA regulates transcription of approximately 48 genes including the lexA and recA genes.[34] The SOS response is known to be widespread in the Bacteria domain, but it is mostly absent in some bacterial phyla, like the Spirochetes.[35] The most common cellular signals activating the SOS response are regions of single-stranded DNA (ssDNA), arising from stalled replication forks or double-strand breaks, which are processed by DNA helicase to separate the two DNA strands.[30] In the initiation step, RecA protein binds

DNA repair to ssDNA in an ATP hydrolysis driven reaction creating RecAssDNA filaments. RecAssDNA filaments activate LexA autoprotease activity, which ultimately leads to cleavage of LexA dimer and subsequent LexA degradation. The loss of LexA repressor induces transcription of the SOS genes and allows for further signal induction, inhibition of cell division and an increase in levels of proteins responsible for damage processing. In Escherichia coli, SOS boxes are 20-nucleotide long sequences near promoters with palindromic structure and a high degree of sequence conservation. In other classes and phyla, the sequence of SOS boxes varies considerably, with different length and composition, but it is always highly conserved and one of the strongest short signals in the genome.[35] The high information content of SOS boxes permits differential binding of LexA to different promoters and allows for timing of the SOS response. The lesion repair genes are induced at the beginning of SOS response. The error-prone translesion polymerases, for example, UmuCD2 (also called DNA polymerase V), are induced later on as a last resort.[36] Once the DNA damage is repaired or bypassed using polymerases or through recombination, the amount of single-stranded DNA in cells is decreased, lowering the amounts of RecA filaments decreases cleavage activity of LexA homodimer, which then binds to the SOS boxes near promoters and restores normal gene expression.

365

Eukaryotic transcriptional responses to DNA damage

Eukaryotic cells exposed to DNA damaging agents also activate important defensive pathways by inducing multiple proteins involved in DNA repair, cell cycle checkpoint control, protein trafficking and degradation. Such genome wide transcriptional response is very complex and tightly regulated, thus allowing coordinated global response to damage. Exposure of yeast Saccharomyces cerevisiae to DNA damaging agents results in overlapping but distinct transcriptional profiles. Similarities to environmental shock response indicates that a general global stress response pathway exist at the level of transcriptional activation. In contrast, different human cell types respond to damage differently indicating an absence of a common global response. The probable explanation for this difference between yeast and human cells may be in the heterogeneity of mammalian cells. In an animal different types of cells are distributed among different organs that have evolved different sensitivities to DNA damage.[36] In general global response to DNA damage involves expression of multiple genes responsible for postreplication repair, homologous recombination, nucleotide excision repair, DNA damage checkpoint, global transcriptional activation, genes controlling mRNA decay, and many others. A large amount of damage to a cell leaves it with an important decision: undergo apoptosis and die, or survive at the cost of living with a modified genome. An increase in tolerance to damage can lead to an increased rate of survival that will allow a greater accumulation of mutations. Yeast Rev1 and human polymerase are members of [Y family translesion DNA polymerases present during global response to DNA damage and are responsible for enhanced mutagenesis during a global response to DNA damage in eukaryotes.[30]

DNA repair

366

DNA repair and aging

Pathological effects of poor DNA repair
Experimental animals with genetic deficiencies in DNA repair often show decreased life span and increased cancer incidence.[15] For example, mice deficient in the dominant NHEJ pathway and in telomere maintenance mechanisms get lymphoma and infections more often, and, as a consequence, have shorter lifespans than wild-type mice.[37] In similar manner, mice deficient in a key repair and transcription protein that unwinds DNA helices have premature onset of aging-related diseases and consequent shortening of lifespan.[38] However, not every DNA repair deficiency creates exactly the predicted effects; mice deficient in the NER pathway exhibited shortened life span without correspondingly higher rates of mutation.[39]

DNA repair rate is an important determinant of cell pathology

If the rate of DNA damage exceeds the capacity of the cell to repair it, the accumulation of errors can overwhelm the cell and result in early senescence, apoptosis, or cancer. Inherited diseases associated with faulty DNA repair functioning result in premature aging,[15] increased sensitivity to carcinogens, and correspondingly increased cancer risk (see below). On the other hand, organisms with enhanced DNA repair systems, such as Deinococcus radiodurans, the most radiation-resistant known organism, exhibit remarkable resistance to the double-strand break-inducing effects of radioactivity, likely due to enhanced efficiency of DNA repair and especially NHEJ.[40]

Longevity and caloric restriction

A number of individual genes have been identified as influencing variations in life span within a population of organisms. The effects of these genes is strongly dependent on the environment, in particular, on the organism's diet. Caloric restriction reproducibly results in extended lifespan in a variety of organisms, likely via nutrient sensing pathways and decreased metabolic rate. The molecular mechanisms by which such restriction results in lengthened lifespan are as yet unclear (see[41] for some discussion); however, the behavior of many genes known to be involved in DNA repair is altered under conditions of caloric restriction. For example, increasing the gene dosage of the gene SIR-2, which regulates DNA packaging in the nematode worm Caenorhabditis elegans, can significantly extend lifespan.[42] The mammalian homolog of SIR-2 is known to induce downstream DNA repair factors involved in NHEJ, an activity that is especially promoted under conditions of caloric restriction.[43] Caloric restriction has been closely linked to the rate of base excision repair in the nuclear DNA of rodents,[44] although similar effects have not been observed in mitochondrial DNA.[45]

Most life span influencing genes affect the rate of DNA damage

It is interesting to note that the C. elegans gene AGE-1, an upstream effector of DNA repair pathways, confers dramatically extended life span under free-feeding conditions but leads to a decrease in reproductive fitness under conditions of caloric restriction.[46] This observation supports the pleiotropy theory of the biological origins of aging, which suggests that genes conferring a large survival advantage early in life will be selected for even if they carry a

DNA repair corresponding disadvantage late in life.

367

Medicine and DNA repair modulation

Hereditary DNA repair disorders
Defects in the NER mechanism are responsible for several genetic disorders, including: Xeroderma pigmentosum: hypersensitivity to sunlight/UV, resulting in increased skin cancer incidence and premature aging Cockayne syndrome: hypersensitivity to UV and chemical agents Trichothiodystrophy: sensitive skin, brittle hair and nails Mental retardation often accompanies the latter two disorders, suggesting increased vulnerability of developmental neurons. Other DNA repair disorders include: Werner's syndrome: premature aging and retarded growth Bloom's syndrome: sunlight hypersensitivity, high incidence of malignancies (especially leukemias). Ataxia telangiectasia: sensitivity to ionizing radiation and some chemical agents All of the above diseases are often called "segmental progerias" ("accelerated aging diseases") because their victims appear elderly and suffer from aging-related diseases at an abnormally young age, while not manifesting all the symptoms of old age. Other diseases associated with reduced DNA repair function include Fanconi's anemia, hereditary breast cancer and hereditary colon cancer.

DNA repair and cancer

Inherited mutations that affect DNA repair genes are strongly associated with high cancer risks in humans. Hereditary nonpolyposis colorectal cancer (HNPCC) is strongly associated with specific mutations in the DNA mismatch repair pathway. BRCA1 and BRCA2, two famous mutations conferring a hugely increased risk of breast cancer on carriers, are both associated with a large number of DNA repair pathways, especially NHEJ and homologous recombination. Cancer therapy procedures such as chemotherapy and radiotherapy work by overwhelming the capacity of the cell to repair DNA damage, resulting in cell death. Cells that are most rapidly dividing - most typically cancer cells - are preferentially affected. The side-effect is that other non-cancerous but rapidly dividing cells such as stem cells in the bone marrow are also affected. Modern cancer treatments attempt to localize the DNA damage to cells and tissues only associated with cancer, either by physical means (concentrating the therapeutic agent in the region of the tumor) or by biochemical means (exploiting a feature unique to cancer cells in the body).

DNA repair and evolution

The basic processes of DNA repair are highly conserved among both prokaryotes and eukaryotes and even among bacteriophage (viruses that infect bacteria); however, more complex organisms with more complex genomes have correspondingly more complex repair mechanisms.[47] The ability of a large number of protein structural motifs to catalyze relevant chemical reactions has played a significant role in the elaboration of repair mechanisms during evolution. For an extremely detailed review of hypotheses relating to the evolution of DNA repair, see.[48] The fossil record indicates that single-cell life began to proliferate on the planet at some point during the Precambrian period, although exactly when recognizably modern life first emerged is unclear. Nucleic acids became the sole and universal means of encoding genetic information, requiring DNA repair mechanisms that in their basic

DNA repair form have been inherited by all extant life forms from their common ancestor. The emergence of Earth's oxygen-rich atmosphere (known as the "oxygen catastrophe") due to photosynthetic organisms, as well as the presence of potentially damaging free radicals in the cell due to oxidative phosphorylation, necessitated the evolution of DNA repair mechanisms that act specifically to counter the types of damage induced by oxidative stress.

368

Rate of evolutionary change

On some occasions, DNA damage is not repaired, or is repaired by an error-prone mechanism that results in a change from the original sequence. When this occurs, mutations may propagate into the genomes of the cell's progeny. Should such an event occur in a germ line cell that will eventually produce a gamete, the mutation has the potential to be passed on to the organism's offspring. The rate of evolution in a particular species (or, in a particular gene) is a function of the rate of mutation. As a consequence, the rate and accuracy of DNA repair mechanisms have an influence over the process of evolutionary change.[49]

References
[1] Lodish H, Berk A, Matsudaira P, Kaiser CA, Krieger M, Scott MP, Zipursky SL, Darnell J. (2004). Molecular Biology of the Cell, p963. WH Freeman: New York, NY. 5th ed. [2] Acharya, PV (1971). "The isolation and partial characterization of age-correlated oligo-deoxyribo-ribonucleotides with covalently linked aspartyl-glutamyl polypeptides.". Johns Hopkins medical journal. Supplement (1): 25460. PMID5055816. [3] Bjorksten, J; Acharya, PV; Ashman, S; Wetlaufer, DB (1971). "Gerogenic fractions in the tritiated rat.". Journal of the American Geriatrics Society 19 (7): 56174. PMID5106728. [4] Browner, WS; Kahn, AJ; Ziv, E; Reiner, AP; Oshima, J; Cawthon, RM; Hsueh, WC; Cummings, SR. (2004). "The genetics of human longevity". Am J Med 117 (11): 85160. doi:10.1016/j.amjmed.2004.06.033. PMID15589490. [5] Roulston A, Marcellus RC, Branton PE (1999). "Viruses and apoptosis" (http:/ / arjournals. annualreviews. org/ doi/ abs/ 10. 1146/ annurev. micro. 53. 1. 577?url_ver=Z39. 88-2003& rfr_id=ori:rid:crossref. org& rfr_dat=cr_pub=ncbi. nlm. nih. gov). Annu. Rev. Microbiol. 53: 577628. doi:10.1146/annurev.micro.53.1.577. PMID10547702. . Retrieved 2008-12-20. [6] Acharya, PVN; The Effect of Ionizing Radiation on the Formation of Age-Correlated Oligo Deoxyribo Nucleo Phospheryl Peptides in Mammalian Cells; 10th International Congress of Gerontology, Jerusalem. Abstract No. 1; January 1975. Work done while employed by Dept. of Pathology, University of Wisconsin, Madison. [7] Acharya, PVN; Implicatons of The Action of Low-Level Ionizing Radiation on the Inducement of Irreparable DNA Damage Leading to Mammalian Aging and Chemical Carcinogenesis.; 10th International Congress of Biochemistry, Hamburg, Germany. Abstract No. 01-1-079; July 1976. Work done while employed by Dept. of Pathology, University of Wisconsin, Madison. [8] Acharya, PV Narasimh; Irreparable DNA-Damage by Industrial Pollutants in Pre-mature Aging, Chemical Carcinogenesis and Cardiac Hypertrophy: Experiments and Theory; 1st International Meeting of Heads of Clinical Biochemistry Laboratories, Jerusalem, Israel. April 1977. Work conducted at Industrial Safety Institute and Behavioral Cybernetics Laboratory, University of Wisconsin, Madison. [9] Madigan MT, Martino JM (2006). Brock Biology of Microorganisms (11th ed.). Pearson. pp.136. ISBN0-13-196893-9. [10] Ohta, Toshihiro; Shin-ichi, Tokishita; Mochizuki, Kayo; Kawase, Jun; Sakahira, Masahide; Yamagata, Hideo (2006). "UV Sensitivity and Mutagenesis of the Extremely Thermophilic Eubacterium Thermus thermophilus HB27" (http:/ / www. jstage. jst. go. jp/ article/ jemsge/ 28/ 2/ 56/ _pdf). Genes and Environment 28 (2): 5661. doi:10.3123/jemsge.28.56. . [11] DNA Lesions That Require Repair: http:/ / www. ncbi. nlm. nih. gov/ books/ bv. fcgi?highlight=lesion& rid=mcb. table. 3236 [12] Braig, M; Schmitt, CA. (2006). "Oncogene-induced senescence: putting the brakes on tumor development". Cancer Res 66 (6): 28812884. doi:10.1158/0008-5472.CAN-05-4006. PMID16540631. [13] Lynch, MD. (2006). "How does cellular senescence prevent cancer?". DNA Cell Biol 25 (2): 6978. doi:10.1089/dna.2006.25.69. PMID16460230. [14] Campisi J, d'Adda di Fagagna F (2007). "Cellular senescence: when bad things happen to good cells.". Rev Mol Cell Biol. 8 (9): 72940. doi:10.1038/nrm2233. PMID17667954. [15] Best,BP (2009). "Nuclear DNA damage as a direct cause of aging" (http:/ / www. benbest. com/ lifeext/ Nuclear_DNA_in_Aging. pdf) (PDF). Rejuvenation Research 12 (3): 199208. doi:10.1089/rej.2009.0847. PMID19594328. . [16] Sancar, A. (2003). "Structure and function of DNA photolyase and cryptochrome blue-light photoreceptors". Chem Rev 103 (6): 220337. doi:10.1021/cr0204348. PMID12797829. [17] Watson JD, Baker TA, Bell SP, Gann A, Levine M, Losick R. (2004). Molecular Biology of the Gene, ch. 9 and 10. Peason Benjamin Cummings; CSHL Press. 5th ed. [18] Volkert, MR. (1988). "Adaptive response of Escherichia coli to alkylation damage". Environ Mol Mutagen 11 (2): 24155. doi:10.1002/em.2850110210. PMID3278898. [19] Wilson, TE; Grawunder, U; Lieber, MR (1997). "Yeast DNA ligase IV mediates non-homologous DNA end joining.". Nature 388 (6641): 4958. doi:10.1038/41365. PMID9242411.

DNA repair
[20] Moore, JK; Haber, JE (1996). "Cell cycle and genetic requirements of two pathways of nonhomologous end-joining repair of double-strand breaks in Saccharomyces cerevisiae". Molecular and cellular biology 16 (5): 216473. PMC231204. PMID8628283. [21] Boulton, SJ; Jackson, SP (1996). "Saccharomyces cerevisiae Ku70 potentiates illegitimate DNA double-strand break repair and serves as a barrier to error-prone DNA repair pathways". The EMBO journal 15 (18): 5093103. PMC452249. PMID8890183. [22] Wilson, TE; Lieber, MR (1999). "Efficient processing of DNA ends during yeast nonhomologous end joining. Evidence for a DNA polymerase beta (Pol4)-dependent pathway". The Journal of biological chemistry 274 (33): 23599609. doi:10.1074/jbc.274.33.23599. PMID10438542. [23] Budman, J; Chu, G (2005). "Processing of DNA for nonhomologous end-joining by cell-free extract". The EMBO journal 24 (4): 84960. doi:10.1038/sj.emboj.7600563. PMC549622. PMID15692565. [24] Wang, H; Wang, H; Perrault, AR; Takeda, Y; Qin, W; Iliakis, G. (2003). "Biochemical evidence for Ku-independent backup pathways of NHEJ". Nucleic Acids Res 31 (18): 537788. doi:10.1093/nar/gkg728. PMC203313. PMID12954774. [25] Jung, D; Alt, FW (2004). "Unraveling V(D)J recombination; insights into gene regulation". Cell 116 (2): 299311. doi:10.1016/S0092-8674(04)00039-X. PMID14744439. [26] Zahradka K, Slade D, Bailone A, Sommer S, Averbeck D, Petranovic M, Lindner AB, Radman M (2006). "Reassembly of shattered chromosomes in Deinococcus radiodurans". NATURE 443 (7111): 569573. doi:10.1038/nature05160. PMID17006450. [27] Waters LS, Minesinger BK, Wiltrout ME, D'Souza S, Woodruff RV, Walker GC (March 2009). "Eukaryotic Translesion Polymerases and Their Roles and Regulation in DNA Damage Tolerance". Microbiol. Mol. Biol. Rev. 73 (1): 13454. doi:10.1128/MMBR.00034-08. PMC2650891. PMID19258535. [28] http:/ / research. chem. psu. edu/ sjbgroup/ projects/ translesion. htm [29] Wang, Z (2001). "Translesion synthesis by the UmuC family of DNA polymerases". Mutat. Res. 486 (2): 5970. doi:10.1016/S0921-8777(01)00089-1. PMID11425512. [30] Friedberg EC, Walker GC, Siede W, Wood RD, Schultz RA, Ellenberger T. (2006). DNA Repair and Mutagenesis, part 3. ASM Press. 2nd ed. [31] Bakkenist, CJ; Kastan, MB. (2003). "DNA damage activates ATM through intermolecular autophosphorylation and dimer dissociation". Nature 421 (6922): 499506. doi:10.1038/nature01368. PMID12556884. [32] Wei, Qingyi; Lei Li, David Chen (2007). DNA Repair, Genetic Instability, and Cancer. World Scientific. ISBN981-270-014-5. [33] Schonthal, Axel H. (2004). Checkpoint Controls and Cancer. Humana Press. ISBN1-58829-500-1. [34] Janion, C. (2001). "Some aspects of the SOS response system-a critical survey". Acta Biochim Pol. 48 (3): 599610. PMID11833768. [35] Erill, I; Campoy, S; Barb, J (2007). "Aeons of distress: an evolutionary perspective on the bacterial SOS response". FEMS Microbiol Rev. 31 (6): 637656. doi:10.1111/j.1574-6976.2007.00082.x. PMID17883408. [36] Schlacher, K; Pham, P; Cox, MM; Goodman, MF. (2006). "Roles of DNA Polymerase V and RecA Protein in SOS Damage-Induced Mutation". Chem. Rev 106 (2): 406419. doi:10.1021/cr0404951. PMID16464012. [37] Espejel, S; Martin, M; Klatt, P; Martin-Caballero, J; Flores, JM; Blasco, MA. (2004). "Shorter telomeres, accelerated ageing and increased lymphoma in DNA-PKcs-deficient mice". EMBO Rep 5 (5): 5039. doi:10.1038/sj.embor.7400127. PMC1299048. PMID15105825. [38] De Boer, J; Andressoo, JO; De Wit, J; Huijmans, J; Beems, RB; Van Steeg, H; Weeda, G; Van Der Horst, GT et al. (2002). "Premature aging in mice deficient in DNA repair and transcription". Science 296 (5571): 12769. doi:10.1126/science.1070174. PMID11950998. [39] Dolle, ME; Busuttil, RA; Garcia, AM; Wijnhoven, S; van Drunen, E; Niedernhofer, LJ; der Horst, G; Hoeijmakers, JH et al. (2006). "Increased genomic instability is not a prerequisite for shortened life span in DNA repair deficient mice". Mutation Research 596 (12): 2235. doi:10.1016/j.mrfmmm.2005.11.008. PMID16472827. [40] Kobayashi, Y; Narumi, I; Satoh, K; Funayama, T; Kikuchi, M; Kitayama, S; Watanabe, H. (2004). "Radiation response mechanisms of the extremely radioresistant bacterium Deinococcus radiodurans". Biol Sci Space 18 (3): 1345. PMID15858357. [41] Spindler, SR. (2005). "Rapid and reversible induction of the longevity, anticancer and genomic effects of caloric restriction". Mech Ageing Dev 126 (9): 9606. doi:10.1016/j.mad.2005.03.016. PMID15927235. [42] Tissenbaum, HA; Guarente, L. (2001). "Increased dosage of a sir-2 gene extends life span in Caenorhabditis elegans". Nature 410 (6825): 22730. doi:10.1038/35065638. PMID11242085. [43] Cohen, HY; Miller, C; Bitterman, KJ; Wall, NR; Hekking, B; Kessler, B; Howitz, KT; Gorospe, M et al. (2004). "Calorie restriction promotes mammalian cell survival by inducing the SIRT1 deacetylase". Science 305 (5682): 3902. doi:10.1126/science.1099196. PMID15205477. [44] Cabelof, DC; Yanamadala, S; Raffoul, JJ; Guo, Z; Soofi, A; Heydari, AR. (2003). "Caloric restriction promotes genomic stability by induction of base excision repair and reversal of its age-related decline". DNA Repair (Amst.) 2 (3): 295307. doi:10.1016/S1568-7864(02)00219-7. PMID12547392. [45] Stuart, JA; Karahalil, B; Hogue, BA; Souza-Pinto, NC; Bohr, VA. (2004). "Mitochondrial and nuclear DNA base excision repair are affected differently by caloric restriction". FASEB J 18 (3): 5957. doi:10.1096/fj.03-0890fje. PMID14734635. [46] Walker, DW; McColl, G; Jenkins, NL; Harris, J; Lithgow, GJ. (2000). "Evolution of lifespan in C. elegans". Nature 405 (6784): 2967. doi:10.1038/35012693. PMID10830948. [47] Cromie, GA; Connelly, JC; Leach, DR (2001). "Recombination at double-strand breaks and DNA ends: conserved mechanisms from phage to humans". Mol Cell. 8 (6): 116374. doi:10.1016/S1097-2765(01)00419-1. PMID11779493. [48] O'Brien, PJ. (2006). "Catalytic promiscuity and the divergent evolution of DNA repair enzymes". Chem Rev 106 (2): 72052. doi:10.1021/cr040481v. PMID16464022.

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DNA repair
[49] Maresca, B; Schwartz, JH (2006). "Sudden origins: a general mechanism of evolution based on stress protein concentration and rapid environmental change". Anat Rec B New Anat. 289 (1): 3846. doi:10.1002/ar.b.20089. PMID16437551.

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External links
Roswell Park Cancer Institute DNA Repair Lectures (http://asajj.roswellpark.org/huberman/DNA_Repair/ DNA_Repair.htm) A comprehensive list of Human DNA Repair Genes (http://www.cgal.icnet.uk/DNA_Repair_Genes.html) 3D structures of some DNA repair enzymes (http://www.biochem.ucl.ac.uk/bsm/xtal/teach/repair/tibs3. html) Human DNA repair diseases (http://www.scielo.br/scielo.php?pid=S0100-84551997000400032& script=sci_arttext&tlng=en) DNA repair special interest group (http://tango01.cit.nih.gov/sig/home.taf?_function=main& SIGInfo_SIGID=32) DNA Repair (http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/D/DNArepair.html) DNA Damage and DNA Repair (http://www.benbest.com/lifeext/aging.html#dna) Segmental Progeria (http://www.benbest.com/lifeext/aging.html#progeria) DNA-damage repair; the good, the bad, and the ugly (https://www.researchgate.net/publication/ 5565866_DNA-damage_repair_the_good_the_bad_and_the_ugly)

Oncogenes
An oncogene is a gene that has the potential to cause cancer.[1] In tumor cells, they are often mutated or expressed at high levels.[2] An oncogene is a gene found in the chromosomes of tumor cells whose activation is associated with the initial and continuing conversion of normal cells into cancer cells. Most normal cells undergo a programmed form of death (apoptosis). Activated oncogenes can cause those cells that ought to die to survive and proliferate instead.[3] Most oncogenes require an additional step, such as mutations in another gene, or environmental factors, such as viral infection, to cause cancer. Since the 1970s, dozens of oncogenes have been identified in human cancer. Many cancer drugs target the proteins encoded by oncogenes.[2] [4]
[5] [6]

Proto-oncogene
A proto-oncogene is a normal gene that can become an oncogene due to mutations or increased expression. The resultant protein may be termed an oncoprotein.[7] Proto-oncogenes code for proteins that help to regulate cell growth and differentiation. Proto-oncogenes are often involved in signal transduction and execution of mitogenic signals, usually through their protein products. Upon activation, a proto-oncogene (or its product) becomes a tumor-inducing agent, an oncogene.[8] Examples of proto-oncogenes include RAS, WNT, MYC, ERK, and TRK. The MYC gene is implicated in Burkitt's Lymphoma, which starts when a chromosomal translocation moves an enhancer sequence within the vicinity of the myc gene. The myc gene codes for widely used transcription factors. When the enhancer sequence is wrongly placed, these transcription factors are produced at much higher rates. Another example of an oncogene is the Bcr-Abl gene found on the Philadelphia Chromosome, a piece of genetic material seen in Chronic Myelogenous Leukemia caused by the translocation of pieces from chromosomes 9 and 22. Bcr-Abl codes for a receptor tyrosine kinase, which is constitutively active, leading to uncontrolled cell proliferation.

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Activation
The proto-oncogene can become an oncogene by a relatively small modification of its original function. There are three basic activation types: A mutation within a proto-oncogene can cause a change in the protein structure, causing an increase in protein (enzyme) activity a loss of regulation An increase in protein concentration, caused by an increase of protein expression (through misregulation) an increase of protein (mRNA) stability, prolonging its existence and thus its activity in the cell a gene duplication (one type of chromosome abnormality), resulting in an increased amount of protein in the cell A chromosomal translocation (another type of chromosome abnormality), causing an increased gene expression in the wrong cell type or at wrong times the expression of a constitutively active hybrid protein. This type of aberration in a dividing stem cell in the bone marrow leads to adult leukemia The expression of oncogenes can be regulated by microRNAs (miRNAs), small RNAs 21-25 nucleotides in length that control gene expression by downregulating them.[9] Mutations in such microRNAs (known as oncomirs) can lead to activation of oncogenes.[10] Antisense messenger RNAs could theoretically be used to block the effects of oncogenes.

Classification
There are several systems for classifying oncogenes,[11] [12] but there is not yet a widely accepted standard. They are sometimes grouped both spatially (moving from outside the cell inwards) and chronologically (parallelling the "normal" process of signal transduction). There are several categories that are commonly used:
Category Growth factors, or mitogens Examples c-Sis Description Usually secreted by specialized cells to induce cell proliferation in themselves, nearby cells, or distant cells. An oncogene may cause a cell to secrete growth factors even though it does not normally do so. It will thereby induce its own uncontrolled proliferation (autocrine loop), and proliferation of neighboring cells. It may also cause production of growth hormones in other parts of the body. Kinases add phosphate groups to other proteins to turn them on or off. Receptor kinases add phosphate groups to receptor proteins at the surface of the cell (which receive protein signals from outside the cell and transmit them to the inside of the cell). Tyrosine kinases add phosphate groups to the amino acid tyrosine in the target protein. They can cause cancer by turning the receptor permanently on (constitutively), even without signals from outside the cell. -

Receptor tyrosine kinases

epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), and vascular endothelial growth factor receptor (VEGFR), HER2/neu

Cytoplasmic tyrosine kinases

Src-family, Syk-ZAP-70 family, and BTK family of tyrosine kinases, the Abl gene in CML - Philadelphia chromosome Raf kinase, and cyclin-dependent kinases (through overexpression).

Cytoplasmic Serine/threonine kinases and their regulatory subunits

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Ras protein Ras is a small GTPase that hydrolyses GTP into GDP and phosphate. Ras is activated by growth factor signaling (i.e., EGF, TGFalpha) and acting like a binary switch (on/off) in growth signaling pathways. Downstream effectors of Ras include Raf, MEK, MEKK, MAPK, ERK, most of which in turn regulate genes that mediate cell proliferation. -They regulate transcription of genes that induce cell proliferation.

Regulatory GTPases

Transcription factors

myc gene

Conversion of proto-oncogenes
There are two mechanisms by which proto-oncogenes can be converted to cellular oncogenes: Quantitative: Tumor formation is induced by an increase in the absolute number of proto-oncogene products or by its production in inappropriate cell types. Qualitative: Conversion from proto-oncogene to transforming gene (c-onc) with changes in the nucleotide sequence that are responsible for the acquisition of the new properties.[13]

History
The first oncogene was discovered in 1970 and was termed src (pronounced sarc as in sarcoma). Src was in fact first discovered as an oncogene in a chicken retrovirus. Experiments performed by Dr G. Steve Martin of the University of California, Berkeley demonstrated that the SRC was indeed the oncogene of the virus. The first nucleotide sequence of v-src was sequenced 1980 by A.P. Czernilofsky et al. (Nature Vol 287, pp 198-203). In 1976 Drs. Dominique Stehelin, J. Michael Bishop and Harold E. Varmus of the University of California, San Francisco demonstrated that oncogenes were activated proto-oncogenes, found in many organisms including humans. For this discovery Bishop and Varmus were awarded the Nobel Prize in Physiology or Medicine in 1989.[14]

References
[1] Wilbur, Beth, editor. The World of the Cell, Becker, W.M., et al., 7th ed. San Francisco, CA; 2009. [2] Kimball's Biology Pages. (http:/ / users. rcn. com/ jkimball. ma. ultranet/ BiologyPages/ O/ Oncogenes. html) "Oncogenes" Free full text [3] The Nobel Prize in Physiology or Medicine 2002. (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 2002/ illpres/ implications. html) Illustrated presentation. [4] Croce CM (Jan 2008). "Oncogenes and cancer" (http:/ / content. nejm. org/ cgi/ content/ full/ 358/ 5/ 502). N Engl J Med. 358 (5): 50211. doi:10.1056/NEJMra072367. PMID18234754. . [5] Yokota J (Mar 2000). "Tumor progression and metastasis" (http:/ / carcin. oxfordjournals. org/ cgi/ content/ full/ 21/ 3/ 497). Carcinogenesis. 21 (3): 497503. doi:10.1093/carcin/21.3.497. PMID10688870. . [6] The Nobel Prize in Physiology or Medicine 1989 (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1989/ press. html) to J. Michael Bishop and Harold E. Varmus for their discovery of "the cellular origin of retroviral oncogenes". [7] Chapter 20 - NEOPLASMS OF THE THYROID - in: Mitchell, Richard Sheppard; Kumar, Vinay; Abbas, Abul K.; Fausto, Nelson. Robbins Basic Pathology. Philadelphia: Saunders. ISBN1-4160-2973-7. 8th edition. [8] Todd R, Wong DT (1999). "Oncogenes". Anticancer Res. 19 (6A): 472946. PMID10697588. [9] Negrini M, Ferracin M, Sabbioni S, Croce CM (Jun 2007). "MicroRNAs in human cancer: from research to therapy". J Cell Sci. 120 (Pt 11): 183340. doi:10.1242/jcs.03450. PMID17515481. [10] Esquela-Kerscher A, Slack FJ (Apr 2006). "Oncomirs - microRNAs with a role in cancer". Nat Rev Cancer 6 (4): 25969. doi:10.1038/nrc1840. PMID16557279. [11] THE Medical Biochemistry Page (http:/ / web. indstate. edu/ thcme/ mwking/ oncogene. html#classes) [12] Classification of Oncogene Function (http:/ / www. qub. ac. uk/ cm/ pat/ education/ Carcinogenesis/ tsld014. htm) [13] Emery, Alan E. H.; Mueller, Robert Francis; Young, Ian T.; Ian D., MD Young (2001). "Oncogene". Emery's elements of medical genetics. Edinburgh: Churchill Livingstone. ISBN0-443-07125-X. [14] Nobel Prize in Physiology or Medicine for 1989 jointly to J. Michael Bishop and Harold E. Varmus for their discovery of "the cellular origin of retroviral oncogenes". (http:/ / nobelprize. org/ nobel_prizes/ medicine/ laureates/ 1989/ press. html) Press Release.

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External links
Drosophila Oncogenes and Tumor Suppressors - The Interactive Fly (http://www.sdbonline.org/fly/aignfam/ tumorsup.htm) List of web sites - oncogenes tables (http://microb230.med.upenn.edu/protocols/cancergenes.html)

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RNA synthesis and processing

Transcription
Transcription is the process of creating a complementary RNA copy of a sequence of DNA.[1] Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes. During transcription, a DNA sequence is read by RNA polymerase, which produces a complementary, antiparallel RNA strand. As opposed to DNA replication, transcription results in an RNA complement that includes uracil (U) in all instances where thymine (T) would have occurred in a DNA complement. Transcription can be explained easily in 4 or 5 steps, each moving like a wave along the DNA. 1. RNA Polymerase unwinds/"unzips" the DNA by breaking the hydrogen bonds between complementary nucleotides. 2. RNA Polymerase adds matching RNA nucleotides that are paired with complementary DNA bases. 3. RNA sugar-phosphate backbone forms with assistance from RNA polymerase. 4. Hydrogen bonds of the untwisted RNA+DNA helix break, freeing the newly synthesized RNA strand. 5. If the cell has a nucleus, the RNA is further processed (addition of a 3' poly-A tail and a 5' cap) and exits through to the cytoplasm through the nuclear pore complex. Transcription is the first step leading to gene expression. The stretch of DNA transcribed into an RNA molecule is called a transcription unit and encodes at least one gene. If the gene transcribed encodes a protein, the result of transcription is messenger RNA (mRNA), which will then be used to create that protein via the process of translation. Alternatively, the transcribed gene may encode for either ribosomal RNA (rRNA) or transfer RNA (tRNA), other components of the protein-assembly process, or other ribozymes. A DNA transcription unit encoding for a protein contains not only the sequence that will eventually be directly translated into the protein (the coding sequence) but also regulatory sequences that direct and regulate the synthesis of that protein. The regulatory sequence before (upstream from) the coding sequence is called the five prime untranslated region (5'UTR), and the sequence following (downstream from) the coding sequence is called the three prime untranslated region (3'UTR). Transcription has some proofreading mechanisms, but they are fewer and less effective than the controls for copying DNA; therefore, transcription has a lower copying fidelity than DNA replication.[2] As in DNA replication, DNA is read from 3' 5' during transcription. Meanwhile, the complementary RNA is created from the 5' 3' direction. This means its 5' end is created first in base pairing. Although DNA is arranged as two antiparallel strands in a double helix, only one of the two DNA strands, called the template strand, is used for transcription. This is because RNA is only single-stranded, as opposed to double-stranded DNA. The other DNA strand is called the coding (lagging) strand, because its sequence is the same as the newly created RNA transcript (except for the substitution of uracil for thymine). The use of only the 3' 5' strand eliminates the need for the Okazaki fragments seen in DNA replication. Transcription is divided into 5 stages: pre-initiation, initiation, promoter clearance, elongation and termination.

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Major steps
Pre-initiation
In eukaryotes, RNA polymerase, and therefore the initiation of transcription, requires the presence of a core promoter sequence in the DNA. Promoters are regions of DNA that promote transcription and, in eukaryotes, are found at -30, -75, and -90 base pairs upstream from the transcription start site (abbreviated to TSS). Core promoters are sequences within the promoter that are essential for transcription initiation. RNA polymerase is able to bind to core promoters in the presence of various specific transcription factors. The most characterized type of core promoter in eukaryotes is a short DNA sequence known as a TATA box, found 25-30 base pairs upstream from the TSS. The TATA box, as a core promoter, is the binding site for a transcription factor known as TATA-binding protein (TBP), which is itself a subunit of another transcription factor, called Transcription Factor II D (TFIID). After TFIID binds to the TATA box via the TBP, five more transcription factors and RNA polymerase combine around the TATA box in a series of stages to form a preinitiation complex. One transcription factor, DNA helicase, has helicase activity and so is involved in the separating of opposing strands of double-stranded DNA to provide access to a single-stranded DNA template. However, only a low, or basal, rate of transcription is driven by the preinitiation complex alone. Other proteins known as activators and repressors, along with any associated coactivators or corepressors, are responsible for modulating transcription rate. Thus, preinitiation complex contains: 1. Core Promoter Sequence 2. Transcription Factors 3. DNA Helicase 4. RNA Polymerase 5. Activators and Repressors The transcription preinitiation in archaea is, in essence, homologous to that of eukaryotes, but is much less complex.[3] The archaeal preinitiation complex assembles at a TATA-box binding site; however, in archaea, this complex is composed of only RNA polymerase II, TBP, and TFB (the archaeal homologue of eukaryotic transcription factor II B (TFIIB)).[4] [5]

Initiation
In bacteria, transcription begins with the binding of RNA polymerase to the promoter in DNA. RNA polymerase is a core enzyme consisting of five subunits: 2 subunits, 1 subunit, 1 ' subunit, and 1 subunit. At the start of initiation, the core enzyme is Simple diagram of transcription initiation. RNAP = RNA polymerase associated with a sigma factor that aids in finding the appropriate -35 and -10 base pairs downstream of promoter sequences.[6] When the sigma factor and RNA polymerase combine, they form a holoenzyme. Transcription initiation is more complex in eukaryotes. Eukaryotic RNA polymerase does not directly recognize the core promoter sequences. Instead, a collection of proteins called transcription factors mediate the binding of RNA polymerase and the initiation of transcription. Only after certain transcription factors are attached to the promoter does the RNA polymerase bind to it. The completed assembly of transcription factors and RNA polymerase bind to the promoter, forming a transcription initiation complex. Transcription in the archaea domain is similar to transcription in eukaryotes.[7]

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Promoter clearance
After the first bond is synthesized, the RNA polymerase must clear the promoter. During this time there is a tendency to release the RNA transcript and produce truncated transcripts. This is called abortive initiation and is common for both eukaryotes and prokaryotes.[8] Abortive initiation continues to occur until the factor rearranges, resulting in the transcription elongation complex (which gives a 35 bp moving footprint). The factor is released before 80 nucleotides of mRNA are synthesized.[9] Once the transcript reaches approximately 23 nucleotides, it no longer slips and elongation can occur. This, like most of the remainder of transcription, is an energy-dependent process, consuming adenosine triphosphate (ATP). Promoter clearance coincides with phosphorylation of serine 5 on the carboxy terminal domain of RNA Pol in eukaryotes, which is phosphorylated by TFIIH.

Elongation
One strand of the DNA, the template strand (or noncoding strand), is used as a template for RNA synthesis. As transcription proceeds, RNA polymerase traverses the template Simple diagram of transcription elongation strand and uses base pairing complementarity with the DNA template to create an RNA copy. Although RNA polymerase traverses the template strand from 3' 5', the coding (non-template) strand and newly-formed RNA can also be used as reference points, so transcription can be described as occurring 5' 3'. This produces an RNA molecule from 5' 3', an exact copy of the coding strand (except that thymines are replaced with uracils, and the nucleotides are composed of a ribose (5-carbon) sugar where DNA has deoxyribose (one less oxygen atom) in its sugar-phosphate backbone). Unlike DNA replication, mRNA transcription can involve multiple RNA polymerases on a single DNA template and multiple rounds of transcription (amplification of particular mRNA), so many mRNA molecules can be rapidly produced from a single copy of a gene. Elongation also involves a proofreading mechanism that can replace incorrectly incorporated bases. In eukaryotes, this may correspond with short pauses during transcription that allow appropriate RNA editing factors to bind. These pauses may be intrinsic to the RNA polymerase or due to chromatin structure.

Termination
Bacteria use two different strategies for transcription termination. In Rho-independent transcription termination, RNA transcription stops when the newly synthesized RNA molecule forms a G-C-rich hairpin loop followed by a run of Us. When Simple diagram of transcription termination the hairpin forms, the mechanical stress breaks the weak rU-dA bonds, now filling the DNA-RNA hybrid. This pulls the poly-U transcript out of the active site of the RNA polymerase, in effect, terminating transcription. In the "Rho-dependent" type of termination, a protein factor called "Rho" destabilizes the interaction between the template and the mRNA, thus releasing the newly synthesized mRNA from the elongation complex.[10] Transcription termination in eukaryotes is less understood but involves cleavage of the new transcript followed by template-independent addition of As at its new 3' end, in a process called polyadenylation.[11]

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Measuring and detecting transcription

Transcription can be measured and detected in a variety of ways: Nuclear Run-on assay: measures the relative abundance of newly formed transcripts RNase protection assay and ChIP-Chip of RNAP: detect active transcription sites RT-PCR: measures the absolute abundance of total or nuclear RNA levels, which may however differ from transcription rates DNA microarrays: measures the relative abundance of the global total or nuclear RNA levels; however, these may differ from transcription rates In situ hybridization: detects the presence of a transcript MS2 tagging: by incorporating RNA stem loops, such as MS2, into a gene, these become incorporated into newly synthesized RNA. The stem loops can then be detected using a fusion of GFP and the Electron micrograph of the ribosomal transcription process. The forming mRNA strands MS2 coat protein, which has a high affinity, sequence-specific are visible as branches from the main DNA interaction with the MS2 stem loops. The recruitment of GFP to the strand. site of transcription is visualised as a single fluorescent spot. This remarkable new approach has revealed that transcription occurs in discontinuous bursts, or pulses (see Transcriptional bursting). With the notable exception of in situ techniques, most other methods provide cell population averages, and are not capable of detecting this fundamental property of genes.[12] Northern blot: the traditional method, and until the advent of RNA-Seq, the most quantitative RNA-Seq: applies next-generation sequencing techniques to sequence whole transcriptomes, which allows the measurement of relative abundance of RNA, as well as the detection of additional variations such as fusion genes, post-translational edits and novel splice sites

Transcription factories
Active transcription units are clustered in the nucleus, in discrete sites called transcription factories or euchromatin. Such sites can be visualized by allowing engaged polymerases to extend their transcripts in tagged precursors (Br-UTP or Br-U) and immuno-labeling the tagged nascent RNA. Transcription factories can also be localized using fluorescence in situ hybridization or marked by antibodies directed against polymerases. There are ~10,000 factories in the nucleoplasm of a HeLa cell, among which are ~8,000 polymerase II factories and ~2,000 polymerase III factories. Each polymerase II factory contains ~8 polymerases. As most active transcription units are associated with only one polymerase, each factory usually contains ~8 different transcription units. These units might be associated through promoters and/or enhancers, with loops forming a cloud around the factor.

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History
A molecule that allows the genetic material to be realized as a protein was first hypothesized by Franois Jacob and Jacques Monod. RNA synthesis by RNA polymerase was established in vitro by several laboratories by 1965; however, the RNA synthesized by these enzymes had properties that suggested the existence of an additional factor needed to terminate transcription correctly. In 1972, Walter Fiers became the first person to actually prove the existence of the terminating enzyme. Roger D. Kornberg won the 2006 Nobel Prize in Chemistry "for his studies of the molecular basis of eukaryotic transcription".[13]

Reverse transcription
Some viruses (such as HIV, the cause of AIDS), have the ability to transcribe RNA into DNA. HIV has an RNA genome that is duplicated into DNA. The resulting DNA can be merged with the DNA genome of the host cell. The main enzyme responsible for synthesis of DNA from an RNA template is called reverse transcriptase. In the case of HIV, reverse transcriptase is responsible for synthesizing a complementary DNA strand (cDNA) to the viral RNA genome. An associated enzyme, ribonuclease H, digests the RNA strand, and reverse transcriptase synthesises a complementary strand of DNA Scheme of reverse transcription to form a double helix DNA structure. This cDNA is integrated into the host cell's genome via another enzyme (integrase) causing the host cell to generate viral proteins that reassemble into new viral particles. In HIV, subsequent to this, the host cell undergoes programmed cell death, apoptosis of T cells.[14] However, in other retroviruses, the host cell remains intact as the virus buds out of the cell. Some eukaryotic cells contain an enzyme with reverse transcription activity called telomerase. Telomerase is a reverse transcriptase that lengthens the ends of linear chromosomes. Telomerase carries an RNA template from which it synthesizes DNA repeating sequence, or "junk" DNA. This repeated sequence of DNA is important because, every time a linear chromosome is duplicated, it is shortened in length. With "junk" DNA at the ends of chromosomes, the shortening eliminates some of the non-essential, repeated sequence rather than the protein-encoding DNA sequence farther away from the chromosome end. Telomerase is often activated in cancer cells to enable cancer cells to duplicate their genomes indefinitely without losing important protein-coding DNA sequence. Activation of telomerase could be part of the process that allows cancer cells to become immortal. However, the true in vivo significance of telomerase has still not been empirically proven.

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References
[1] MedicineNet.com. "Transcription definition" (http:/ / www. medterms. com/ script/ main/ art. asp?articlekey=5835). . Retrieved 11 October 2009. [2] Berg J, Tymoczko JL, Stryer L (2006). Biochemistry (6th ed.). San Francisco: W. H. Freeman. ISBN0716787245. [3] Littlefield, O., Korkhin, Y., and Sigler, P.B. (1999). "The structural basis for the oriented assembly of a TBP/TFB/promoter complex". PNAS 96 (24): 1366813673. doi:10.1073/pnas.96.24.13668. PMC24122. PMID10570130. [4] Hausner, W; Thomm, M (2001). "Events during Initiation of Archaeal Transcription: Open Complex Formation and DNA-Protein Interactions". Journal of Bacteriology 183 (10): 30253031. doi:10.1128/JB.183.10.3025-3031.2001. PMC95201. PMID11325929. [5] Qureshi, SA; Bell, SD; Jackson, SP (1997). "Factor requirements for transcription in the archaeon Sulfolobus shibatae". EMBO Journal 16 (10): 29272936. doi:10.1093/emboj/16.10.2927. PMC1169900. PMID9184236. [6] Raven, Peter H. (2011). Biology: ninth edition. New York: McGraw-Hill. pp.278301. ISBN9780073532226. [7] Mohamed Ouhammouch, Robert E. Dewhurst, Winfried Hausner, Michael Thomm, and E. Peter Geiduschek (2003). "Activation of archaeal transcription by recruitment of the TATA-binding protein". Proceedings of the National Academy of Sciences of the United States of America 100 (9): 50975102. doi:10.1073/pnas.0837150100. PMC154304. PMID12692306. [8] Goldman, R.; Ebright, H.; Nickels, E. (May 2009). "Direct detection of abortive RNA transcripts in vivo". Science 324 (5929): 927928. Bibcode2009Sci...324..927G. doi:10.1126/science.1169237. ISSN0036-8075. PMC2718712. PMID19443781. [9] Dvir, A (Sep 2002). "Promoter escape by RNA polymerase II". Biochimica et Biophysica Acta 1577 (2): 208223. ISSN0006-3002. PMID12213653. [10] Richardson J. Rho-dependent termination and ATPases in transcript termination. Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression. 2002;1577(2):251-260. Available at: http:/ / dx. doi. org/ 10. 1016/ S0167-4781(02)00456-6 [Accessed March 5, 2011]. [11] Lykke-Andersen S, Jensen TH. Overlapping pathways dictate termination of RNA polymerase II transcription. Biochimie. 2007;89(10):1177-82. Available at: http:/ / dx. doi. org/ 10. 1016/ j. biochi. 2007. 05. 007 [Accessed August 5, 2010]. [12] Raj, A. and van Oudenaarden, A. (2008). Nature, nurture, or chance: stochastic gene expression and its consequences. Cell 135, 216-26. [13] "Chemistry 2006" (http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 2006/ ). Nobel Foundation. . Retrieved 2007-03-29. [14] Kolesnikova I. N. (2000 .). "Some patterns of apoptosis mechanism during HIV-infection" (http:/ / www. dissercat. com/ content/ nekotorye-osobennosti-mekhanizmov-apoptoza-pri-vich-infektsii) (in ru). Dissertation. . Retrieved 2011-02-20.

External links
Interactive Java simulation of transcription initiation. (http://cmol.nbi.dk/models/dynamtrans/dynamtrans. html) From Center for Models of Life (http://cmol.nbi.dk/) at the Niels Bohr Institute. Interactive Java simulation of transcription interference--a game of promoter dominance in bacterial virus. (http:/ /cmol.nbi.dk/models/dna/rnap.html) From Center for Models of Life (http://cmol.nbi.dk/) at the Niels Bohr Institute. Biology animations about this topic under Chapter 15 and Chapter 18 (http://highered.mcgraw-hill.com/sites/ dl/free/0072437316/120060/ravenanimation.html) Virtual Cell Animation Collection, Introducing Transcription (http://vcell.ndsu.nodak.edu/animations/ transcription/index.htm)

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Regulation of gene expression

Gene modulation redirects here. For information on therapeutic regulation of gene expression, see therapeutic gene modulation. For vocabulary, see Glossary of gene expression terms Regulation of gene expression (or gene regulation) includes the processes that cells and viruses use to regulate the way that the information in genes is turned into gene products. Although a functional gene product can be an RNA, the majority of known mechanisms regulate protein coding genes. Any step of the gene's expression may be modulated, from DNA-RNA transcription to the post-translational modification of a protein. Gene regulation is essential for viruses, prokaryotes and eukaryotes as it increases the versatility and adaptability of an organism by allowing Diagram showing at which stages in the DNA-mRNA-protein pathway expression can be the cell to express protein when needed. Although in 1951, Barbara controlled McClintock showed interaction between two genetic loci, Activator (Ac) and Dissociator (Ds), the first discovery of a gene regulation system is widely considered to be the identification in 1961 of the lac operon, discovered by Jacques Monod, in which protein involved in lactose metabolism are expressed by E. coli only in the presence of lactose and absence of glucose. Furthermore, gene regulation drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms where the different types of cells may possess different gene expression profiles though they all possess the same genome sequence.

Regulated stages of gene expression

Any step of gene expression may be modulated, from the DNA-RNA transcription step to post-translational modification of a protein. The following is a list of stages where gene expression is regulated, the most extensively utilised point is Transcription Initiation: Chromatin domains Transcription Post-transcriptional modification RNA transport Translation mRNA degradation

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Modification of DNA
In eukaryotes, the accessibility of large regions of DNA can depend on its chromatin structure, which can be altered as a result of histone modifications directed by DNA methylation, ncRNA, or DNA-binding protein. Hence these modifications may up or down regulate the expression of gene. Certain of these modifications that regulate gene expression are inheritable and are referred to as epigenetic regulation.

Chemical
Methylation of DNA is a common method of gene silencing. DNA is typically methylated by methyltransferase enzymes on cytosine nucleotides in a CpG dinucleotide sequence (also called "CpG islands" when densely clustered). Analysis of the pattern of methylation in a given region of DNA (which can be a promoter) can be achieved through a method called bisulfite mapping. Methylated cytosine residues are unchanged by the treatment, whereas unmethylated ones are changed to uracil. The differences are analyzed by DNA sequencing or by methods developed to quantify SNPs, such as Pyrosequencing (Biotage) or MassArray (Sequenom), measuring the relative amounts of C/T at the CG dinucleotide. Abnormal methylation patterns are thought to be involved in oncogenesis.

Structural
Transcription of DNA is dictated by its structure. In general, the density of its packing is indicative of the frequency of transcription. Octameric protein complexes called nucleosomes are responsible for the amount of supercoiling of DNA, and these complexes can be temporarily modified by processes such as phosphorylation or more permanently modified by processes such as methylation. Such modifications are considered to be responsible for more or less permanent changes in gene expression levels. Histone acetylation is also an important process in transcription. Histone acetyltransferase enzymes (HATs) such as CREB-binding protein also dissociate the DNA from the histone complex, allowing transcription to proceed. Often, DNA methylation and histone deacetylation work together in gene silencing. The combination of the two seems to be a signal for DNA to be packed more densely, lowering gene expression.

Regulation of transcription
Regulation of transcription controls when transcription occurs and how much RNA is created. Transcription of a gene by RNA polymerase can be regulated by at least five mechanisms: Specificity factors alter the specificity of RNA polymerase for a given promoter or set of promoters, making it more or less likely to bind to them (i.e., sigma factors used in prokaryotic transcription). Repressors bind to non-coding sequences on the DNA strand that are close to or overlapping the promoter region, impeding RNA polymerase's progress along the strand, thus impeding the expression of the gene. General transcription factors position RNA polymerase at the start of a protein-coding sequence and then release the polymerase to transcribe the mRNA. Activators enhance the interaction between RNA polymerase and a particular promoter, encouraging the expression of the gene. Activators do this by increasing the attraction of RNA polymerase for the promoter, through interactions with subunits of the RNA polymerase or indirectly by changing the structure of the DNA. Enhancers are sites on the DNA helix that are bound to by activators in order to loop the DNA bringing a specific promoter to the initiation complex. Enhancers are much more common in eukaryote than prokaryotes, where only a few examples exist (to date).[1]

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Post-transcriptional regulation
After the DNA is transcribed and mRNA is formed, there must be some sort of regulation on how much the mRNA is translated into proteins. Cells do this by modulating the capping, splicing, addition of a Poly(A) Tail, the sequence-specific nuclear export rates, and, in several contexts, sequestration of the RNA transcript. These processes occur in eukaryotes but not in prokaryotes. This modulation is a result of a protein or transcript that, in turn, is regulated and may have an affinity for certain sequences.

Regulation of translation
The translation of mRNA can also be controlled by a number of mechanisms, mostly at the level of initiation. Recruitment of the small ribosomal subunit can indeed be modulated by mRNA secondary structure, antisense RNA binding, or protein binding. In both prokaryotes and eukaryotes, a large number of RNA binding proteins exist, which often are directed to their target sequence by the secondary structure of the transcript, which may change depending on certain conditions, such as temperature or presence of a ligand (aptamer). Some transcripts act as ribozymes and self-regulate their expression.

Examples of gene regulation

Enzyme induction is a process in which a molecule (e.g., a drug) induces (i.e., initiates or enhances) the expression of an enzyme. The induction of heat shock proteins in the fruit fly Drosophila melanogaster. The Lac operon is an interesting example of how gene expression can be regulated. Viruses, despite having only a few genes, possess mechanisms to regulate their gene expression, typically into an early and late phase, using collinear systems regulated by anti-terminators (lambda phage) or splicing modulators (HIV).

Developmental biology
A large number of studied regulatory systems come from developmental biology. Examples include: The colinearity of the Hox gene cluster with their nested antero-posterior patterning It has been speculated that pattern generation of the hand (digits - interdigits) The gradient of Sonic hedgehog (secreted inducing factor) from the zone of polarizing activity in the limb, which creates a gradient of active Gli3, which activates Gremlin, which inhibits BMPs also secreted in the limb, resulting in the formation of an alternating pattern of activity as a result of this reaction-diffusion system. Somitogenesis is the creation of segments (somites) from a uniform tissue (Pre-somitic Mesoderm, PSM). They are formed sequentially from anterior to posterior. This is achieved in amniotes possibly by means of two opposing gradients, Retinoic acid in the anterior (wavefront) and Wnt and Fgf in the posterior, coupled to an oscillating pattern (segmentation clock) composed of FGF + Notch and Wnt in antiphase.[2] Sex determination in the soma of a Drosophila requires the sensing of the ratio of autosomal genes to sex chromosome-encoded genes, which results in the production of sexless splicing factor in females, resulting in the female isoform of doublesex.[3]

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Circuitry
Up-regulation and down-regulation
Up-regulation is a process that occurs within a cell triggered by a signal (originating internal or external to the cell), which results in increased expression of one or more genes and as a result the protein(s) encoded by those genes. On the converse, down-regulation is a process resulting in decreased gene and corresponding protein expression. Up-regulation occurs, for example, when a cell is deficient in some kind of receptor. In this case, more receptor protein is synthesized and transported to the membrane of the cell and, thus, the sensitivity of the cell is brought back to normal, reestablishing homeostasis. Down-regulation occurs, for example, when a cell is overstimulated by a neurotransmitter, hormone, or drug for a prolonged period of time, and the expression of the receptor protein is decreased in order to protect the cell (see also tachyphylaxis).

Inducible vs. repressible systems

Gene Regulation can be summarized by the response of the respective system: Inducible systems - An inducible system is off unless there is the presence of some molecule (called an inducer) that allows for gene expression. The molecule is said to "induce expression". The manner by which this happens is dependent on the control mechanisms as well as differences between prokaryotic and eukaryotic cells. Repressible systems - A repressible system is on except in the presence of some molecule (called a corepressor) that suppresses gene expression. The molecule is said to "repress expression". The manner by which this happens is dependent on the control mechanisms as well as differences between prokaryotic and eukaryotic cells.

Theoretical circuits
Repressor/Inducer: an activation of a sensor results in the change of expression of a gene negative feedback: the gene product downregulates its own production directly or indirectly, which can result in keeping transcript levels constant/proportional to a factor inhibition of run-away reactions when coupled with a positive feedback loop creating an oscillator by taking advantage in the time delay of transcription and translation, given that the mRNA and protein half-life is shorter positive feedback: the gene product upregulates its own production directly or indirectly, which can result in signal amplification bistable switches when two genes inhibit each other and both have positive feedback pattern generation

Methods
In general, most experiments investigating differential expression used whole cell extracts of RNA, called steady-state levels, to determine which genes changed and by how much they did. These are, however, not informative of where the regulation has occurred and may actually mask conflicting regulatory processess (see post-transcriptional regulation), but it is still the most commonly analysed (QPCR and DNA microarray). When studying gene expression, there are several methods to look at the various stages. In eukaryotes these include: The local chromatin environment of the region can be determined by ChIP-chip analysis by pulling down RNA Polymerase II, Histone 3 modifications, Trithorax-group protein, Polycomb-group protein, or any other DNA-binding element to which a good antibody is available. Epistatic interactions can be investigated by synthetic genetic array analysis

Regulation of gene expression Due to post-transcriptional regulation, transcription rates and total RNA levels differ significantly. To measure the transcription rates nuclear run-on assays can be done and newer high-throughput methods are being developed, using thiol labelling instead of radioactivity.[4] Only 5% of the RNA polymerised in the nucleus actually exists,[5] and not only introns, abortive products, and non-sense transcripts are degradated. Therefore, the differences in nuclear and cytoplasmic levels can be see by separating the two fractions by gentle lysis.[6] Alternative splicing can be analysed with a splicing array or with a tiling array (see DNA microarray). All in vivo RNA is complexed as RNPs. The quantity of transcripts bound to specific protein can be also analysed by RIP-Chip. For example, DCP2 will give an indication of sequestered protein; ribosome-bound gives and indication of transcripts active in transcription (although it should be noted that a more dated method, called polysome fractionation, is still popular in some labs) Protein levels can be analysed by Mass spectrometry, which can be compared only to QPCR data, as microarray data is relative and not absolute. RNA and protein degradation rates are measured by means of transcription inhibitors (actinomycin D or -amanitin) or translation inhibitors (Cycloheximide), respectively.

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References
[1] Austin S, Dixon R (June 1992). "The prokaryotic enhancer binding protein NTRC has an ATPase activity which is phosphorylation and DNA dependent". EMBO J. 11 (6): 221928. PMC556689. PMID1534752. [2] Dequant ML, Pourqui O. Segmental patterning of the vertebrate embryonic axis. Nat Rev Genet. 2008 May;9(5):370-82. PMID 18414404 [3] Gilbert SF (2003). Developmental biology, 7th ed., Sunderland, Mass: Sinauer Associates, 656. ISBN 0-87893-258-5. [4] Cheadle C, Fan J, Cho-Chung YS, Werner T, Ray J, Do L, Gorospe M, Becker KG (2005). "Control of gene expression during T cell activation: alternate regulation of mRNA transcription and mRNA stability". BMC Genomics 6: 75. doi:10.1186/1471-2164-6-75. PMC1156890. PMID15907206. [5] Jackson DA, Pombo A, Iborra F (2000). "The balance sheet for transcription: an analysis of nuclear RNA metabolism in mammalian cells" (http:/ / www. fasebj. org/ cgi/ content/ abstract/ 14/ 2/ 242). FASEB J. 14 (2): 24254. PMID10657981. . [6] Schwanekamp JA, Sartor MA, Karyala S, Halbleib D, Medvedovic M, Tomlinson CR (2006). "Genome-wide analyses show that nuclear and cytoplasmic RNA levels are differentially affected by dioxin". Biochim. Biophys. Acta 1759 (89): 388402. doi:10.1016/j.bbaexp.2006.07.005. PMID16962184.

Further reading
Latchman, David S. (2005). Gene regulation: a eukaryotic perspective (http://books.google.com/ books?id=4x3ZzLNyfDsC). Psychology Press. ISBN9780415365109.

External links
Cellular Darwinism (http://www.scitopics.com/ Stochastic_gene_expression_and_cell_differentiation_during_embryo_development.html) MeSH Regulation of Gene Expression (http://www.nlm.nih.gov/cgi/mesh/2011/MB_cgi?mode=& term=Regulation+of+Gene+Expression)

385

Protein synthesis and modifications

Translation
In molecular biology and genetics, translation is the third stage of protein biosynthesis (part of the overall process of gene expression). In translation, messenger RNA (mRNA) produced by transcription is decoded by the ribosome to produce a specific amino acid chain, or polypeptide, that will later fold into an active protein. In Bacteria, translation occurs in the cell's cytoplasm, where the large and small subunits of the ribosome are located, and bind to the mRNA. In Eukaryotes, translation occurs across the membrane of the endoplasmic reticulum in a process called vectorial synthesis. The ribosome Diagram showing the translation of mRNA and the synthesis of proteins by a ribosome. facilitates decoding by inducing the binding of tRNAs with complementary anticodon sequences to that of the mRNA. The tRNAs carry specific amino acids that are chained together into a polypeptide as the mRNA passes through and is "read" by the ribosome in a fashion reminiscent to that of a stock ticker and ticker tape. In many instances, the entire ribosome/mRNA complex will bind to the outer membrane of the rough endoplasmic reticulum and release the nascent protein polypeptide inside for later vesicle transport and secretion outside of the cell. Many types of transcribed RNA, such as transfer RNA, ribosomal RNA, and small nuclear RNA, do not undergo translation into proteins. Translation proceeds in four phases: activation, initiation, elongation and termination (all describing the growth of the amino acid chain, or polypeptide that is the product of translation). Amino acids are brought to ribosomes and assembled into proteins. In activation, the correct amino acid is covalently bonded to the correct transfer RNA (tRNA). The amino acid is joined by its carboxyl group to the 3' OH of the tRNA by an ester bond. When the tRNA has an amino acid linked to it, it is termed "charged". Initiation involves the small subunit of the ribosome binding to the 5' end of mRNA with the help of initiation factors (IF). Termination of the polypeptide happens when the A site of the ribosome faces a stop codon (UAA, UAG, or UGA). No tRNA can recognize or bind to this codon. Instead, the stop codon induces the binding of a release factor protein that prompts the disassembly of the entire ribosome/mRNA complex. A number of antibiotics act by inhibiting translation; these include anisomycin, cycloheximide, chloramphenicol, tetracycline, streptomycin, erythromycin, and puromycin, among others. Prokaryotic ribosomes have a different structure from that of eukaryotic ribosomes, and thus antibiotics can specifically target bacterial infections without any detriment to a eukaryotic host's cells.

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Basic mechanisms
Further information: Prokaryotic translation and Eukaryotic translation The basic process of protein production is addition of one amino acid at a time to the end of a protein. This operation is performed by a ribosome. The choice of amino acid type to add is determined by an mRNA molecule. Each amino acid added is matched to a three nucleotide subsequence of the mRNA. For each such triplet possible, only one particular amino acid type is accepted. The successive amino acids added to the chain are matched to successive nucletide triplets in the mRNA. In this way the sequence of nucletides in the template mRNA chain determines the sequence of amino acids in the generated amino acid chain.[1] The mRNA carries genetic information encoded as a ribonucleotide sequence from the chromosomes to the ribosomes. The ribonucleotides are "read" by translational machinery in a sequence of nucleotide triplets called codons. Each of those triplets codes for a specific amino acid.
Tertiary structure of tRNA. CCA tail in orange, Acceptor stem in purple, D arm in red, Anticodon arm in blue with Anticodon in black, T arm in green.

The ribosome molecules translate this code to a specific sequence of amino acids. The ribosome is a multisubunit structure containing rRNA and proteins. It is the "factory" where amino acids are assembled into proteins. tRNAs are small noncoding RNA chains (74-93 nucleotides) that transport amino acids to the ribosome. tRNAs have a site for amino acid attachment, and a site called an anticodon. The anticodon is an RNA triplet complementary to the mRNA triplet that codes for their cargo amino acid. Aminoacyl tRNA synthetase (an enzyme) catalyzes the bonding between specific tRNAs and the amino acids that their anticodon sequences call for. The product of this reaction is an aminoacyl-tRNA molecule. This aminoacyl-tRNA travels inside the ribosome, where mRNA codons are matched through complementary base pairing to specific tRNA anticodons. The ribosome has three sites for tRNA to bind. They are the aminoacyl site (abbreviated A), the peptidyl site (abbreviated P) and the exit site (abbreviated E). With respect to the mRNA, the three sites are oriented 5to 3 E-P-A, because ribosomes move in a 3 to 5 fashion. The A site binds the incoming tRNA with the complementary codon on the mRNA. The P site holds the tRNA with the growing polypeptide chain. The E site holds the tRNA without its amino acid. When an aminoacyl-tRNA initially binds to its corresponding codon on the mRNA, it is in the A site. Then, a peptide bond forms between the amino acid of the tRNA in the A site and the amino acid of the charged tRNA in the P site. The growing polypeptide chain is transferred to the tRNA in the A site. Translocation occurs, moving the tRNA in the P site, now without an amino acid, to the E site; the tRNA that was in the A site, now charged with the polypeptide chain, is moved to the P site. The tRNA in the E site leaves and another aminoacyl-tRNA enters the A site to repeat the process. [2] After the new amino acid is added to the chain, the energy provided by the hydrolysis of a GTP bound to the translocase EF-G (in prokaryotes) and eEF-2 (in eukaryotes) moves the ribosome down one codon towards the 3' end. The energy required for translation of proteins is significant. For a protein containing n amino acids, the number of high-energy Phosphate bonds required to translate it is 4n-1 . The rate of translation varies; it is significantly higher in prokaryotic cells (up to 17-21 amino acid residues per second) than in eukaryotic cells (up to 6-9 amino acid residues per second).[3]

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Genetic code
Whereas other aspects such as the 3D structure, called tertiary structure, of protein can only be predicted using sophisticated algorithms, the amino acid sequence, called primary structure, can be determined solely from the nucleic acid sequence with the aid of a translation table. This approach may not give the correct amino acid composition of the protein, in particular if unconventional amino acids such as selenocysteine are incorporated into the protein, which is coded for by a conventional stop codon in combination with a downstream hairpin (SElenoCysteine Insertion Sequence, or SECIS). There are many computer programs capable of translating a DNA/RNA sequence into a protein sequence. Normally this is performed using the Standard Genetic Code; many bioinformaticians have written at least one such program at some point in their education. However, few programs can handle all the "special" cases, such as the use of the alternative initiation codons. For instance, the rare alternative start codon CTG codes for Methionine when used as a start codon, and for Leucine in all other positions. Example: Condensed translation table for the Standard Genetic Code (from the NCBI Taxonomy webpage [4]). AAs Starts Base1 Base2 Base3 = = = = = FFLLSSSSYY**CC*WLLLLPPPPHHQQRRRRIIIMTTTTNNKKSSRRVVVVAAAADDEEGGGG ---M---------------M---------------M---------------------------TTTTTTTTTTTTTTTTCCCCCCCCCCCCCCCCAAAAAAAAAAAAAAAAGGGGGGGGGGGGGGGG TTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGG TCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAG

Translation tables
Even when working with ordinary Eukaryotic sequences such as the Yeast genome, it is often desired to be able to use alternative translation tablesnamely for translation of the mitochondrial genes. Currently the following translation tables are defined by the NCBI Taxonomy Group [5] for the translation of the sequences in GenBank: 1: 2: 3: 4: 5: 6: 9: 10: 11: 12: 13: 14: 15: 16: 21: 22: 23: = The Standard The Vertebrate Mitochondrial Code The Yeast Mitochondrial Code The Mold, Protozoan, and Coelenterate Mitochondrial Code and the Mycoplasma/Spiroplasma Code The Invertebrate Mitochondrial Code The Ciliate, Dasycladacean and Hexamita Nuclear Code The Echinoderm and Flatworm Mitochondrial Code The Euplotid Nuclear Codecbn dxh The Bacterial and Plant Plastid Code The Alternative Yeast Nuclear Code The Ascidian Mitochondrial Code The Alternative Flatworm Mitochondrial Code Blepharisma Nuclear Code Chlorophycean Mitochondrial Code Trematode Mitochondrial Code Scenedesmus obliquus mitochondrial Code Thraustochytrium Mitochondrial Code

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References
[1] Neill, Campbell (1996). Biology; Fourth edition. The Benjamin/Cummings Publishing Company. p.309,310. ISBN0-8053-1940-9. [2] Griffiths, Anthony (2008). "9". Introduction to Genetic Analysis (9th ed.). New York: W.H. Freeman and Company. pp.335-339. ISBN978-0-7167-6887-6. [3] Ross JF, Orlowski M (February 1982). "Growth-rate-dependent adjustment of ribosome function in chemostat-grown cells of the fungus Mucor racemosus". J. Bacteriol. 149 (2): 6503. PMC216554. PMID6799491. [4] http:/ / www. ncbi. nlm. nih. gov/ Taxonomy/ Utils/ wprintgc. cgi?mode=c [5] http:/ / www. ncbi. nlm. nih. gov/ Taxonomy/

Further reading
Champe, Pamela C; Harvey, Richard A; Ferrier, Denise R (2004). Lippincott's Illustrated Reviews: Biochemistry (3rd ed.). Hagerstwon, MD: Lippincott Williams & Wilkins. ISBN0-7817-2265-9. Cox, Michael; Nelson, David R.; Lehninger, Albert L (2005). Lehninger principles of biochemistry (4th ed.). San Francisco...: W.H. Freeman. ISBN0-7167-4339-6. Malys N, McCarthy JEG (2010). "Translation initiation: variations in the mechanism can be anticipated". Cellular and Molecular Life Sciences 68 (6): 9911003. doi:10.1007/s00018-010-0588-z. PMID21076851.

External links
Virtual Cell Animation Collection: Introducing Translation (http://vcell.ndsu.nodak.edu/animations/ translation/index.htm) mRNA to Amino Acid translator (http://www.bluetulip.org/discrepancy/aminotrans.php)

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Posttranslational modification
Posttranslational modification (PTM) is the chemical modification of a protein after its translation. It is one of the later steps in protein biosynthesis, and thus gene expression, for many proteins. A protein (also called a polypeptide) is a chain of amino acids. During protein synthesis, 20 different amino acids can be incorporated to become a protein. After translation, the posttranslational modification of amino acids extends the range of functions of the protein by attaching it to other biochemical functional groups (such as acetate, phosphate, various lipids and carbohydrates), changing the chemical nature of an amino acid (e.g. citrullination), or making structural changes (e.g. formation of disulfide bridges). Also, enzymes may remove amino acids from the amino end of the protein, or cut the peptide chain in the middle. For instance, the peptide hormone insulin is cut twice after disulfide bonds are formed, and a propeptide is removed from the middle of the chain; the resulting protein consists of two polypeptide chains connected by disulfide bonds. Also, most nascent polypeptides start with the amino acid methionine because the "start" codon on mRNA also codes for this amino acid. This amino acid is usually taken off during post-translational modification. Other modifications, like phosphorylation, are part of common mechanisms for controlling the behavior of a protein, for instance activating or inactivating an enzyme.
The bottom of this diagram shows the modification of primary structure of insulin, as described.

Post-translational modification of proteins is detected by mass spectrometry or Eastern blotting.

Posttranslational modification

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PTMs involving addition of functional groups

PTMs involving addition by an enzyme in vivo
PTMs involving addition of hydrophobic groups for membrane localization myristoylation, attachment of myristate, a C14 saturated acid palmitoylation, attachment of palmitate, a C16 saturated acid isoprenylation or prenylation, the addition of an isoprenoid group (e.g. farnesol and geranylgeraniol) farnesylation geranylgeranylation glypiation, glycosylphosphatidylinositol (GPI) anchor formation via an amide bond to C-terminal tail PTMs involving addition of cofactors for enhanced enzymatic activity lipoylation, attachment of a lipoate (C8) functional group flavin moiety (FMN or FAD) may be covalently attached
The genetic code diagram

[1]

showing the amino acid residues as target of modification.

heme C attachment via thioether bonds with cysteins phosphopantetheinylation, the addition of a 4'-phosphopantetheinyl moiety from coenzyme A, as in fatty acid, polyketide, non-ribosomal peptide and leucine biosynthesis retinylidene Schiff base formation PTMs involving unique modifications of translation factors diphthamide formation (on a histidine found in eEF2) ethanolamine phosphoglycerol attachment (on glutamte found in eEF1)[2] hypusine formation (on conserved lysine of eIF5A (eukaryotic) and aIF5A (archeal)) PTMs involving addition of smaller chemical groups acylation, e.g. O-acylation (esters), N-acylation (amides), S-acylation (thioesters) acetylation, the addition of an acetyl group, either at the N-terminus [3] of the protein or at lysine residues.[4] See also histone acetylation.[5] [6] The reverse is called deacetylation. formylation alkylation, the addition of an alkyl group, e.g. methyl, ethyl methylation the addition of a methyl group, usually at lysine or arginine residues. The reverse is called demethylation. amide bond formation

Posttranslational modification amidation at C-terminus amino acid addition arginylation, a tRNA-mediation addition polyglutamylation, covalent linkage of glutamic acid residues to the N-terminus of tubulin and some other proteins.[7] (See tubulin polyglutamylase) polyglycylation, covalent linkage of one to more than 40 glycine residues to the tubulin C-terminal tail gamma-carboxylation dependent on Vitamin K[8] glycosylation, the addition of a glycosyl group to either asparagine, hydroxylysine, serine, or threonine, resulting in a glycoprotein. Distinct from glycation, which is regarded as a nonenzymatic attachment of sugars. polysialylation, addition of polysialic acid, PSA, to NCAM ADP-ribosylation hydroxylation iodination (e.g. of thyroglobulin) oxidation phosphate ester (O-linked) or phosphoramidate (N-linked) formation phosphorylation, the addition of a phosphate group, usually to serine, threonine, and tyrosine (O-linked), or histidine (N-linked) adenylylation, the addition of an adenylyl moiety, usually to tyrosine (O-linked), or histidine and lysine (N-linked) pyroglutamate formation S-glutathionylation S-nitrosylation sulfation, the addition of a sulfate group to a tyrosine. selenoylation (co-translational incorporation of selenium in selenoproteins)

391

PTMs involving non-enzymatic additions in vivo

glycation, the addition of a sugar molecule to a protein without the controlling action of an enzyme.

PTMs involving non-enzymatic additions in vitro

biotinylation, acylation of conserved lysine residues with a biotin appendage pegylation

PTMs involving addition of other proteins or peptides

ISGylation, the covalent linkage to the ISG15 protein (Interferon-Stimulated Gene 15)[9] SUMOylation, the covalent linkage to the SUMO protein (Small Ubiquitin-related MOdifier)[10] ubiquitination, the covalent linkage to the protein ubiquitin. Neddylation, the covalent linkage to Nedd

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PTMs involving changing the chemical nature of amino acids

citrullination, or deimination, the conversion of arginine to citrulline deamidation, the conversion of glutamine to glutamic acid or asparagine to aspartic acid eliminylation, the conversion to an alkene by beta-elimination of phosphothreonine and phosphoserine, or dehydration of threonine and serine, as well as by decarboxylation of cysteine [11] carbamylation, the conversion of lysine to homocitrulline [12]

PTMs involving structural changes

disulfide bridges, the covalent linkage of two cysteine amino acids proteolytic cleavage, cleavage of a protein at a peptide bond racemization of proline by prolyl isomerase

Post-translational modification statistics

Recently statistics of each post-translational modification experimentally and putatively detected have been compiled using proteome-wide information from the Swiss-Prot database.[13] These statistics can be found at http:/ / selene.princeton.edu/PTMCuration/.

Case examples
Cleavage and formation of disulfide bridges during the production of insulin PTM of histones as regulation of transcription: RNA polymerase control by chromatin structure PTM of RNA polymerase II as regulation of transcription Cleavage of polypeptide chains [14] as crucial for lectin specificity

External links
List of posttranslational modifications in ExPASy [15] Statistics of each post-translational modification from the Swiss-Prot database [16] AutoMotif Server [17] - A Computational Protocol for Identification of Post-Translational Modifications in Protein Sequences [18] Functional analyses for site-specific phosphorylation of a target protein in cells [19] Detection of Post-Translational Modifications after high-accuracy MSMS [20]

References
[1] Gramatikoff K. in Abgent Catalog (2004-5) p.263 [2] Whiteheart SW, Shenbagamurthi P, Chen L, et al. (1989). "Murine elongation factor 1 alpha (EF-1 alpha) is posttranslationally modified by novel amide-linked ethanolamine-phosphoglycerol moieties. Addition of ethanolamine-phosphoglycerol to specific glutamic acid residues on EF-1 alpha.". J. Biol. Chem. 264 (24): 1433441. PMID2569467. [3] Polevoda B, Sherman F (2003). "N-terminal acetyltransferases and sequence requirements for N-terminal acetylation of eukaryotic proteins". J Mol Biol 325 (4): 595622. doi:10.1016/S0022-2836(02)01269-X. PMID12507466. [4] Yang XJ, Seto E (2008). "Lysine acetylation: codified crosstalk with other posttranslational modifications". Mol Cell 31 (4): 44961. doi:10.1016/j.molcel.2008.07.002. PMC2551738. PMID18722172. [5] Brtov E, Krejc J, Harnicarov A, Galiov G, Kozubek S (2008). "Histone modifications and nuclear architecture: a review". J Histochem Cytochem 56 (8): 71121. doi:10.1369/jhc.2008.951251. PMC2443610. PMID18474937. [6] Glozak MA, Sengupta N, Zhang X, Seto E (2005). "Acetylation and deacetylation of non-histone proteins". Gene 363: 1523. doi:10.1016/j.gene.2005.09.010. PMID16289629. [7] Edd B, Rossier J, Le Caer JP, Desbruyres E, Gros F, Denoulet P (1990). "Posttranslational glutamylation of alpha-tubulin". Science 247 (4938): 835. Bibcode1990Sci...247...83E. doi:10.1126/science.1967194. PMID1967194.

Posttranslational modification
[8] Walker CS, Shetty RP, Clark K, et al. (2001). "On a potential global role for vitamin K-dependent gamma-carboxylation in animal systems. Evidence for a gamma-glutamyl carboxylase in Drosophila". J. Biol. Chem. 276 (11): 776974. doi:10.1074/jbc.M009576200. PMID11110799. [9] Malakhova, Oxana A.; Yan, Ming; Malakhov, Michael P.; Yuan, Youzhong; Ritchie, Kenneth J.; Kim, Keun Il; Peterson, Luke F.; Shuai, Ke; and Dong-Er Zhang (2003). "Protein ISGylation modulates the JAK-STAT signaling pathway" (http:/ / www. genesdev. org/ cgi/ content/ full/ 17/ 4/ 455). Genes & Development 17 (4): 45560. doi:10.1101/gad.1056303. PMC195994. PMID12600939. . [10] Van G. Wilson (Ed.) (2004). Sumoylation: Molecular Biology and Biochemistry (http:/ / www. horizonpress. com/ hsp/ books/ sumo. html). Horizon Bioscience. ISBN 0-9545232-8-8. [11] Brennan DF, Barford D (2009). "Eliminylation: a post-translational modification catalyzed by phosphothreonine lyases". Trends in Biochemical Sciences 34 (3): 108114. doi:10.1016/j.tibs.2008.11.005. PMID19233656. [12] Mydel P, et al. (2010). "Carbamylation-dependent activation of T cells: a novel mechanism in the pathogenesis of autoimmune arthritis.". Journal of Immunology 184 (12): 68826890. doi:10.4049/ jimmunol.1000075. PMC2925534. PMID20488785. [13] Khoury, George A.; Baliban, Richard C.; and Christodoulos A. Floudas (2011). "Proteome-wide post-translational modification statistics: frequency analysis and curation of the swiss-prot database" (http:/ / www. nature. com/ srep/ 2011/ 110913/ srep00090/ full/ srep00090. html). Scientific Reports 1 (90). doi:10.1038/srep00090. . [14] http:/ / www. proteopedia. org/ wiki/ index. php/ 1tp8 [15] http:/ / www. uniprot. org/ docs/ ptmlist [16] http:/ / selene. princeton. edu/ PTMCuration/ [17] http:/ / ams2. bioinfo. pl/ [18] http:/ / www. natureprotocols. com/ 2007/ 03/ 23/ automotif_server_a_computation. php [19] http:/ / www. natureprotocols. com/ 2007/ 01/ 10/ functional_analyses_for_sitesp. php [20] http:/ / www. detectorvision. com/ deltaMasses. html

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Proteolysis
Proteolysis is the directed degradation (digestion) of proteins by cellular enzymes called proteases or by intramolecular digestion.

Purposes
Proteolysis is used by the cell for several purposes. They include: Removal of N-terminal methionine residues after translation. Removal of the signal sequence of peptides after their transport through a membrane Separation of viral proteins that were translated from a polycistronic mRNA Digestion of proteins from foods as a source of amino acids Conversion of predecessor-proteins (proenzymes, zymogens, prehormones) into their final structures. Degradation of unneeded or damaged proteins for example cyclins at different stages of the cell cycle.

Proteolysis is also used in research and diagnostic applications: In-gel digestion of proteins after separation by gel electrophoresis for the identification by mass spectrometry. Digestion of proteins in solution for proteome analysis by liquid chromatography-mass spectrometry (LC-MS).

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Examples
Examples of serine proteases include: trypsin chymotrypsin elastase papain

Venoms
Certain types of venom, such as those produced by venomous snakes, can also cause proteolysis. These venoms are, in fact, complex digestive fluids that begin their work outside of the body. Proteolytic venoms cause a wide range of toxic effects,[1] including effects that are: cytotoxic (cell-destroying) hemotoxic (blood-destroying) myotoxic (muscle-destroying) hemorrhagic (bleeding)

References
[1] Hayes WK. 2005. Research on Biological Roles and Variation of Snake Venoms. (http:/ / www. llu. edu/ llu/ grad/ natsci/ hayes/ research-c-venom. html?PHPSESSID=55842bf3eeb83dcdfec66c45b91925fc) Loma Linda University.

External links
Proteolysis (http://www.emedicinehealth.com/script/main/srchcont_dict.asp?src=Proteolysis) at eMedicine Dictionary Proteolysis MAP from Center on Proteolytic Pathways (http://www.proteolysis.org/)

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Proteasome
Proteasomes are very large protein complexes inside all eukaryotes and archaea, and in some bacteria. In eukaryotes, they are located in the nucleus and the cytoplasm.[1] The main function of the proteasome is to degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks peptide bonds. Enzymes that carry out such reactions are called proteases. Proteasomes are part of a major mechanism by which cells regulate the concentration of particular proteins and degrade misfolded proteins. The degradation process yields peptides of about seven to eight amino acids long, which can then be further degraded into amino acids and used in synthesizing new proteins.[2] Proteins are tagged for degradation with a small protein called ubiquitin. The tagging reaction is catalyzed by enzymes called ubiquitin ligases. Once a protein is tagged with a single ubiquitin molecule, this is a signal to other ligases to attach additional ubiquitin molecules. The result is a polyubiquitin chain that is bound by the proteasome, allowing it to degrade the tagged protein.[2] In structure, the proteasome is a cylindrical complex containing a "core" of four stacked rings around a central pore. Each ring is composed of seven individual proteins. The inner two rings are made of seven subunits that contain three to seven protease active sites. These sites are located on the interior surface of the rings, so that the target protein must enter the central pore before it is degraded. The outer two rings each contain seven subunits whose function is to maintain a "gate" through which proteins enter the barrel. These subunits are controlled by binding to "cap" structures or regulatory particles that recognize polyubiquitin tags attached to protein substrates and initiate the degradation process. The overall system of ubiquitination and proteasomal degradation is known as the ubiquitin-proteasome system. The proteasomal degradation pathway is essential for many cellular processes, including the cell cycle, the regulation of gene expression, and responses to oxidative stress. The importance of proteolytic degradation inside cells and the role of ubiquitin in proteolytic pathways was acknowledged in the award of the 2004 Nobel Prize in Chemistry to Aaron Ciechanover, Avram Hershko and Irwin Rose.[3]

Cartoon representation of a proteasome. Its active sites are sheltered inside the tube (blue). The caps (red; in this case, 11S regulatory particles) on the ends regulate entry into the destruction chamber, where the protein is degraded.

Discovery
Before the discovery of the ubiquitin proteasome system, protein degradation in cells was thought to rely mainly on lysosomes, membrane-bound organelles with acidic and protease-filled interiors
Top view of the proteasome above.

Proteasome that can degrade and then recycle exogenous proteins and aged or damaged organelles.[2] However, work by Alfred Goldberg in 1977 on ATP-dependent protein degradation in reticulocytes, which lack lysosomes, suggested the presence of a second intracellular degradation mechanism.[4] This was shown in 1978 to be composed of several distinct protein chains, a novelty among proteases at the time.[5] Later work on modification of histones led to the identification of an unexpected covalent modification of the histone protein by a bond between a lysine side chain of the histone and the C-terminal glycine residue of ubiquitin, a protein that had no known function.[6] It was then discovered that a previously identified protein associated with proteolytic degradation, known as ATP-dependent proteolysis factor 1 (APF-1), was the same protein as ubiquitin.[7] Later, the ATP-dependent proteolytic complex that was responsible for ubiquitin-dependent protein degradation was discovered and was called the 26S proteasome.[8] [9] Much of the early work leading up to the discovery of the ubiquitin proteasome system occurred in the late 1970s and early 1980s at the Technion in the laboratory of Avram Hershko, where Aaron Ciechanover worked as a graduate student. Hershko's year-long sabbatical in the laboratory of Irwin Rose at the Fox Chase Cancer Center provided key conceptual insights, though Rose later downplayed his role in the discovery.[10] The three shared the 2004 Nobel Prize in Chemistry for their work in discovering this system.[3] Although electron microscopy data revealing the stacked-ring structure of the proteasome became available in the mid-1980s,[11] the first structure of the proteasome core particle was not solved by X-ray crystallography until 1994.[12] As of 2006, no structure has been solved of the core particle in complex with the most common form of regulatory cap.

396

Structure and organization

The proteasome subcomponents are often referred to by their Svedberg sedimentation coefficient (denoted S). The most common form of the proteasome is known as the 26S proteasome, which is about 2000 kilodaltons (kDa) in molecular mass and contains one 20S core particle structure and two 19S regulatory caps. The core is hollow and provides an enclosed cavity in which proteins are degraded; openings at the two ends of the core allow the target protein to enter. Each end of the core particle associates with a 19S regulatory subunit that contains multiple ATPase active sites and ubiquitin binding sites; it is this structure that recognizes polyubiquitinated proteins and transfers them to the catalytic core. An alternative form of regulatory subunit called the 11S particle can associate with the core in essentially the same manner as the 19S particle; the 11S may play a role in degradation of foreign peptides such as those produced after infection by a virus.[13]

20S core particle

The number and diversity of subunits contained in the 20S core particle depends on the organism; the number of distinct and specialized subunits is larger in multicellular than unicellular organisms and larger in eukaryotes than in prokaryotes. All 20S particles consist of four stacked heptameric

A schematic diagram of the proteasome 20S core particle viewed from one side. The subunits that make up the outer two rings are shown in green, and the subunits that make up the inner two rings are shown in blue.

ring

structures

that

are

Proteasome

397

themselves composed of two different types of subunits; subunits are structural in nature, whereas subunits are predominantly catalytic. The outer two rings in the stack consist of seven subunits each, which serve as docking domains for the regulatory particles and the alpha subunits N-termini form a gate that blocks unregulated access of substrates to the interior cavity.[14] The inner two rings each consist of seven subunits and contain the protease active sites that perform the proteolysis reactions. The size of the proteasome is relatively conserved and is about 150 angstroms () by 115. The interior chamber is at most 53 wide, though the entrance can be as narrow as 13, suggesting that substrate proteins must be at least partially unfolded to enter.[15]

In archaea such as Thermoplasma acidophilum, all the and all the subunits are identical, whereas eukaryotic proteasomes such as those in yeast contain seven distinct types of each subunit. In mammals, the 1, 2, and 5 subunits are catalytic; although they share a common mechanism, they have three distinct substrate specificities considered chymotrypsin-like, trypsin-like, and peptidyl-glutamyl peptide-hydrolyzing (PHGH).[16] Alternative forms denoted 1i, 2i, and 5i can be expressed in hematopoietic cells in response to exposure to pro-inflammatory signals such as cytokines, in particular, interferon gamma. The proteasome assembled with these alternative subunits is known as the immunoproteasome, whose substrate specificity is altered relative to the normal proteasome.[15]

Top view of the same schematic, illustrating the seven-fold symmetry of the rings.

19S regulatory particle

The 19S particle in eukaryotes consists of 19 individual proteins and is divisible into two subassemblies, a 10-protein base that binds directly to the ring of the 20S core particle, and a 9-protein lid where polyubiquitin is bound. Six of the ten base proteins are ATPase subunits from the AAA Family, and an evolutionary homolog of these ATPases exists in archaea, called PAN (Proteasome-Activating Nucleotidase).[17] The association of the 19S and 20S particles requires the binding of ATP to the 19S ATPase subunits, and ATP hydrolysis is required for the assembled complex to degrade folded and ubiquitinated proteins. Note that only the step of substrate unfolding requires energy from ATP hydrolysis, while ATP-binding alone can support all the other steps required for protein degradation (e.g., complex assembly, gate opening, translocation, and proteolysis).[18] [19] In fact, ATP binding to the ATPases by itself supports the rapid degradation of unfolded proteins. However, while ATP hydrolysis is required for unfolding only, it is not yet clear whether this energy may be used in the coupling of some of these steps.[19] [20] As of 2011, the atomic structure of the 26S proteasome has not been solved, despite massive efforts to do so. Nevertheless, it is understood, in general, how the 19S associates with and regulates the 20S core particle.[14] [21] In fact, the 19S and 11S particles bind to the same sites in the rings of the 20S core particle although, they each induce gate opening by different mechanism.[14]

Regulation of the 20S by the 19S

The 19S regulatory particle is responsible for stimulating the 20S to degrade proteins. A primary function of the 19S regulatory ATPases is to open the gate in the 20S that blocks the entry of substrates into the degradation chamber.[22] The mechanism by which the proteasomal ATPase open this gate has been recently elucidated.[14] 20S gate opening, and thus substrate degradation, requires the C-termini of the proteasomal ATPases, which contains a specific motif (i.e., HbYX motif). The ATPases C-termini bind into pockets in the top of the 20S, and tether the ATPase complex to the 20S proteolytic complex, thus joining the substrate unfolding equipment with the 20S degradation machinery. Binding of these C-termini into these 20S pockets by themselves stimulates opening of the gate in the 20S in much

Proteasome the same way that a "key-in-a-lock" opens a door.[14] The precise mechanism by which this "key-in-a-lock" mechanism functions has been structurally elucidated.[23]

398

11S regulatory particle

20S proteasomes can also associate with a second type of regulatory particle, the 11S regulatory particle, a heptameric structure that does not contain any ATPases and can promote the degradation of short peptides but not of complete proteins. It is presumed that this is because the complex cannot unfold larger substrates. This structure is also known as PA28 or REG. The mechanisms by which it binds to the core particle through the C-terminal tails of its subunits and induces -ring conformational changes to open the 20S gate suggest a similar mechanism for the 19S particle.[24] The expression of the 11S particle is induced by interferon gamma and is responsible, in conjunction with the immunoproteasome subunits, for the generation of peptides that bind to the major histocompatibility complex.[13]

Assembly
The assembly of the proteasome is a complex process due to the number of subunits that must associate to form an active complex. The subunits are synthesized with N-terminal "propeptides" that are post-translationally modified during the assembly of the 20S particle to expose the proteolytic active site. The 20S particle is assembled from two half-proteasomes, each of which consists of a seven-membered pro- ring attached to a seven-membered ring. The association of the rings of the two half-proteasomes triggers threonine-dependent autolysis of the propeptides to expose the active site. These interactions are mediated mainly by salt bridges and hydrophobic interactions between conserved alpha helices whose disruption by mutation damages the proteasome's ability to assemble.[25] The assembly of the half-proteasomes, in turn, is initiated by the assembly of the subunits into their heptameric ring, forming a template for the association of the corresponding pro- ring. The assembly of subunits has not been characterized.[26] In general, less is known about the assembly and maturation of the 19S regulatory particles. They are believed to assemble as two distinct subcomponents, the ATPase-containing base and the ubiquitin-recognizing lid. The six ATPases in the base may assemble in a pairwise manner mediated by coiled-coil interactions.[27] The order in which the nineteen subunits of the regulatory particle are bound is a likely regulatory mechanism that prevents exposure of the active site before assembly is complete.[21]

Proteasome

399

The protein degradation process

Ubiquitylation and targeting
Proteins are targeted for degradation by the proteasome by covalent modification of a lysine residue that requires the coordinated reactions of three enzymes. In the first step, a ubiquitin-activating enzyme (known as E1) hydrolyzes ATP and adenylylates a ubiquitin molecule. This is then transferred to E1's active-site cysteine residue in concert with the adenylylation of a second ubiquitin.[28] This adenylylated ubiquitin is then transferred to a cysteine of a second enzyme, ubiquitin-conjugating enzyme (E2). In the last step, a Ribbon diagram of ubiquitin, the highly conserved protein that serves member of a highly diverse class of enzymes known as as a molecular tag targeting proteins for degradation by the proteasome ubiquitin ligases (E3) recognizes the specific protein to be ubiquitinated and catalyzes the transfer of ubiquitin from E2 to this target protein. A target protein must be labeled with at least four ubiquitin monomers (in the form of a polyubiquitin chain) before it is recognized by the proteasome lid.[29] It is therefore the E3 that confers substrate specificity to this system.[30] The number of E1, E2, and E3 proteins expressed depends on the organism and cell type, but there are many different E3 enzymes present in humans, indicating that there is a huge number of targets for the ubiquitin proteasome system. The mechanism by which a polyubiquitinated protein is targeted to the proteasome is not fully understood. Ubiquitin-receptor proteins have an N-terminal ubiquitin-like (UBL) domain and one or more ubiquitin-associated (UBA) domains. The UBL domains are recognized by the 19S proteasome caps and the UBA domains bind ubiquitin via three-helix bundles. These receptor proteins may escort polyubiquitinated proteins to the proteasome, though the specifics of this interaction and its regulation are unclear.[31] The ubiquitin protein itself is 76 amino acids long and was named due to its ubiquitous nature, as it has a highly conserved sequence and is found in all known eukaryotic organisms. The genes encoding ubiquitin in eukaryotes are arranged in tandem repeats, possibly due to the heavy transcription demands on these genes to produce enough ubiquitin for the cell. It has been proposed that ubiquitin is the slowest-evolving protein identified to date.[32]

Unfolding and translocation

After a protein has been ubiquitinated, it is recognized by the 19S regulatory particle in an ATP-dependent binding step.[19] The substrate protein must then enter the interior of the 20S particle to come in contact with the proteolytic active sites. Because the 20S particle's central channel is narrow and gated by the N-terminal tails of the ring subunits, the substrates must be at least partially unfolded before they enter the core. The passage of the unfolded substrate into the core is called The ubiquitination pathway translocation and necessarily occurs after deubiquitination.[19] However, the order in which substrates are deubiquitinated and unfolded is not yet clear.[33]

Proteasome Which of these processes is the rate-limiting step in the overall proteolysis reaction depends on the specific substrate; for some proteins, the unfolding process is rate-limiting, while deubiquitination is the slowest step for other proteins.[18] The extent to which substrates must be unfolded before translocation is not known, but substantial tertiary structure, and in particular nonlocal interactions such as disulfide bonds, are sufficient to inhibit degradation.[34] The gate formed by the subunits prevents peptides longer than about four residues from entering the interior of the 20S particle. The ATP molecules bound before the initial recognition step are hydrolyzed before translocation. While energy is needed for substrate unfolding, it is not required for translocation.[35] [36] The assembled 26S proteasome can degrade unfolded proteins in the presence of a non-hydrolyzable ATP analog, but cannot degrade folded proteins, indicating that energy from ATP hydrolysis is used for substrate unfolding.[35] Passage of the unfolded substrate through the opened gate occurs via facilitated diffusion if the 19S cap is in the ATP-bound state.[37] The mechanism for unfolding of globular proteins is necessarily general, but somewhat dependent on the amino acid sequence. Long sequences of alternating glycine and alanine have been shown to inhibit substrate unfolding decreasing the efficiency of proteasomal degradation; this results in the release of partially degraded byproducts, possibly due to the decoupling of the ATP hydrolysis and unfolding steps.[38] Such glycine-alanine repeats are also found in nature, for example in silk fibroin; in particular, certain Epstein-Barr virus gene products bearing this sequence can stall the proteasome, helping the virus propagate by preventing antigen presentation on the major histocompatibility complex.[39]

400

Proteolysis
The mechanism of proteolysis by the subunits of the 20S core particle is through a threonine-dependent nucleophilic attack. This mechanism may depend on an associated water molecule for deprotonation of the reactive threonine hydroxyl. Degradation occurs within the central chamber formed by the association of the two rings and normally does not release partially degraded products, instead reducing the substrate to short polypeptides typically 79 residues long, though they can range from 4 to 25 residues depending on the organism and substrate. The biochemical mechanism that determines product length is not fully characterized.[40] Although the three catalytic subunits have a common mechanism, they have slightly different substrate specificities, which are considered chymotrypsin-like, trypsin-like, and peptidyl-glutamyl peptide-hydrolyzing (PHGH)-like. These variations in specificity are the result of interatomic contacts with local residues near the active sites of each subunit. Each catalytic subunit also possesses a conserved lysine residue required for proteolysis.[16]

Although the proteasome normally produces very short peptide fragments, in some cases these products are themselves biologically active and functional molecules. Certain transcription factors regulating the expression of specific genes, including one component of the mammalian complex NF-B, are synthesized as inactive precursors whose ubiquitination and subsequent proteasomal degradation converts them to an

A cutaway view of the proteasome 20S core particle illustrating the locations of the active sites. The subunits are represented as green spheres and the subunits as protein backbones colored by individual polypeptide chain. The small pink spheres represent the location of the active-site threonine residue in each subunit. Light blue chemical structures are the inhibitor bortezomib bound to the active sites.

Proteasome active form. Such activity requires the proteasome to cleave the substrate protein internally: rather than processively degrading it from one terminus. It has been suggested that long loops on these proteins' surfaces serve as the proteasomal substrates and enter the central cavity, while the majority of the protein remains outside.[41] Similar effects have been observed in yeast proteins; this mechanism of selective degradation is known as regulated ubiquitin/proteasome dependent processing (RUP).[42]

401

Ubiquitin-independent degradation
Although most proteasomal substrates must be ubiquitinated before being degraded, there are some exceptions to this general rule, especially when the proteasome plays a normal role in the post-translational processing of the protein. The proteasomal activation of NF-B by processing p105 into p50 via internal proteolysis is one major example.[41] Some proteins that are hypothesized to be unstable due to intrinsically unstructured regions,[43] are degraded in a ubiquitin-independent manner. The most well-known example of a ubiquitin-independent proteasome substrate is the enzyme ornithine decarboxylase.[39] Ubiquitin-independent mechanisms targeting key cell cycle regulators such as p53 have also been reported, although p53 is also subject to ubiquitin-dependent degradation.[44] Finally, structurally abnormal, misfolded, or highly oxidized proteins are also subject to ubiquitin-independent and 19S-independent degradation under conditions of cellular stress.[45]

Evolution
The 20S proteasome is both ubiquitous and essential in eukaryotes. Some prokaryotes, including many archaea and the bacterial order Actinomycetales also share homologs of the 20S proteasome, whereas most bacteria possess heat shock genes hslV and hslU, whose gene products are a multimeric protease arranged in a two-layered ring and an ATPase.[46] The hslV protein has been hypothesized to resemble the likely ancestor of the 20S proteasome.[47] In general, HslV is not essential in bacteria, and not all bacteria possess it, whereas some protists possess both the 20S and the hslV systems.[46] Sequence analysis suggests that the catalytic subunits diverged earlier in evolution than the predominantly structural subunits. The assembled complex of hslV (blue) and hslU (red) In bacteria that express a 20S proteasome, the subunits have high from E. coli. This complex of heat shock proteins is sequence identity to archaeal and eukaryotic subunits, whereas thought to resemble the ancestor of the modern the sequence identity is much lower. The presence of 20S proteasome. proteasomes in bacteria may result from lateral gene transfer, while the diversification of subunits among eukaryotes is ascribed to multiple gene duplication events.[46]

Cell cycle control

Cell cycle progression is controlled by ordered action of cyclin-dependent kinases (CDKs), activated by specific cyclins that demarcate phases of the cell cycle. Mitotic cyclins, which persist in the cell for only a few minutes, have one of the shortest life spans of all intracellular proteins.[2] After a CDK-cyclin complex has performed its function, the associated cyclin is polyubiquitinated and destroyed by the proteasome, which provides directionality for the cell cycle. In particular, exit from mitosis requires the proteasome-dependent dissociation of the regulatory component cyclin B from the mitosis promoting factor complex.[48] In vertebrate cells, "slippage" through the mitotic checkpoint leading to premature M phase exit can occur despite the delay of this exit by the spindle checkpoint.[49]

Proteasome Earlier cell cycle checkpoints such as post-restriction point check between G1 phase and S phase similarly involve proteasomal degradation of cyclin A, whose ubiquitination is promoted by the anaphase promoting complex (APC), an E3 ubiquitin ligase.[50] The APC and the Skp1/Cul1/F-box protein complex (SCF complex) are the two key regulators of cyclin degradation and checkpoint control; the SCF itself is regulated by the APC via ubiquitination of the adaptor protein, Skp2, which prevents SCF activity before the G1-S transition.[51] Individual components of the 19S particle have their own regulatory roles. Gankyrin, a recently identified oncoprotein, is one of the 19S subcomponents that also tightly binds the cyclin-dependent kinase CDK4 and plays a key role in recognizing ubiquitinated p53, via its affinity for the ubiquitin ligase MDM2. Gankyrin is anti-apoptotic and has been shown to be overexpressed in some tumor cell types such as hepatocellular carcinoma.[52]

402

Regulation of plant growth

In plants, signaling by auxins, or phytohormones that order the direction and tropism of plant growth, induces the targeting of a class of transcription factor repressors known as Aux/IAA proteins for proteasomal degradation. These proteins are ubiquitinated by SCFTIR1, or SCF in complex with the auxin receptor TIR1. Degradation of Aux/IAA proteins derepresses transcription factors in the auxin-response factor (ARF) family and induces ARF-directed gene expression.[53] The cellular consequences of ARF activation depend on the plant type and developmental stage, but are involved in directing growth in roots and leaf veins. The specific response to ARF derepression is thought to be mediated by specificity in the pairing of individual ARF and Aux/IAA proteins.[54]

Apoptosis
Both internal and external signals can lead to the induction of apoptosis, or programmed cell death. The resulting deconstruction of cellular components is primarily carried out by specialized proteases known as caspases, but the proteasome also plays important and diverse roles in the apoptotic process. The involvement of the proteasome in this process is indicated by both the increase in protein ubiquitination, and of E1, E2, and E3 enzymes that is observed well in advance of apoptosis,[28] [55] [56] During apoptosis, proteasomes localized to the nucleus have also been observed to translocate to outer membrane blebs characteristic of apoptosis.[57] Proteasome inhibition has different effects on apoptosis induction in different cell types. In general, the proteasome is not required for apoptosis, although inhibiting it is pro-apoptotic in most cell types that have been studied. Apoptosis is mediated through disrupting the regulated degradation of pro-growth cell cycle proteins.[58] However, some cell lines in particular, primary cultures of quiescent and differentiated cells such as thymocytes and neurons are prevented from undergoing apoptosis on exposure to proteasome inhibitors. The mechanism for this effect is not clear, but is hypothesized to be specific to cells in quiescent states, or to result from the differential activity of the pro-apoptotic kinase JNK.[59] The ability of proteasome inhibitors to induce apoptosis in rapidly dividing cells has been exploited in several recently developed chemotherapy agents such as bortezomib and salinosporamide A.

Response to cellular stress

In response to cellular stresses such as infection, heat shock, or oxidative damage heat shock proteins that identify misfolded or unfolded proteins and target them for proteasomal degradation are expressed. Both Hsp27 and Hsp90chaperone proteins have been implicated in increasing the activity of the ubiquitin-proteasome system, though they are not direct participants in the process.[60] Hsp70, on the other hand, binds exposed hydrophobic patches on the surface of misfolded proteins and recruits E3 ubiquitin ligases such as CHIP to tag the proteins for proteasomal degradation.[61] The CHIP protein (carboxyl terminus of Hsp70-interacting protein) is itself regulated via inhibition of interactions between the E3 enzyme CHIP and its E2 binding partner.[62] Similar mechanisms exist to promote the degradation of oxidatively damaged proteins via the proteasome system. In particular, proteasomes localized to the nucleus are regulated by PARP and actively degrade inappropriately

Proteasome oxidized histones.[63] Oxidized proteins, which often form large amorphous aggregates in the cell, can be degraded directly by the 20S core particle without the 19S regulatory cap and do not require ATP hydrolysis or tagging with ubiquitin.[45] However, high levels of oxidative damage increases the degree of cross-linking between protein fragments, rendering the aggregates resistant to proteolysis. Larger numbers and sizes of such highly oxidized aggregates are associated with aging.[64] Dysregulation of the ubiquitin proteasome system may contribute to several neural diseases. It may lead to brain tumors such as astrocytomas.[65] In some of the late-onset neurodegenerative diseases that share aggregation of misfolded proteins as a common feature, such as Parkinson's disease and Alzheimer's disease, large insoluble aggregates of misfolded proteins can form and then result in neurotoxicity, through mechanisms that are not yet well understood. Decreased proteasome activity has been suggested as a cause of aggregation and Lewy body formation in Parkinson's.[66] This hypothesis is supported by the observation that yeast models of Parkinson's are more susceptible to toxicity from -synuclein, the major protein component of Lewy bodies, under conditions of low proteasome activity.[67] Impaired proteasomal activity may underlie cognitive disorders such as the autism spectrum disorders, and muscle and nerve diseases such as inclusion body myopathy.[65]

403

Role in the immune system

The proteasome plays a straightforward but critical role in the function of the adaptive immune system. Peptide antigens are displayed by the major histocompatibility complex class I (MHC) proteins on the surface of antigen-presenting cells. These peptides are products of proteasomal degradation of proteins originated by the invading pathogen. Although constitutively expressed proteasomes can participate in this process, a specialized complex composed of proteins whose expression is induced by interferon gamma produces peptides of the optimal size and composition for MHC binding. These proteins whose expression increases during the immune response include the 11S regulatory particle, whose main known biological role is regulating the production of MHC ligands, and specialized subunits called 1i, 2i, and 5i with altered substrate specificity. The complex formed with the specialized subunits is known as the immunoproteasome.[13] Another 5i variant subunit, 5t, is expressed in the thymus, leading to a thymus-specific "thymoproteasome" whose function is as yet unclear.[68] The strength of MHC class I ligand binding is dependent on the composition of the ligand C-terminus, as peptides bind by hydrogen bonding and by close contacts with a region called the "B pocket" on the MHC surface. Many MHC class I alleles prefer hydrophobic C-terminal residues, and the immunoproteasome complex is more likely to generate hydrophobic C-termini.[69] Due to its role in generating the activated form of NF-B, an anti-apoptotic and pro-inflammatory regulator of cytokine expression, proteasomal activity has been linked to inflammatory and autoimmune diseases. Increased levels of proteasome activity correlate with disease activity and have been implicated in autoimmune diseases including systemic lupus erythematosus and rheumatoid arthritis.[13] The proteasome is also involved in Intracellular antibody-mediated proteolysis of antibody bound virions. In this neutralisation pathway, TRIM21 (a protein of the tripartite motif family) binds with immunoglobulin G to direct the virion to the proteasome where it is degraded.[70]

Proteasome

404

Proteasome inhibitors
Proteasome inhibitors have effective anti-tumor activity in cell culture, inducing apoptosis by disrupting the regulated degradation of pro-growth cell cycle proteins.[58] This approach of selectively inducing apoptosis in tumor cells has proven effective in animal models and human trials. Bortezomib, a molecule developed by Millennium Pharmaceuticals and marketed as Velcade, is the first proteasome inhibitor to reach clinical use as a chemotherapy agent.[71] Bortezomib is used in the treatment of multiple myeloma.[72] Notably, multiple myeloma has been observed to result in increased proteasome levels in blood serum that decrease to normal levels in response to successful chemotherapy.[73] Studies in animals have indicated that bortezomib may also have clinically significant effects in pancreatic cancer.[74] [75] Preclinical and early clinical studies have been started to examine bortezomib's effectiveness in treating other B-cell-related cancers,[76] particularly some types of non-Hodgkin's lymphoma.[77] The molecule ritonavir, marketed as Norvir, was developed as a protease inhibitor and used to target HIV infection. However, it has been shown to inhibit proteasomes as well as free proteases; to be specific, the chymotrypsin-like activity of the proteasome is inhibited by ritonavir, while the trypsin-like activity is somewhat enhanced.[78] Studies in animal models suggest that ritonavir may have inhibitory effects on the growth of glioma cells.[79] Proteasome inhibitors have also shown promise in treating autoimmune diseases in animal models. For example, studies in mice bearing human skin grafts found a reduction in the size of lesions from psoriasis after treatment with a proteasome inhibitor.[80] Inhibitors also show positive effects in rodent models

Chemical structure of bortezomib, a proteasome inhibitor used in chemotherapy that is particularly effective against multiple myeloma

Bortezomib bound to the core particle in a yeast proteasome. The bortezomib molecule is in the center colored by atom type (carbon = pink, nitrogen = blue, oxygen = red, boron = yellow), surrounded by the local protein surface. The blue patch is the catalytic threonine residue whose activity is blocked by the presence of bortezomib.

of asthma.[81] Labeling and inhibition of the proteasome is also of interest in laboratory settings for both in vitro and in vivo study of proteasomal activity in cells. The most commonly used laboratory inhibitors are lactacystin, a natural product synthesized by Streptomyces bacteria,[59] and peptide MG132. Fluorescent inhibitors have also been developed to specifically label the active sites of the assembled proteasome.[82]

References
[1] Peters, Jan-Michael; Franke, Werner W.; Kleinschmidt, Jiirgen A. (March 1994). "Distinct 19 S and 20 S subcomplexes of the 26 S proteasome and their distribution in the nucleus and the cytoplasm" (http:/ / www. jbc. org/ cgi/ pmidlookup?view=long& pmid=8125997). The Journal of Biological Chemistry 269 (10): 770918. PMID8125997. . [2] Lodish H, Berk A, Matsudaira P, Kaiser CA, Krieger M, Scott MP, Zipursky SL, Darnell J (2004). "3". Molecular cell biology (5th ed.). New York: W.H. Freeman and CO. pp.6672. ISBN0-7167-4366-3. [3] Nobel Prize Committee (2004). "Nobel Prize Awardees in Chemistry, 2004" (http:/ / nobelprize. org/ nobel_prizes/ chemistry/ laureates/ 2004/ ). . Retrieved 2006-12-11. [4] Etlinger, Joseph D.; Goldberg, Alfred L. (January 1977). "A soluble ATP-dependent proteolytic system responsible for the degradation of abnormal proteins in reticulocytes". PNAS 74 (1): 548. doi:10.1073/pnas.74.1.54. PMC393195. PMID264694.

Proteasome
[5] Ciehanover, Aharon; Hod, Yaacov; Hershko, Avram (1978). "A heat-stable polypeptide component of an ATP-dependent proteolytic system from reticulocytes". Biochemical and Biophysical Research Communications 81 (4): 11005. doi:10.1016/0006-291X(78)91249-4. PMID666810. [6] Goldknopf, Ira L.; Busch, Harris (1977). "Isopeptide linkage between nonhistone and histone 2A polypeptides of chromosomal conjugate-protein A24". PNAS 74 (3): 8648. doi:10.1073/pnas.74.3.864. PMC430507. PMID265581. [7] Ciechanover, Aaron (2000). "Early work on the ubiquitin proteasome system, an interview with Aaron Ciechanover". Cell Death and Differentiation 12 (9): 116777. doi:10.1038/sj.cdd.4401691. PMID16094393. [8] Tanaka, Keiji; Waxman, Lloyd; Goldbert, Alfred L. (June 1983). "ATP serves two distinct roles in protein degradation in reticulocytes, one requiring and one independent of ubiquitin". The Journal of Cell Biology 96 (6): 15805. doi:10.1083/jcb.96.6.1580. PMC2112434. PMID6304111. [9] Hough, Ronald; Pratt, Gregory; Rechsteiner, Martin (June 1987). "Purification of two high molecular weight proteases from rabbit reticulocyte lysate" (http:/ / www. jbc. org/ cgi/ pmidlookup?view=long& pmid=3298229). The Journal of Biological Chemistry 262 (17): 830313. PMID3298229. . [10] Hershko A (2005). "Early work on the ubiquitin proteasome system, an interview with Avram Hershko". Cell Death Differ 12 (9): 11581161. doi:10.1038/sj.cdd.4401709. PMID16094391. [11] Kopp F et al. (1986). "Size and shape of the multicatalytic proteinase from rat skeletal muscle". Biochim Biophys Acta 872 (3): 25360. doi:10.1016/0167-4838(86)90278-5. PMID3524688. [12] Lwe J et al. (1995). "Crystal structure of the 20S proteasome from the archaeon T. acidophilum at 3.4 resolution". Science 268 (5210): 533539. doi:10.1126/science.7725097. PMID7725097. [13] Wang J, Maldonado MA (2006). "The Ubiquitin-Proteasome System and Its Role in Inflammatory and Autoimmune Diseases". Cell Mol Immunol 3 (4): 25561. PMID16978533. [14] Smith DM, Chang SC, Park S, Finley D, Cheng Y, Goldberg AL (September 2007). "Docking of the proteasomal ATPases' carboxyl termini in the 20S proteasome's alpha ring opens the gate for substrate entry". Mol. Cell 27 (5): 73144. doi:10.1016/j.molcel.2007.06.033. PMC2083707. PMID17803938. [15] Nandi D et al. (2006). "The ubiquitin-proteasome system". J Biosci 31 (1): 13755. doi:10.1007/BF02705243. PMID16595883. [16] Heinemeyer W et al. (1997). "The active sites of the eukaryotic 20 S proteasome and their involvement in subunit precursor processing". J Biol Chem 272 (40): 252009. doi:10.1074/jbc.272.40.25200. PMID9312134. [17] Zwickl P, Ng D, Woo KM, Klenk HP, Goldberg AL (September 1999). "An archaebacterial ATPase, homologous to ATPases in the eukaryotic 26 S proteasome, activates protein breakdown by 20 S proteasomes" (http:/ / www. jbc. org/ cgi/ pmidlookup?view=long& pmid=10473546). J. Biol. Chem. 274 (37): 2600814. doi:10.1074/jbc.274.37.26008. PMID10473546. . [18] Smith DM, Kafri G, Cheng Y, Ng D, Walz T, Goldberg AL (December 2005). "ATP binding to PAN or the 26S ATPases causes association with the 20S proteasome, gate opening, and translocation of unfolded proteins". Mol. Cell 20 (5): 68798. doi:10.1016/j.molcel.2005.10.019. PMID16337593. [19] Liu CW et al. (2006). "ATP binding and ATP hydrolysis play distinct roles in the function of 26S proteasome". Mol Cell 24 (1): 3950. doi:10.1016/j.molcel.2006.08.025. PMID17018291. [20] Lam YA et al. (2002). "A proteasomal ATPase subunit recognizes the polyubiquitin degradation signal". Nature 416 (6882): 7637. doi:10.1038/416763a. PMID11961560. [21] Sharon M et al. (2006). "Structural Organization of the 19S Proteasome Lid: Insights from MS of Intact Complexes". PLoS Biol 4 (8): e267. doi:10.1371/journal.pbio.0040267. PMC1523230. PMID16869714. [22] Khler A, Cascio P, Leggett DS, Woo KM, Goldberg AL, Finley D (June 2001). "The axial channel of the proteasome core particle is gated by the Rpt2 ATPase and controls both substrate entry and product release" (http:/ / linkinghub. elsevier. com/ retrieve/ pii/ S1097-2765(01)00274-X). Mol. Cell 7 (6): 114352. doi:10.1016/S1097-2765(01)00274-X. PMID11430818. . [23] Rabl J, Smith DM, Yu Y, Chang SC, Goldberg AL, Cheng Y (May 2008). "Mechanism of gate opening in the 20S proteasome by the proteasomal ATPases". Mol. Cell 30 (3): 3608. doi:10.1016/j.molcel.2008.03.004. PMID18471981. [24] Forster A et al. (2005). "The 1.9 Structure of a Proteasome-11S Activator Complex and Implications for Proteasome-PAN/PA700 Interactions". Mol Cell 18 (5): 58999. doi:10.1016/j.molcel.2005.04.016. PMID15916965. [25] Witt S et al. (2006). "Proteasome assembly triggers a switch required for active-site maturation". Structure 14 (7): 117988. doi:10.1016/j.str.2006.05.019. PMID16843899. [26] Kruger E et al. (2001). "20S proteasome biogenesis". Biochimie 83 (34): 28993. doi:10.1016/S0300-9084(01)01241-X. PMID11295488. [27] Gorbea C et al. (1999). "Assembly of the regulatory complex of the 26S proteasome". Mol Biol Rep 26 (12): 159. doi:10.1023/A:1006957802028. PMID10363641. [28] Haas AL, Warms JV, Hershko A, Rose IA (March 1982). "Ubiquitin-activating enzyme. Mechanism and role in protein-ubiquitin conjugation.". The Journal of biological chemistry 257 (5): 25438. PMID6277905. [29] Thrower JS et al. (2000). "Recognition of the polyubiquitin proteolytic signal". EMBO J 19 (1): 94102. doi:10.1093/emboj/19.1.94. PMC1171781. PMID10619848. [30] Risseeuw EP et al. (2003). "Protein interaction analysis of SCF ubiquitin E3 ligase subunits from Arabidopsis". Plant J 34 (6): 75367. doi:10.1046/j.1365-313X.2003.01768.x. PMID12795696. [31] Elsasser S, Finley D (2005). "Delivery of ubiquitinated substrates to protein-unfolding machines". Nat Cell Biol 7 (8): 7429. doi:10.1038/ncb0805-742. PMID16056265.

405

Proteasome
[32] Sharp PM, Li WH (1987). "Ubiquitin genes as a paradigm of concerted evolution of tandem repeats". J Mol Evol 25 (1): 14321432. doi:10.1007/BF02100041. PMID3041010. [33] Zhu Q et al. (2005). "Deubiquitination by proteasome is coordinated with substrate translocation for proteolysis in vivo". Exp Cell Res 307 (2): 43651. doi:10.1016/j.yexcr.2005.03.031. PMID15950624. [34] Wenzel T, Baumeister W (1995). "Conformational constraints in protein degradation by the 20S proteasome". Nat Struct Biol 2 (3): 199204. doi:10.1038/nsb0395-199. PMID7773788. [35] Smith DM et al. (2005). "ATP binding to PAN or the 26S ATPases causes association with the 20S proteasome, gate opening, and translocation of unfolded proteins". Mol Cell 20 (5): 68798. doi:10.1016/j.molcel.2005.10.019. PMID16337593. [36] Liu CW, Li X, Thompson D, et al. (October 2006). "ATP binding and ATP hydrolysis play distinct roles in the function of 26S proteasome". Mol. Cell 24 (1): 3950. doi:10.1016/j.molcel.2006.08.025. PMID17018291. [37] Smith DM et al. (2006). "Proteasomes and their associated ATPases: a destructive combination". J Struct Biol 156 (1): 7283. doi:10.1016/j.jsb.2006.04.012. PMID16919475. [38] Hoyt MA et al. (2006). "Glycine-alanine repeats impair proper substrate unfolding by the proteasome". EMBO J 25 (8): 17209. doi:10.1038/sj.emboj.7601058. PMC1440830. PMID16601692. [39] Zhang M, Coffino P (2004). "Repeat sequence of Epstein-Barr virus-encoded nuclear antigen 1 protein interrupts proteasome substrate processing". J Biol Chem 279 (10): 863541. doi:10.1074/jbc.M310449200. PMID14688254. [40] Voges D et al. (1999). "The 26S proteasome: a molecular machine designed for controlled proteolysis". Annu Rev Biochem 68: 10151068. doi:10.1146/annurev.biochem.68.1.1015. PMID10872471. [41] Rape M, Jentsch S (2002). "Taking a bite: proteasomal protein processing". Nat Cell Biol 4 (5): E1136. doi:10.1038/ncb0502-e113. PMID11988749. [42] Rape M, Jentsch S (2004). "Productive RUPture: activation of transcription factors by proteasomal processing". Biochim Biophys Acta 1695 (13): 20913. doi:10.1016/j.bbamcr.2004.09.022. PMID15571816. [43] Asher G, Reuven N, Shaul Y (2006). "20S proteasomes and protein degradation "by default"". Bioessays 28 (8): 8449. doi:10.1002/bies.20447. PMID16927316. [44] Asher G, Shaul Y (2005). "p53 proteasomal degradation: poly-ubiquitination is not the whole story". Cell Cycle 4 (8): 10158. doi:10.4161/cc.4.8.1900. PMID16082197. [45] Shringarpure R et al. (2003). "Ubiquitin conjugation is not required for the degradation of oxidized proteins by proteasome". J Biol Chem 278 (1): 3118. doi:10.1074/jbc.M206279200. PMID12401807. [46] Gille C et al. (2003). "A Comprehensive View on Proteasomal Sequences: Implications for the Evolution of the Proteasome". J Mol Biol 326 (5): 14371448. doi:10.1016/S0022-2836(02)01470-5. PMID12595256. [47] Bochtler M et al. (1999). "The proteasome". Annu. Rev. Biophys. Biomol. Struct. 28: 295317. doi:10.1146/annurev.biophys.28.1.295. PMID10410804. [48] Chesnel F et al. (2006). "Cyclin B dissociation from CDK1 precedes its degradation upon MPF inactivation in mitotic extracts of Xenopus laevis embryos". Cell Cycle 5 (15): 168798. doi:10.4161/cc.5.15.3123. PMID16921258. [49] Brito DA, Rieder CL (2006). "Mitotic checkpoint slippage in humans occurs via cyclin B destruction in the presence of an active checkpoint". Curr Biol 16 (12): 1194200. doi:10.1016/j.cub.2006.04.043. PMC2749311. PMID16782009. [50] Havens CG et al. (2006). "Regulation of late G1/S phase transition and APC Cdh1 by reactive oxygen species". Mol Cell Biol 26 (12): 470111. doi:10.1128/MCB.00303-06. PMC1489138. PMID16738333. [51] Bashir T et al. (2004). "Control of the SCF(Skp2-Cks1) ubiquitin ligase by the APC/C(Cdh1) ubiquitin ligase". Nature 428 (6979): 1903. doi:10.1038/nature02330. PMID15014502. [52] Higashitsuji H et al. (2005). "The oncoprotein gankyrin negatively regulates both p53 and RB by enhancing proteasomal degradation". Cell Cycle 4 (10): 13357. doi:10.4161/cc.4.10.2107. PMID16177571. [53] Dharmasiri S, Estelle M (2002). "The role of regulated protein degradation in auxin response". Plant Mol Biol 49 (34): 4019. doi:10.1023/A:1015203013208. PMID12036263. [54] Weijers D et al. (2005). "Developmental specificity of auxin response by pairs of ARF and Aux/IAA transcriptional regulators". EMBO J 24 (10): 187485. doi:10.1038/sj.emboj.7600659. PMC1142592. PMID15889151. [55] Schwartz LM et al. (1990). "Activation of polyubiquitin gene expression during developmentally programmed cell death". Neuron 5 (4): 4119. doi:10.1016/0896-6273(90)90080-Y. PMID2169771. [56] Lw P et al. (1997). "Expression of a 26S proteasome ATPase subunit, MS73, in muscles that undergo developmentally programmed cell death, and its control by ecdysteroid hormones in the insect Manduca sexta". FEBS Lett 400 (3): 3459. doi:10.1016/S0014-5793(96)01413-5. PMID9009228. [57] Pitzer F et al. (1996). "Removal of proteasomes from the nucleus and their accumulation in apoptotic blebs during programmed cell death". FEBS Lett 394 (1): 4750. doi:10.1016/0014-5793(96)00920-9. PMID8925925. [58] Adams J et al. (1999). "Proteasome inhibitors: a novel class of potent and effective antitumor agents". Cancer Res 59 (11): 261522. PMID10363983. [59] Orlowski RZ (1999). "The role of the ubiquitin-proteasome pathway in apoptosis". Cell Death Differ 6 (4): 303313. doi:10.1038/sj.cdd.4400505. PMID10381632. [60] Garrido C, Brunet M, Didelot C, Zermati Y, Schmitt E, Kroemer G (2006). "Heat Shock Proteins 27 and 70: Anti-Apoptotic Proteins with Tumorigenic Properties". Cell Cycle 5 (22): 22. PMID17106261.

406

Proteasome
[61] Park SH et al. (2007). "The cytoplasmic Hsp70 chaperone machinery subjects misfolded and endoplasmic reticulum import-incompetent proteins to degradation via the ubiquitin-proteasome system". Mol Biol Cell 18 (1): 15365. doi:10.1091/mbc.E06-04-0338. PMC1751312. PMID17065559. [62] Dai Q et al. (2005). "Regulation of the cytoplasmic quality control protein degradation pathway by BAG2". J Biol Chem 280 (46): 3867381. doi:10.1074/jbc.M507986200. PMID16169850. [63] Bader N, Grune T (2006). "Protein oxidation and proteolysis". Biol Chem 387 (1011): 13515. doi:10.1515/BC.2006.169. PMID17081106. [64] Davies KJ (2003). "Degradation of oxidized proteins by the 20S proteasome". Biochimie 83 (34): 30110. doi:10.1016/S0300-9084(01)01250-0. PMID11295490. [65] Lehman NL (2009). "The ubiquitin proteasome system in neuropathology". Acta Neuropathol 118 (3): 32947. doi:10.1007/s00401-009-0560-x. PMC2716447. PMID19597829. [66] McNaught KS et al. (2006). "Proteasomal dysfunction in sporadic Parkinson's disease". Neurology 66 (10 Suppl 4): S3749. doi:10.1212/01.wnl.0000221745.58886.2e. PMID16717251. [67] Sharma N et al. (2006). "Alpha-Synuclein budding yeast model: toxicity enhanced by impaired proteasome and oxidative stress". J Mol Neurosci 28 (2): 16178. doi:10.1385/JMN:28:2:161. PMID16679556. [68] Murata S, Sasaki K, Kishimoto T, et al. (June 2007). "Regulation of CD8+ T cell development by thymus-specific proteasomes" (http:/ / www. sciencemag. org/ cgi/ pmidlookup?view=long& pmid=17540904). Science 316 (5829): 134953. doi:10.1126/science.1141915. PMID17540904. . Retrieved 2009-03-26. [69] Cascio P, Hilton C, Kisselev AF, Rock KL, Goldberg AL (May 2001). "26S proteasomes and immunoproteasomes produce mainly N-extended versions of an antigenic peptide". EMBO J. 20 (10): 235766. doi:10.1093/emboj/20.10.2357. PMC125470. PMID11350924. [70] Donna L Mallery, William A McEwan, Susanna R Bidgood, Greg J Towers, Chris M Johnson, and Leo C James, (November 2010). "Antibodies mediate intracellular immunity through tripartite motif-containing 21 (TRIM21)" (http:/ / www. pnas. org/ content/ early/ 2010/ 11/ 01/ 1014074107). PNAS 107 (46): 1998519990. doi:10.1073/pnas.1014074107. PMC2993423. PMID21045130. . Retrieved 2010-03-11. [71] United States Food and Drug Administration press release (http:/ / www. fda. gov/ bbs/ topics/ NEWS/ 2003/ NEW00905. html) 13 May 2003. Access date 29 December 2006. See also FDA Velcade information page (http:/ / www. fda. gov/ cder/ drug/ infopage/ velcade/ ). [72] Fisher RI et al. (2006). "Multicenter phase II study of bortezomib in patients with relapsed or refractory mantle cell lymphoma". J Clin Oncol 24 (30): 486774. doi:10.1200/JCO.2006.07.9665. PMID17001068. [73] Jakob C et al. (2007). "Circulating proteasome levels are an independent prognostic factor for survival in multiple myeloma". Blood 109 (5): 21005. doi:10.1182/blood-2006-04-016360. PMID17095627. [74] Shah SA et al. (2001). "26S proteasome inhibition induces apoptosis and limits growth of human pancreatic cancer". J Cell Biochem 82 (1): 11022. doi:10.1002/jcb.1150. PMID11400168. [75] Nawrocki ST et al. (2004). "The proteasome inhibitor bortezomib enhances the activity of docetaxel in orthotopic human pancreatic tumor xenografts". Mol Cancer Ther 3 (1): 5970. PMID14749476. [76] Schenkein D (2002). "Proteasome inhibitors in the treatment of B-cell malignancies". Clin Lymphoma 3 (1): 4955. doi:10.3816/CLM.2002.n.011. PMID12141956. [77] O'Connor OA et al. (2005). "Phase II clinical experience with the novel proteasome inhibitor bortezomib in patients with indolent non-Hodgkin's lymphoma and mantle cell lymphoma". J Clin Oncol 23 (4): 67684. doi:10.1200/JCO.2005.02.050. PMID15613699. [78] Schmidtke G et al. (1999). "How an inhibitor of the HIV-I protease modulates proteasome activity". J Biol Chem 274 (50): 3573440. doi:10.1074/jbc.274.50.35734. PMID10585454. [79] Laurent N et al. (2004). "Effects of the proteasome inhibitor ritonavir on glioma growth in vitro and in vivo". Mol Cancer Ther 3 (2): 12936. PMID14985453. [80] Zollner TM et al. (2002). "Proteasome inhibition reduces superantigenmediated T cell activation and the severity of psoriasis in a SCID-hu model". J Clin Invest. 109 (5): 671679. doi:10.1172/JCI12736. PMC150886. PMID11877475. [81] Elliott PJ et al. (1999). "Proteasome inhibition: A novel mechanism to combat asthma". J Allergy Clin Immunol. 104 (2 Pt 1): 294300. doi:10.1016/S0091-6749(99)70369-6. PMID10452747. [82] Verdoes M et al. (2006). "A fluorescent broad-spectrum proteasome inhibitor for labeling proteasomes in vitro and in vivo". Chem Biol 13 (11): 121726. doi:10.1016/j.chembiol.2006.09.013. PMID17114003.

407

Proteasome

408

External links
Glickman, Michael H.; Adir, Noam (2004). "The Proteasome and the Delicate Balance between Destruction and Rescue". PLoS Biology 2 (1): e13. doi:10.1371/journal.pbio.0020013. PMC314468. PMID14737189. The Yeast 26S Proteasome with list of subunits and pictures (http://biochemie.web.med.uni-muenchen.de/ feldmann/proteasome_units.html) Ciechanover, A (2005). "Early work on the ubiquitin proteasome system, an interview with Aaron Ciechanover". Cell Death and Differentiation 12 (9): 116777. doi:10.1038/sj.cdd.4401691. PMID16094393. Hershko, A (2005). "Early work on the ubiquitin proteasome system, an interview with Avram Hershko". Cell Death and Differentiation 12 (9): 115861. doi:10.1038/sj.cdd.4401709. PMID16094391. Adams, J (2005). "Early work on the ubiquitin proteasome system, an interview with Irwin Rose". Cell Death and Differentiation 12 (9): 11626. doi:10.1038/sj.cdd.4401700. PMID16094392. Cvek, B; Dvorak, Z (2007). "Targeting of nuclear factor-kappaB and proteasome by dithiocarbamate complexes with metals." (http://www.benthamdirect.org/pages/content.php?CPD/2007/00000013/00000030/0010B. SGM). Current pharmaceutical design 13 (30): 315567. doi:10.2174/138161207782110390. PMID17979756.

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Aldado, Ffirehorse, Figma, Figure, Firetrap9254, Fishingpal99, Flavaflav1005, Florentino floro, Fnielsen, Forluvoft, Freakofnurture, FreplySpang, Friendly Neighbour, Frostyservant, Fruge, Fs, Fvasconcellos, G3pro, GAThrawn22, GHe, GODhack, GVnayR, Gaara san, Gakrivas, Galoubet, Gary King, Gatortpk, Gazibara, GeeJo, Gene Nygaard, GeoMor, Giftiger wunsch, Giftlite, Gilisa, Gilliam, Gimmetrow, Gjuggler, Glen Hunt's DNA, Glenn, Gmaxwell, GoEThe, Goatasaur, Gogo Dodo, GoldRingChip, Golnazfotohabadi, GordonWatts, GorillaWarfare, Gracenotes, Graeme Bartlett, GraemeL, Grafikm fr, Graft, Graham87, GrahamColm, Grandegrandegrande, GregorB, Grover cleveland, Gurko, Gustav von Humpelschmumpel, Gutza, Gwsrinme, HJ Mitchell, Hadal, Hagerman, Hairchrm, Hairwheel, Hammersoft, Hannes Rst, Harianto, HayleyJohnson21, Headbomb, Heathhunnicutt, Hephaestos, Heron, Hersfold, Hersfold tool account, Heyheyhack, Hockey21dude, Horatio, Hu, Hughdbrown, Hurricanehink, Hut 8.5, Hvn0413, Hzh, I hate DNA, Ialsoagree, Iapetus, Icairns, Ilia Kr., Impamiizgraa, InShaneee, Inge-Lyubov, IronGargoyle, Isilanes, Isis07, Itub, Ixfd64, Izehar, JForget, JHMM13, JWSchmidt, JWSurf, JZuehlke, Jacek Kendysz, Jackrm, JamesMLane, JamesMt1984, Janejellyroll, Jarhed, Javert, Jaxl, Jeka911, Jer ome, JeremyA, Jerzy, Jetsetpainter, Jh51681, Jiddisch, Jim1138, Jimriz, Jimwong, Jlh29, Jls043, Jmcc150, Jnorton7558, JoanneB, Joconnol, Joeywallace9, Johanvs, John Mackenzie Burke, JohnArmagh, Johntex, Johnuniq, Jojit fb, JonMoulton, Jonrunles, Jonverve, Jorge Stolfi, Jorvik, JoshuaZ, Josq, Jossi, Jrtayloriv, Jstech, Jujutacular, Julian Diamond, Jumbo Snails, Junes, Juneythomas, Jwrosenzweig, Kahlfin, Kapow, Karrmann, Katyare, Kaushlendratripathi, Kazkaskazkasako, Kbh3rd, Kbrose, Keegan, Keepweek, Keilana, Kelly Martin, Kemyou, Kendrick7, KerryO77, Kevin B12, Kevmitch, Kghose, Kholdstare99, Kierano, KimvdLinde, King of Hearts, KingTT, Kingturtle, Kitch, Knowledge Seeker, KnowledgeOfSelf, Koavf, KrakatoaKatie, Kums, Kungfuadam, Kuru, Kwamikagami, Kwekubo, L Kensington, LA2, LFaraone, La goutte de pluie, Lascorz, Latka, Lavateraguy, Lee Daniel Crocker, Leevanjackson, Lemchesvej, Leptictidium, Lerdsuwa, Leuko, Lexor, Lfh, Lhenslee, Lia Todua, LightFlare, Lightmouse, Lightspeedchick, Ligulem, LikeLakers2, Lincher, Lion Wilson, Lir, Llongland, Llull, Lockesdonkey, Logical2u, Loginbuddy, Looxix, Loren36, Loris, Low-frequency internal, Luigi30, Luk, Lumos3, Luna Santin, Luuva, MER-C, MKoltnow, MONGO, MSGJ, Mac, Madeleine Price Ball, Madhero88, Magadan, Magister Mathematicae, Magnus Manske, Majorly, Malcolm rowe, Malo, Mandarax, Mandyj61596, Mantissa128, Marcus.aerlous, Marj Tiefert, Martin.Budden, MarvPaule, Mashin6, Master dingley, Materialscientist, Mattbr, Mattbrundage, Mattjblythe, Maurice Carbonaro, Mav, Max Baucus' DNA, Max Naylor, McDogm, Medessec, Medos2, Melaen, Melchoir, Mentalmaniac07, Mgiganteus1, Mgtoohey, Mhking, Michael Devore, MichaelHa, Michaelas10, Michigan user, MidgleyDJ, Midnightblueowl, Midoriko, Mika293, Mike Rosoft, Mikker, Mikko Paananen, Minimac, Mintman16, MisfitToys, Misza13, Mithent, Mjpieters, Mleefs7, Moink, Moorice, Mortene, Motley Crue Rocks, Mr Bungle, Mr Meow Meow, Mr Stephen, MrErku, Mstislavl, Mstroeck, Mulad, Munita

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Prasad, Muro de Aguas, Mwanner, Mxn, Nakon, Nanettea, Nanodance, Narayanese, Natalie Erin, Natarajanganesan, Nate1028, NatureA16, Nauseam, Nbauman, Neckro, Netkinetic, Netoholic, Neutrality, Never give in, NewEnglandYankee, Nick Number, Nighthawk380, NighthawkJ, Nihiltres, Nirajrm, Nishkid64, Nitecrawler, Nitramrekcap, No Guru, NoIdeaNick, NochnoiDozor, Nohat, Nono64, Northfox, NorwegianBlue, Nthornberry, NuclearWarfare, Nunh-huh, OBloodyHell, OOODDD, Obli, Oblivious, Ocaasi, Ocolon, OekelWm, Ojl, Omicronpersei8, Onco p53, Opabinia regalis, Opelio, Orphan Wiki, Orrin Hatch's DNA, Orthologist, Ortolan88, Ouishoebean, Outriggr, OwenX, P99am, PDH, PFHLai, PaePae, Pakaran, Pascal666, Patrick0Moran, Patrick2480, Patstuart, Paul Foxworthy, Paul venter, Paulinho28, Pcb21, Pde, Peak, Pedro, Perseus, Son of Zeus, Persian Poet Gal, Peter Isotalo, Peter K., Peter Winnberg, Pgan002, Philip Trueman, PhilipO, Phoenix Hacker, Pierceno, PierreAbbat, Pifactorial, Pigman, Pigmietheclub, Pilotguy, Pkank, Pkirlin, Poor Yorick, Portugue6927, Potatoswatter, Prathfig, Preston47, Prestonmag, Priscilla 95925, Pristontale111, Pro crast in a tor, Prodego, Psora, PsyMar, Psymier, Pumpkingrower05, Pyrospirit, Quebec99, Quickbeam, Qutezuce, Qxz, R'n'B, R. 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YanWong, Yansa, Yaser al-Nabriss, Yasha, Yikrazuul, Yobol, Yomama9753, Younusporteous, Yurik, ZScout370, Zahid Abdassabur, Zahiri, Zanaq, Zazou, Zell Miller's DNA, Zephyris, Zoicon5, Zouavman Le Zouave, Zsinj, Zven, 1391 anonymous edits Protein Source: http://en.wikipedia.org/w/index.php?oldid=462352893 Contributors: 0, 162.129.26.xxx, 168..., 200itlove, 8472, A K AnkushKumar, A-giau, AA, ABF, Aarktica, Aaron Schulz, AaronY, Abce2, Achilles.g, Acroterion, ActivExpression, AdamRetchless, Adashiel, Adenosine, AdjustShift, Agent Smith (The Matrix), Ageo020, Agesworth, Agilemolecule, Ahoerstemeier, Aitias, Alansohn, Ale jrb, Alexjohnc3, Alison, Alpha Quadrant, Alphachimp, Altenmann, Altruistic Egotist, Alvestrand, Amb sib, Amotz, Ams80, AnOddName, Anclation, Andonic, Andre Engels, AndrewWTaylor, Andrewpmk, Andries, Angr, Annabel, Antandrus, Anthony, Aragorn2, Arakunem, Arcadian, Arkuat, Ascidian, Ashdog137, Asher196, Aspenandgwen, AssistantX, Astrojan, AuburnPilot, AutoFire, AvicAWB, 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S. 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Article Sources and Contributors

Kaushlendratripathi, Keenan Pepper, Keilana, Kengo1040, Kevinallencrittenden, Khajidha, Khooly59, Kijitah, Kingpin13, Kjkolb, Knivusa, Knutux, Kostisl, Kpjas, Kubra, Kuru, L-Tyrosine, La goutte de pluie, Leeyc0, Lesterama, Leszek Jaczuk, Lexor, Lfh, Lights, LikeHolyWater, LilHelpa, Ling.Nut, Llull, Looxix, LosDorks, Lovey73, LuckyWizard, Lumos3, M1ss1ontomars2k4, MD87, Maartend8, MacGyverMagic, Magister Mathematicae, Magnus Manske, Majorclanger, Malljaja, Mani1, Marj Tiefert, Master gopher, Materialscientist, Matt Fitzpatrick, Matteo2000, Matthew Yeager, Mattisse, Maurice Carbonaro, Mav, Maxamegalon2000, Maximaximax, Mayalld, McGeddon, Memset, Methcub, Mikael Hggstrm, Mike2vil, Miyagawa, Molybdenumblue, Monurkar, Moryoav, MrOllie, Mushin, Myanw, N5iln, NHRHS2010, Naive cynic, Nandesuka, Narayanese, Natalie Erin, Naturespace, NerdyScienceDude, Nevin Janzen, Nihiltres, Nikki311, Nilfanion, Nlu, Nomad2u001, Nono64, Nucleophilic, Nuklear, Oasisbob, Obradovic Goran, Olivier, Omegatron, Onco p53, Opabinia regalis, Opelio, Openlander, Orangutan, Otisjimmy1, Otivaeey, Oystertoadfish, Pansi gupta, Patstuart, Pauli133, Peak, PeterSymonds, Pgan002, Philip Trueman, Physchim62, PierreAbbat, Pinethicket, Possum, Prilla, Pro crast in a tor, Prodego, Proofreader77, Qef, Qxz, RE73, Rada, Radon210, Ravi12346, Res2216firestar, Rewster, RexNL, Rich Farmbrough, Richard Arthur Norton (1958- ), Richard001, Rifleman 82, Rittard, Rjwilmsi, Rlee1185, Rmosler2100, Roadrunner, Roadsoap, Robma, Rory096, RoyBoy, Sakkura, Sam Hocevar, SchnitzelMannGreek, Seans Potato Business, Selket, Serinde, Shaddack, ShadowBlade1000, Shadowfax0, Sheehan, Shekharsuman, Sikkema, SimonP, Sionus, SirGrant, Skoddet, Sligocki, Slow Riot, Slp1, Smithbrenon, Smokefoot, Snowmanradio, Soks1, Soren.harward, SpLoT, SpaceFlight89, Squidonius, SriMesh, Stephentaylor, Stewartadcock, Stillnotelf, Stismail, Stone, Stormhawk, Superarthur, Switchsonic, Sydney naturopath, TBadger, Tarawneh, Tarotcards, Teddks, Teles, Thaseint, Thehelpfulone, Tide rolls, Tiggerjay, TimVickers, Tjod, Tobias Hoevekamp, Tonderai, Tony1, TonyTheTiger, Toon05, Toyz, Transhumanist, Tremello, Trevor Wennblom, Tristanb, Trlblzr, Truthflux, Tryptofish, Ttony21, Tuckerekcut, Turnstep, Tyler, Ultimate sickness, Umleashuh, Unukorno, User A1, Ustye, Utcursch, Uttam Pal, Versageek, Vina, ViolentRage, WVhybrid, Weedwhacker128, White.matthew.09, Whittlewiki, Wickey-nl, Wiki007wiki, William Avery, WillowW, Wimt, Winchelsea, Wizzy, Wolfmankurd, WriterHound, Wrstroud, Wtmitchell, Ww, Wwallacee, Xinit, XoiiLov3Youuxo55, Xook1kai Choa6aur, Yamamoto Ichiro, Yoasif, Yobmod, Yosef1987, Yyy, Zaphraud, Zhou Yu, Zoicon5, ~K, 4101 , anonymous edits Properties of the twenty amino acids Source: http://en.wikipedia.org/w/index.php?oldid=455045516 Contributors: Aa77zz, Arcadian, BenJWoodcroft, Benlisquare, BrentN, Cacycle, Capricorn42, ChipperGuy, Chris Roy, Cpt ricard, DMacks, Dancojocari, Drphilharmonic, Edgar181, Eloil, Erechtheus, Flakinho, Gregogil, Gurch, Hichris, Itub, Jackroven, Keenan Pepper, Kkmurray, Kumorifox, Kupirijo, L-Tyrosine, Lanthanum-138, Leptictidium, MRandazzUCSD, Microball, Mikael Hggstrm, Mindmatrix, Mlaffs, Myceteae, Nagelfar, Pgan002, Phlounder, Phoenix Hacker, Qchristensen, Radagast83, Rich Farmbrough, Scoutfreak, Selket, Stifynsemons, Takometer, TimVickers, Uthbrian, Wiki007wiki, Willking1979, Xact, 49 anonymous edits Myoglobin Source: http://en.wikipedia.org/w/index.php?oldid=457628794 Contributors: Adrian J. Hunter, Aitias, AndrewGNF, Anonymi, Antandrus, AnteaterZot, Arcadian, Atakdoug, Atlant, BerserkerBen, Blindman shady, Bob98133, Boghog, Bogwhistle, BokicaK, Br77rino, Bryan Derksen, C'est moi, CRaines, Cburnett, Clicketyclack, ClockworkSoul, DerHexer, Digfarenough, Dwmyers, El C, Entropic existence, Epbr123, Erexeisen, Etxrge, Excirial, Fvasconcellos, Gilliam, Glacian79, Habj, Herbertxu, Inslid, JWSchmidt, Jack B108, Jfdwolff, John, Jonowen008, Kingpin13, MaerlynsRainbow, Magnus Manske, Marc Venot, Marj Tiefert, Mattis, Mav, Memenen, Metalloid, Mmoneypenny, Mr Adequate, Noca2plus, Nono64, Pdcook, Phil Boswell, Polly, RDBrown, ResearchProteins1, Revth, Rich Farmbrough, Rifleman 82, Rjwilmsi, SV Resolution, Samulili, Shell Kinney, Shu99, Shureg, SkyLined, Some standardized rigour, Spiritia, Splette, Stenzela, Tangpoet, Tavilis, That Guy, From That Show!, The Rambling Man, The Rogue Penguin, TimVickers, Twooars, Vipinhari, Vossman, Wayne Slam, Wikipe-tan, Wisdom89, Zybez, 127 anonymous edits Hemoglobin Source: http://en.wikipedia.org/w/index.php?oldid=464437569 Contributors: AWeenie, Abscissa, Achaemenes, AdrianaW, Ahoerstemeier, Aka ninja69, Alain.michalowicz, Alansohn, Aldis90, Aleph Infinity, Alex.tan, Alison, Alsandro, Alvin Seville, Amb sib, Anatoly IVANOV, Andres, Andrewko, Aneurysmal, Anticipation of a New Lover's Arrival, The, Appraiser, Aragan Jarosalam, Arcadian, Arisa, Atlant, AxelBoldt, Baron von Zandt, Beachbutt66, Becksdrake, Beepu, Belovedfreak, Benamies, Bensaccount, BerserkerBen, Bhca, BiT, Bkell, Black Cat Claws, Blu3d, Bob Castle, Bobo192, Bogey97, Boghog, Bombhead, Brian Huffman, Bucephalus, CIreland, CS46, Cajolingwilhelm, Calm, CalumH93, CanadianLinuxUser, Canderson7, Carlosisgay, Carstensen, Ccevo2010, Cek, Chairman S., Chaldor, Charles Brooking, Christian List, Christopherlin, ChunkySoup, ClockworkSoul, Cmcnicoll, Codwiki, Commit charge, CommonsDelinker, Conversion script, Corinne68, Correctaboot, Cottingham, Crabula, Csv028, Cwtyler, D, DGaw, DRosenbach, DVD R W, Dan Gluck, Dannyt101, Darklilac, Darwinek, Dawn Bard, Dejvid, Dereck poliink, Deviceauthor1, Diberri, Difu Wu, Discospinster, Doc glasgow, Doc.mari, Donarreiskoffer, Doyley, Dr. F.C. Turner, Draeco, Drphilharmonic, Dungodung, Dwmyers, EMSPhydeaux, Edgar181, Edward Z. Yang, Eleassar, Epbr123, Epingchris, Erik9, Escape Orbit, Ettrig, Eug, Excirial, Fabrcio Kury, Fallendarling, FisherQueen, Flyguy649, Forluvoft, Freakmighty, Fresheneesz, Friginator, Funkadan, Funnyfarmofdoom, Fvasconcellos, GPHemsley, Gak, Galoubet, Gator0929, Geoking66, Gilgamesh, Gilliam, Gogo Dodo, Greatpatton, Green Giant, Gurch, Gzkn, H Padleckas, Habj, Hairy Dude, Hamiltondaniel, Hans Dunkelberg, Headleyp, Heathmoor, Heli1062, Helios87, Hellbus, Hellisp, Heron, HexaChord, Hitokirishinji, Hroychow, Hurricanefan24, IGeMiNix, Iepeulas, Immunize, Imnotminkus, Isaac.holeman, Ivan Bajlo, Ixfd64, J.delanoy, JRamlow, Jack B108, Jaknouse, Jamessmith2011, Jared Wilmoth, Jebba, Jfdwolff, Jkl, Joeshmoe123123123, John of Reading, JohnArmagh, JohnCD, Jojocurler, Jonathan.s.kt, Jordanmif, Joyous!, Julesd, Julianva, Jusdafax, Jylothr, KGasso, Kalaiarasy, Kandar, Kbh3rd, Kinberg1, King of the Eagles, Kirika, Kisher07, Kku, Koavf, Kpjas, Kubigula, Kwamikagami, Kyoko, L Kensington, Lateg, LeadSongDog, LeeG, Lemchesvej, Lennert B, Lenov, Leonid 2, Lightmouse, LittleHow, LizardJr8, Loonymonkey, LostLucidity, Low-frequency internal, Luk, Lukas.S, Lunajurai, MER-C, Mac Davis, Macy, Madhero88, Magnus Manske, Mah131, Man12345, Mark Yen, Martin451, MartinezMD, Master of Pies, Master of Puppets, Master of darkness 666, Materialscientist, Matthias.hilty, Mav, Maximaximax, Mejoribus, Melman14, Mentifisto, Metalloid, Michaeldsuarez, Midorilily, Miguel Andrade, Mikael Hggstrm, Mike Rosoft, Mike2vil, Miranda, Mitch Ames, Mmoneypenny, Mogsmar5, Momo san, MottyGlix, Moulder, Muzzypatmn, Mynameisnoted, Myxoma, Natisto, Natkeeran, Naturespace, NawlinWiki, Neptunecradle, New299, NewEnglandYankee, Nibuod, Nick Number, Nick04, Nieren, Nivix, Nixeagle, Nono64, Northfox, Notcart, Nuttycoconut, Opabinia regalis, Orangemarlin, Orphan Wiki, Ouishoebean, Oxymoron83, PDH, PaddyM, Pathew, Pdeitiker, Pedrora, Perditor, Perlygatekeeper, Persian Poet Gal, Petergans, Peterl, Petiatil, Pgan002, Pgk, Philip Trueman, Phoenix2, Pinethicket, Pinot, Pinzo, Pmm530, Prisonblues, Priyankadaga, Pseudomonas, Qaddosh, Quazgaa, RJFJR, Rabend, Radagast83, Ramrajesh, Ramsus, ReallyNiceGuy, Reaper Eternal, RebootThalheimer, Recipe For Hate, Reyps, Rhiani-ani-on, Riana, Rich Farmbrough, Richthewitch, Rifleman 82, Rjwilmsi, Rmessenger, Robert K S, Rrenner, Rubicon, Sam Hocevar, Saravask, Savant13, Sbharris, Sbmehta, Sceptre, Schzmo, Scientificone, Scottalter, Sephirothrr, Sercretions101, Shadowjams, Shanes, Sharksoup, Shawthorn, Sherybatta, Shirik, Sietse Snel, Silivrenion, Silversink, Silversmith, Sir Vicious, Sitush, Sk741, Smodtactical, Snek01, Snowmanradio, SoLando, Some standardized rigour, SpuriousQ, Srnec, Staticd, Stemonitis, Stevertigo, SuperHamster, SwirlBoy39, TaintedMustard, Tarosan, Taucetiman, Taw, Taxman, Teh tennisman, Temporaluser, The Thing That Should Not Be, TheAlmightyEgg, TheTito, Three-quarter-ten, Tide rolls, Tillalb, TimVickers, Tjmayerinsf, Tommy2010, TonySt, Topbanana, Treisijs, Triggtay, Tristanb, Twooars, Uazikoff, Unixer, Vainamainien, Vary, VasilievVV, Vgree 01, Vianello, Vivin, Vrenator, Wavelength, West Brom 4ever, WhatamIdoing, Wickey-nl, WikHead, WikipedianMarlith, Will Beback Auto, Winston365, Wouterblox, Wwallacee, X!, Xiggelee, Yikrazuul, Ym3, Yosaphbridge, Youssefsan, Ysangkok, Zaheer12a, ZarathustraD, ZenSaohu, Zephyris, Zfr, Ziggurat, Ziphon, Zzuuzz, 789 anonymous edits Enzyme catalysis Source: http://en.wikipedia.org/w/index.php?oldid=460817468 Contributors: Arbitrarily0, Arcadian, Banus, Bci2, Beta cafe, BigrTex, Bomac, Clemwang, Colonies Chris, Dratman, Dspoel, Emw, Hughdbrown, Icelight, Jamiejoseph, Jebus989, Jkhwang, Kosigrim, Mion, NorwegianBlue, Pdcook, Perkinsonj, Peter Karlsen, Rich Farmbrough, Rifleman 82, Rjwilmsi, Simon12, Stefano Garibaldi, TimVickers, Vanisheduser12345, Wikidudeman, Woohookitty, Zephyris, 51 anonymous edits Enzyme kinetics Source: http://en.wikipedia.org/w/index.php?oldid=462305976 Contributors: 08cflin, A2a2a2, Adenosine, Adrian J. Hunter, Alan Liefting, Alansohn, Alexandria, AndrewWTaylor, Angmar09, Anylai, Arcadian, BanjoCam, Bbatsell, BecR, Blanchardb, Bobo192, BokicaK, Brighterorange, Brim, Britzingen, Cacycle, CarinaT, Carstensen, Chaser, Chymo72, Clemwang, ClockworkSoul, Cmprince, DRHagen, Dana boomer, Dasdasdase, Davo22, Diberri, Dmol, Domitori, Dr Zak, ESkog, Economo, Eskeptic, Espresso Addict, Evenap, Flyguy649, Forluvoft, Fourthhour, Gene Nygaard, Giftlite, Graft, Gkhan, Hannes Rst, Hede2000, I need a game, Igodard, Itub, Jacobandrew2012, Java13690, Jeff Dahl, JoNo67, Joshwa-hayswa, Karam.Anthony.K, Kimchi.sg, Kmg90, KnightRider, Knights who say ni, Koavf, Kozuch, Kyoko, Laurinavicius, Legalvoice123, Lightmouse, Luna Santin, Manysplintered, Mastercampbell, Mauegd, Michael A. Wilkins, Michael Hardy, Mion, Moonsword, MrOllie, Nigholith, NuclearWarfare, Oda Mari, OpenToppedBus, Outriggr, Poccil, Pro bug catcher, Qef, R'n'B, RDBrown, Rasikraj, Raul654, Rich Farmbrough, Richard001, Rjwilmsi, RobertG, RockMFR, Rschen7754, Ruute, Samsara, SandyGeorgia, Saravask, SebastianHelm, Shauun, Shyamal, Slashme, SnowFire, Sodin, Some standardized rigour, Sonjaaa, Spidrak, Spoladore, Stefano Garibaldi, Stillnotelf, Storm Rider, Suruena, Sxenko, Tbhotch, The Parsnip!, The Thing That Should Not Be, TheTweaker, TheoThompson, TimVickers, ToNToNi, Todd40324, Tom Doniphon, Tommy2010, Ukexpat, V8rik, Wabaman44, WillowW, Wknight94, Wnt, WolfmanSF, Xcomradex, Xiahou, Xijkix, Yaser al-Nabriss, 168 anonymous edits Lipid Source: http://en.wikipedia.org/w/index.php?oldid=464495146 Contributors: 168..., 27 Juni, 52 Pickup, 61 88 131 149a, A8UDI, Actam, Adambro, Addshore, AeonicOmega, Ahoerstemeier, Aitias, Albert galiza, Alice Davidson, Alteripse, Anaxial, Andrewx96, Andycjp, Andymc, Antandrus, Aranherunar, Arcadian, Arthena, Artichoker, Ashill, Ashleyisachild, AubreyEllenShomo, Autocracy, Bagelboi, Bensaccount, Bergamo, Bevo, Big Papa 13, Bobak, Bomac, Bongwarrior, Boosmith, Borgx, BorisTM, Boulaur, BuickCenturyDriver, Bulbeck, Burntsauce, CalebNoble, Can't sleep, clown will eat me, Capricorn42, Casper2k3, Cburnett, Cckkab, Cenarium, Ceyockey, Chaffers, Chairman S., CharlotteWebb, Choij, Chrislk02, Christian75, Cimex, Clicketyclack, ClockworkSoul, Compaddict11, Conversion script, Coppertwig, Costas78, Courcelles, Crazynas, Cruccone, Curiousgeorgethemonkey, DARTH SIDIOUS 2, DVD R W, Daf, Dakilla15, Dan4636, Daniel Cliff, Daniel5127, Dannyhoffman, DarkHorizon, Darklilac, Darth Panda, Dave6, Dcoetzee, DeadEyeArrow, Deor, DerHexer, Derek.cashman, Dina, Donarreiskoffer, Doodle77, Drphilharmonic, DubaiTerminator, Dybdahl, Eboireau, EchoBravo, EconoPhysicist, Edcolins, Edgar181, ElBeeroMan, Eleassar777, Elkman, Enigma55, Eog1916, EscapingLife, Esoltas, Etxrge, Everyking, Evil saltine, Ezhuttukari, FaerieInGrey, Felix Wan, Fenteany, Fieldday-sunday, Fireice, Frankenpuppy, Freiddie, GFP, Garzo, Giants27, Giftlite, Giraffedata, Glane23, Gomm, Gurch, Gustavb, Gwernol, HaeB, Haham hanuka, Hapsiainen, Hardrada, Harvy2004, Headbomb, Hiei-Touya-icedemon, Hodja Nasreddin, HolIgor, Hoops32, Hughesdavidw, Husond, Ikip, Immunize, Ixfd64, JForget, JHMM13, JWSchmidt, Jadeogle, Jauhienij, Javanbakht, Jdude89, Jedidan747, Jfdwolff, JimBrownish, JimVC3, JinJian, Joanjoc, JoeAnderson, Jrtayloriv, Jvcallaway, Kajasudhakarababu, Kandar, Karada, Karol Langner, Katgabrielle, Keilana, Kelly Katula, King003, Knopfkind, Kruusamgi, Krylonblue83, Kupirijo, Kuru, LeaveSleaves, Lemchesvej, Lennert B, Leptonsoup337, Lir, Lmaps, Loren.wilton, Lunchboxhero, Lunkwill42, MEHMEHMEHMEH, MER-C, MZMcBride, Mac, Madfalcon108, Madhero88, Magnus Manske, Mani1, Marc Venot, Marj Tiefert, Markacohen, Maryrebecca, Matanya (renamed), Materialscientist, Mav, Mayooranathan, MchRj2, MedicineMan555, Mgiganteus1, Michael Devore, Michaplot, Mifter, Mikael Hggstrm, Mike2vil, Miller17CU94, Mivadar, Miyagawa, Mr. Lefty, Mrsmosher, Mschel, Muffinman123123, Musselboy, Mxn, Myanw, Myrryam, NCurse, NHRHS2010, NYMets2000, Nakon, Nanominori, Narayanese, NellieBly, Neutrality, Niazac, Nikai, Nikolaevna, Nivix, Nlaeditor, Nlu, NochnoiDozor, Nothing98765432154, Nunh-huh, Ocaasi, Octahedron80, Oiws, Olegkolesnikov, Orbst, OrcaMorgan, Patrick, Pearle, Pedro, Peter Chastain, Pharaoh of the Wizards, Philip Trueman, Physicistjedi, Platonides, Plw, Pradilla94, Pschemp, Pyrospirit, Qst, Quadell, Quantumobserver, Quentonamos, Quidam65, Qxz, R500Mom, Ranveig, Reo On, RexNL, Reywas92, Rich Farmbrough, Richard001, Rifleman 82, Rjwilmsi, Roadnottaken, RobertG, Ronhjones, RoyBoy, Rubylover234, Ryanreednro, S charette, SWAdair, Sakkura, SallyForth123, Sam Korn, Sardanaphalus, Sasata, Sceptre, Sfmammamia, Shaddack,

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Article Sources and Contributors

Shenme, Shirulashem, Showjumpersam, Sion55, Sjakkalle, SpaceFlight89, Special-T, Sphinxlipos, Spook`, Squeezeweasel, Sten, Stephen Gilbert, Steven Weston, Stewartadcock, SubwayEater, Syrthiss, TLF1981, Tahmmo, Taroaldo, Tb, Tbhotch, Tempodivalse, TestPilot, The Rambling Man, The Thing That Should Not Be, ThePointblank, TheTito, TheUnionBlood, Thingg, Thunderboltz, Tide rolls, Tim1357, TimVickers, Tommy2010, Toytoy, Travis.Thurston, Tree Biting Conspiracy, Trusilver, Tullywinters, User A1, V8rik, VMS Mosaic, Vaz222, Velella, Vgranucci, ViperBite, Visium, Vreezkid, WLU, Walkerma, Wavelength, Wayne Slam, We hope, Where, Why My Fleece?, Wickey-nl, Wiki alf, WikipedianMarlith, Wikipelli, Wildnox, WiseManOnTheMountain, Woudloper, Yamamoto Ichiro, Yelsaimery, Zephyris, Zzuuzz, , , 833 anonymous edits Biological membrane Source: http://en.wikipedia.org/w/index.php?oldid=463308948 Contributors: 168..., Aceofhearts1968, Adashiel, Adrian J. Hunter, Aejohnst, Alan Liefting, Alansohn, Alpha Quadrant (alt), Animeronin, Apers0n, Arcadian, Bensaccount, Biophysik, BorisTM, Brghntr, CALR, Chrisahn, Conversion script, Dabombg, Drphilharmonic, EconoPhysicist, Elassint, Gtrmp, Hodja Nasreddin, Icek, JS Hoyer, JerrySteal, JuanitaJP, Jvbishop, Karol Langner, La goutte de pluie, LadyofHats, Lexor, Lunarbunny, Madman Ven Colt II, Magnus Manske, Marj Tiefert, NERIC-Security, Naddy, Nihiltres, PhilKnight, Pschemp, R. S. Shaw, RDBrown, Reo On, Rosentod, RyanGerbil10, Sadalmelik, SimonP, Sjb90, Skittleys, Smartse, Spike Wilbury, Squidonius, Thorncrag, Tide rolls, TimBentley, User A1, WAS 4.250, Wayne Slam, Zidonuke, , 44 anonymous edits Membrane protein Source: http://en.wikipedia.org/w/index.php?oldid=464451295 Contributors: 168..., A09147801, Alansohn, Arcadian, Asymptote, BIONICLE233, Baronnet, Bjrn P Pedersen, Boobermonkey, CalumH93, Connormah, Dcirovic, Drmies, El C, Fritzpoll, FrozenMan, Gosolowe, Graham87, Grendelkhan, Gurch, Hodja Nasreddin, IW.HG, Ixfd64, JWSchmidt, Jag123, Jfdwolff, Jusdafax, Kcgi, Ksalcines, Lalehkam, Lexor, Lfh, Magnus Manske, Michael Devore, Mike of Wikiworld, Namenotek, NellieBly, Pdcook, Pschemp, R'n'B, Radowell, Ramamoor, Road Wizard, Robomaeyhem, Rumadatme, ShaiM, Snapouse, Stepa, Stewartadcock, Tameeria, Thingg, Vojtech.dostal, Yamamoto Ichiro, 58 anonymous edits Cell membrane Source: http://en.wikipedia.org/w/index.php?oldid=464055323 Contributors: 168..., 1exec1, 6osama9, 7cg43m, ABF, AThing, Academic Challenger, Access Denied, Adambiswanger1, Adambro, Adenosine, Adrian.benko, Ajraddatz, Akajune, Akanemoto, Akap007, Alan012, Alansohn, Ale jrb, AlexiusHoratius, Aliyarockzharder, Anaxial, Anchit virmani, Andrea.gf, Andres, Andy85719, Antandrus, Apparition11, Arcadian, Arjun01, ArmadilloFromHell, Armoreno10, Art LaPella, Arthena, Avagad2, Babylonian Armor, BananaFiend, Banaticus, Bart133, Bassbonerocks, Bencherlite, Bendzh, Bensaccount, Bfigura's puppy, Biophysik, Bissinger, BitingHobo, Bjarki S, BlahSpecialK, Bobo192, Bogey97, Boing! said Zebedee, Bongwarrior, BorgQueen, BorisTM, Brauigja, Briannna21a, Bryan Derksen, Burntsauce, Burzmali, Can't sleep, clown will eat me, Canderson7, Carre, Catgut, Cedmudowicz, Ceyockey, Chaldor, Chaos, CharlotteWebb, Chill doubt, Chiros Sunrider, Chris 73, Chris j wood, Chris the speller, Chrislk02, Chronus1029, Cira030, Claire van der Meer, ClickRick, ClockworkSoul, ClockworkTroll, Closedmouth, Collectorben, Conversion script, Coolidays, Cornellrockey, Corpx, Courcelles, Crana, Crohnie, Curiouscorey, Cyclonenim, DARTH SIDIOUS 2, DO11.10, DVD R W, DanielCD, Darkmanx806, Darrentcook, Darth Panda, Daughter of Mmir, Dcandeto, Debresser, Deino Wanthers, DerHexer, Dhatfield, Diberri, Dick Mcgee, Discospinster, Disturbedling, Doctorkismet, Dootv.tv, Doulos Christos, Draeco, Drphilharmonic, Dss971, Dudelock24, Dwmyers, E-lord, E2eamon, EMT1871, Eastssideboy, Echtoran, Elano, Elassint, Eleassar777, Eleuther, Ellywa, Elmyr, Emperor Banh, Enviroboy, Epbr123, Epolk, Erpbridge, Evercat, Everyking, Evil saltine, Evy Surender, Exp HP, Explicit, Faradan, Fat Ted, Fatal!ty, Fenteany, Ferkelparade, Fermion, Fern91, Fieldday-sunday, Firebrother, Firsfron, Flying Canuck, Foochar, Frank, Freakofnurture, Fred Bradstadt, Friginator, G3pro, GB fan, GClugo, Gaelen S., Gaff, Gaius Cornelius, Gd231, Ged UK, Gen. von Klinkerhoffen, Gffkr, Giants27, Giftlite, GillPer, Glagolev, Glane23, GlassCobra, Gludwiczak, Gogo Dodo, GorillaWarfare, Gornly, Graham87, Gwernol, HJ Mitchell, Habj, Hacker312, HaeB, HalfShadow, HangingCurve, Hannah14, HappyCamper, Harvestdancer, HeckXX, Heron, Hodja Nasreddin, Honkbird, HotBabyBoy31097, Howard McCay, Howl5, Hqb, HrishikeshSarode, Hut 8.5, I.am.not.communist, I7988414, Ian Yorston, IcedNut, Immunize, Iridescent, Isfisk, Itfc+canes=me, Ixfd64, J. Spencer, J.delanoy, JLaTondre, JSpudeman, JSpung, JWSchmidt, JYoung130, Jackelfive, Jackfork, Jackol, Jared81, Java7837, Jaw959, Jebba, Jebus989, Jennavecia, Jeppelbaum, JesseW, Jfdwolff, JoanneB, Joao, Joehall45, John Cardinal, John D. Croft, JohnFFields, Johnleemk, JohnnyTehUnicorn, JohnstonDJ, Jonomacdrones, JordanDeLong, Joseph Rudolf, Joshwalker, Jossi, Journeyman, Jugyhng, Juliancolton, Jvbishop, Jyril, Jna runn, Kandar, Karol Langner, Katieh5584, Kavanagh21, Kcox101, Keelan1993, Keilana, Khukri, Knute1232001, Kochipoik, Krashlandon, Krellis, Kubra, Kukini, Kuru, L Kensington, LadyofHats, Lagatapirata, Lahiru k, Laserion, LeaveSleaves, Lectonar, Lee726, Leilimo, Lerdthenerd, Levil, Lightbulb500, Lights, LikeLakers2, LinDrug, Lindmere, Lir, LittleOldMe, Luckyluke, Luigi30, Lukas.S, Luna Santin, MER-C, MONGO, Mac Davis, Magnus Manske, Mani1, MarcoTolo, MarkSutton, Martarius, Masterpiece2000, Materialscientist, MatthiasG, Mav, MaxSem, Maxim, Maxim Razin, Mbtran, McSly, Meaghan, Medical geneticist, Mephistophelian, Merovigla, Mifter, Mikael Hggstrm, Mike Rosoft, Mild Bill Hiccup, Mnlkpoq, Moreschi, Mr. Lefty, Mrees1997, Mrholybrain, Munita Prasad, Myanw, Mygerardromance, Mslimix, N5iln, NSR, Naddy, Nakon, Nasnema, 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Finlay McWalter, GraybeardBiochemist, Gkhan, Ibrmrn3000, Iknewwhereelectricitycomesfrom, Isnow, JaGa, Jag123, Jeppelbaum, Jmarchn, Jojhutton, Jyril, Kawta maderchoud55, Kubanczyk, La goutte de pluie, Leptictidium, Lova Falk, Malljaja, Mcstrother, Michall, Michaplot, Mikael Hggstrm, Mikr18, Mjharrison, Molkhal, Neelix, Nerd, OverlordQ, PL290, Pedrora, Pro crast in a tor, Richard001, Rod57, Rsabbatini, SCEhardt, Sandstein, Scoutersig, Seren-dipper, Shahriyar alavi, Skaaii, Someone87, TFOWR, Taral, TimVickers, Tristanb, Turgonml, Una Smith, Unused0026, Uthbrian, Vojtech.dostal, Wafulz, Waves00, Wimjongman, Wolingfeng, Yale2013, Zanimum, 146 anonymous edits Glycogen Source: http://en.wikipedia.org/w/index.php?oldid=457697601 Contributors: 28421u2232nfenfcenc, AThing, Adamcieslicki, Agathoclea, Ahda, Ahltorp, Akamad, Akxcskier, Ale jrb, Allstarecho, Alteripse, Arbitrarily0, Arcadian, Arthena, BRSM, Banus, Beetstra, Ben Thuronyi, Bencherlite, Bensaccount, Bhadani, BillFlis, Bomac, BorisTM, 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SietskeEN, Sir Vicious, Sjakkalle, Solidpoint, Sonjaaa, Spencerw, Stubblyhead, Taishaku, The sanch, Thegreenj, Tmcclell, Tminus65, Tnxman307, Tonyle, Vedran12, Vrenator, WATerian, Whosasking, Wickey-nl, WikipedianMarlith, Wikiwooroo, WiseUmos, WriterHound, Ww, Xenonice, XxClayTheBossxX, XxNeXuSxX, Zombiejesus, Zuma212, ~K, , 268 anonymous edits Pentose phosphate pathway Source: http://en.wikipedia.org/w/index.php?oldid=455031335 Contributors: -VL-, Adenosine, Anneli1, Arcadian, Bryan Derksen, Cepheus 5, Chtit draco, CopperKettle, Dhorspool, Drphilharmonic, Edgar181, El C, Eras-mus, Eug, Fvasconcellos, Gabbe, GdenBesten, Ge Tianfang, Giftlite, GraybeardBiochemist, Horiavulpe, Isnow, Jag123, Jffre, John Riemann Soong, Kazkaskazkasako, Kupirijo, Lrunge, Materialscientist, MiPe, Narayanese, Nina Gerlach, Nonagonal Spider, PatrciaR, Pcampeau, Pelirojopajaro, Shahriyar alavi, Shoeofdeath, SkyMaja, Steinsky, Tagishsimon, Tahmmo, TimVickers, Time9, Xris0, Yikrazuul, 90 anonymous edits Citric acid cycle Source: http://en.wikipedia.org/w/index.php?oldid=464219279 Contributors: 1029x, 129.186.19.xxx, A bit iffy, AC+79 3888, AManWithNoPlan, Aa77zz, AdamRetchless, Adenosine, AdultSwim, Alansohn, Alba, Aldaron, AlexanderPico, Andrea105, Animum, Antandrus, Arcadian, Arctic Night, Ashcraft, AxelBoldt, Basicsharingwatuknow!, Baxter9, Bdesham, Bensaccount, Bernlomnty, Bluedustmite, Boghog, BorisTM, BrettAllen, Brian0918, Bryan Derksen, Butwhatdoiknow, CDN99, Cadsuane Melaidhrin, CaitlinG, Capricorn42, CardinalDan, Castiron, Catbar, Charleschuck, Chibibrain, Christopher Parham, Christopherlin, ClamDip, Clicketyclack, ClockworkSoul, CoeurDeLion, Conversion script, Cquan, Craigy144, Crana, Curtholr, Cyberman, DGG, DRHagen, DVirus101, Da Joe, DarkoV, Darsie, Davewho2, David D., Dawn Bard, Daycd, DeadEyeArrow, Delta G, Deor, Discospinster, Dj Capricorn, Djmccoul, Dlrohrer2003, Dnvrfantj, Docboat, DrNixon, Drestros power, Drphilharmonic, Dryphi, Dwmyers, ESkog, Eleuther, Elronxenu, 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Croft, John Lake, Johner, Jojit fb, Jose Ramos, Josh Cherry, JuanitaJP, Julianonions, Jyril, Kaidenet, Kandar, Katoa, Kawta maderchoud, Kelovy, Kim Bruning, KoleTang, KrebsCycle, L Kensington, Lachoy11, Lexor, Lir, Luci Sandor, Luigifan, MER-C, Madeinsane, Magicxcian, Magnus Manske, Malljaja, Marek69, Mark Patterson, Marqueed, Mav, MercuryBlue, Mgiganteus1, Michael Shields, Microbrain, MisterDub, Mkweise, Mohawkjohn, Mpatel, Mushin, Narayanese, Nbauman, Nchaimov, Nihiltres, Nike tick, Nilmerg, Nomad2u001, Nucleusboy, ODoyle040, Oatmeal batman, Onco p53, OwenBlacker, Peak, Philip Trueman, Piano non troppo, Pion, Plasmic Physics, Pontificalibus, ProfessorAM, Profoss, Pschemp, Qcomplex5, Rdphair, Rhino8, Rich Farmbrough, Richard001, Rjwilmsi, Rob Hooft, Ronhjones, Ryan Postlethwaite, Ryanreednro, SJP, Sardino, Sbfw, ScAvenger lv, Scattered Lights, Scigatt, Seaphoto, Selket, Semper discens, Serein (renamed because of SUL), Shahriyar alavi, Shrimp wong, Shureg, Skeksys, SkyWalker, Slipstream, Snellios, Sojoho09, Spaully, Spoladore, Stewartadcock, Student7, THEN WHO WAS PHONE?, Tarquin, Technopat, Thatother1dude, The Hybrid, Thegeneralguy, TimVickers, Tknight1011, ToematoeAdmn, Tokhtuev, Trnoriega, Turmerick, Ugen64, Ultratomio, Vanderesch, VsevolodKrolikov, Weird Bird, Wik, WriterHound, YassineMrabet, YellowMonkey, Zarel, Zeeroid, Zephyris, Ziga, 455 anonymous edits

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Oxidative phosphorylation Source: http://en.wikipedia.org/w/index.php?oldid=454686404 Contributors: Abstraktn, Adenosine, Alan Liefting, Alnokta, Antelan, Anton Gutsunaev, Arcadian, Art LaPella, AshLin, Bahar101, Bensaccount, Bobo192, Borgx, BorisTM, Brenont, Brighterorange, Bryan Derksen, Can't sleep, clown will eat me, Chris Rodgers, Clicketyclack, CommonsDelinker, Conversion script, CopperKettle, Crazycomputers, Cst17, DJ Clayworth, DMacks, Danny, Davaith, David D., Dcirovic, Dj Capricorn, Drphilharmonic, Dwmyers, EerieNight, El C, Epbr123, Eribro, Espresso Addict, Etxrge, Eubulides, Faramir333, Fvasconcellos, Gaius Cornelius, Graham87, GraybeardBiochemist, Habj, Harryboyles, Hodja Nasreddin, IKenny, IRP, Ijustam, Imjustmatthew, J.delanoy, JWSchmidt, Jag123, Jamouse, Josh Cherry, Jpg, Julianonions, Jwollbold, Kakli, Kozuch, Laurip, LedgendGamer, Leonard^Bloom, Leptictidium, Lesviolonsdautomne, Lilac Soul, MITBeaverRocks, Macowell, Marj Tiefert, Materialscientist, Maxim Gavrilyuk, Michael Devore, Mikael Hggstrm, Mwucherer, Narayanese, Nhandler, Nono64, Nwbeeson, ParticleMan, Persian Poet Gal, Philgoetz, Piledhigheranddeeper, Pinkadelica, Plugs0, RDBrown, Rakwiki, Rich Farmbrough, Richard001, Richard0612, Rjwilmsi, Semitallsmalls, Shahriyar alavi, Simpsons contributor, Slowpokeiv, Smokefoot, Smurrayinchester, Some jerk on the Internet, Some standardized rigour, StaticGull, Stepa, Stewartadcock, StradivariusTV, Suruena, Tameeria, TimVickers, Treisijs, Ttownfeen, Ulisse0, Verticalstall, Wavelength, Wmpearl, WolfmanSF, WriterHound, Xp54321, Zephyris, 156 anonymous edits Photosynthesis Source: http://en.wikipedia.org/w/index.php?oldid=462428021 Contributors: (jarbarf), *drew, -- April, 1exec1, 28421u2232nfenfcenc, 678910, 9258fahsflkh917fas, @pple, AJRM, ASKingquestions, ATMarsden, Abductive, Abu-Fool Danyal ibn Amir al-Makhiri, Academic Challenger, Accuruss, Adashiel, Adenosine, Agathman, Ahoerstemeier, Aidentbrown, Aitias, Alan Liefting, Alansohn, Alexius08, Ali, 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Anderson, Steven Zhang, Stevenmitchell, Stoic Squirrel, Storkk, Stroppolo, Studentne, SueHay, Sundar, Sunholm, Supernaturalist, Sweet xx, SweetNeo85, Swmcd, Szajci, THEN WHO WAS PHONE?, Tackyleprechaun, Talrias, Tameeria, Tarquin, Tastycheeze, Tattoo275, Tawker, Tbhotch, Tchaika, Tdslk, TedE, Teh wise one, Tellyaddict, Tero, TestPilot, TexasAndroid, Thatotherdude, The Catfish, The Earwig, The Thing That Should Not Be, The sock that should not be, The wub, TheArmadillo, TheTito, Thingg, Tide rolls, TigerShark, Tikoo, Tim Q. Wells, TimVickers, Tito4000, Toadams, Tom Pippens, Tomaschwutz, Tommy2010, Torahjerus14, Touchstone42, Towerofthunder, Tpbradbury, Tra, Tradnor, Trappist, Traroth, Treisijs, Tristanb, Troy 07, Twirligig, U.S.Vevek, Uiteoi, Ulric1313, Uncle Dick, Unclepea, Unyoyega, Upeksha112, Usdi, User A1, Username314, Valrie75, Van helsing, Vanished user, Vary, Vices, Viper777, VolcomRUXC, Vortexrealm, Vsmith, Vuvar1, Waggers, WatermelonPotion, Wavelength, Wejer, WelshMatt, White.matthew.09, Whosyourjudas, Wiitennisxd, WikHead, Wiki alf, WikiLaurent, Wikidenizen, Wikidudeman, Wikipe-tan, WikipedianMarlith, William Avery, Wimt, Wine Guy, Wk muriithi, Woohookitty, Woudloper, WriterHound, Wutsje, Xchbla423, Xezbeth, Xiong Chiamiov, Xoddam, Yamaguchi, Yamamoto Ichiro, Yerpo, Yikrazuul, Yon Yonson, Z 153, Zack, Zap Rowsdower, Zarniwoot, Zedla, Zepheus, Zfr, ZooFari, Zvn, Zxoxm, , 2154 anonymous edits Fatty acid synthesis Source: http://en.wikipedia.org/w/index.php?oldid=450714507 Contributors: AlexanderPico, Arcadian, AvicAWB, Azo bob, Bochyboch, Chodid, Clicketyclack, Colinc719, Cow2001, Ctrlaltdelete200390, D6, Dcirovic, Drphilharmonic, Fizzyfifi, Giraffedata, GraybeardBiochemist, Hubba, Itub, Jaypg, John of Reading, Knopfkind, Kubanczyk, Leafyplant, Luci Sandor, Mtuttle, NikeTenis, OG Clriley, Ojii-san, Panoramix303, Pro crast in a tor, RE73, RShorty30, Rjwilmsi, Skaaii, SkyMaja, Tommy2010, Yikrazuul, Zachlipton, 39 anonymous edits Lipogenesis Source: http://en.wikipedia.org/w/index.php?oldid=459445784 Contributors: Amog, Arcadian, Chodid, Colinc719, Ctrlaltdelete200390, Drphilharmonic, Ed7654, Gensanders, Gkhan, Keenan Pepper, Ksero, Kubanczyk, Leptictidium, LittleT889, Paul August, Pelirojopajaro, Plico, RDBrown, Rich Farmbrough, Shrimp wong, Snellios, 61 anonymous edits Acetyl-CoA carboxylase Source: http://en.wikipedia.org/w/index.php?oldid=447826665 Contributors: Adeez, Alexhlau, Alnokta, Arcadian, Bfx0, Boghog, BorisTM, EagleFan, Edward, Frietjes, GraybeardBiochemist, Im.a.lumberjack, J G Campbell, Jimhsu77479, Luuva, M gehrig2000, Mikael Hggstrm, Muhandes, Mushin, Nw.NPC, RainR, Rb1248, Reaper Eternal, Saxbryn, Sbmehta, Shrimp wong, SimonP, Slaporte, The wub, Tom harrison, Waldroplab, WikHead, Woohookitty, Xris0, Yottie, 29 anonymous edits Fatty acid degradation Source: http://en.wikipedia.org/w/index.php?oldid=418315617 Contributors: Arcadian, Clicketyclack, Haripandit, Kubanczyk, Mikael Hggstrm, Nick Number, Rich Farmbrough, Richard001, Synchronism, ThinkerThoughts, Walkerma, Wmpearl, WolfmanSF, , 4 anonymous edits Beta oxidation Source: http://en.wikipedia.org/w/index.php?oldid=464183482 Contributors: -VL-, AC+79 3888, Abdull, Abrech, Arcadian, Aurista25, Beamoflaser, Bogdangiusca, Bunnyhop11, C.Fred, Chibibrain, Christian75, Clicketyclack, ClockworkSoul, Coolstoryhansel, Darklilac, Dawhitfield, Dcirovic, Dmanagadze, Dono, Dr. Ambitious, Drocra, Drphilharmonic, Eumeng.chong, Grafen, GraybeardBiochemist, Haripandit, HkCaGu, Hyalos, Jeyradan, Jstanley, Krylonblue83, Langthorne, Lecj77, Lemchesvej, LinkinPark, LoneSeeker, Magairlin, Mastee, MauchoEagle, Meaganrock, Meph636, Mushin, NEUROtiker, Narfaniel, Natarajanganesan, Nitroblu, Oddity-, Paxse, Physchim62, RShorty30, Silkblackrose, Spongefrog, Stepa, Su-no-G, Taskualads, TimVickers, Tomifly, V8rik, Velella, Vivekanand23, Vojtech.dostal, Wmpearl, Zerkalox, Zink53, Zoidbergness, 73 anonymous edits Nitrogen fixation Source: http://en.wikipedia.org/w/index.php?oldid=463457356 Contributors: 129.128.164.xxx, 2D, A2Kafir, Alansohn, Andrewlevy, Andyman1125, AnonGuy, Ashmoo, Ashnard, Atubeileh, Aur, Avinashraut, Bakabaka, Beland, Bender235, Bendzh, Bkurl, Bob Burkhardt, Br77rino, Brian Crawford, Bryan Derksen, Butterflied, C S, Cacycle, Callior, CanisRufus, Charfroman, Chester Markel, CinchBug, Clicketyclack, Conversion script, DARTH SIDIOUS 2, DarkAudit, Devtrash, Dj Capricorn, Dougluce, Drmies, Drphilharmonic, Dvorak729, Dysmorodrepanis, Ekawolfram, Epbr123, Excirial, Fancy steve, FrYGuY, Frappyjohn, Fumitol, Gareth.randall, Guarwolf, Haikon, Headbomb, Heathmoor, Heterotheca, Igodard, Ilyushka88, J.delanoy, J04n, JForget, JamesBWatson, Jusdafax, Kappa, Katoa, Kazvorpal, Kbh3rd, Keegan, Keenan Pepper, Kupirijo, L Kensington, LOL, Lab-oratory, M uzair, MPF, Malcolm Farmer, Manuel Anastcio, Mapetite526, Marcjarod, MarcoTolo, Marshman, Melaen, Michael C Price, Michael Hardy, Midnightdreary, Mork the delayer, Mouse Nightshirt, Mspraveen, NawlinWiki, Ninjatacoshell, Nono64, Notinasnaid, Onco p53, Ortolan88, Paleorthid, Parisoil, PhilKnight, Pinethicket, PipOC, Prari, Pseudomonas, Puppy8800, Quercusrobur, RHaden, RandomP, Rich Farmbrough, RobertG, Rosser1954, RucasHost, RyJones, Sabedon, Satyrium, SchoonerGuy, ScottSteiner, Securiger, Seraphim, Shinkolobwe, Shinpah1, SimonP, Skater, Slightsmile, Smack, Smartse, Smokefoot, Smurfsahoy, So God created Manchester, Sonett72, Spencer, Stone, Tetracube, Thomasn1, Tide rolls, Tideflat, Tigerente, TimTL, TimVickers, Tom Pippens, Topbanana, Touchstone42, True Pagan Warrior, Tsugasdad, Utility Monster, V8rik, Veledan, Velella, Vsmith, Wavelength, Weetziekat, Why Not A Duck, Wingedsubmariner, Woohookitty, WormRunner,

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XRK, Yersinia, Yobol, Zazil, Zombiejesus, 270 anonymous edits Amino acid synthesis Source: http://en.wikipedia.org/w/index.php?oldid=441364518 Contributors: Arcadian, Avs5221, Dan East, Diderot's dreams, Drphilharmonic, Edgar181, Elonka, Giftlite, Keenan Pepper, Leptictidium, Littlealien182, Mirabellen, Nutriveg, Pedro-kun, Phlounder, Redheylin, Rockfang, Shrimp wong, Welsh, Wingedsubmariner, Woohookitty, Zephyris, 14 anonymous edits Nucleotide Source: http://en.wikipedia.org/w/index.php?oldid=464450294 Contributors: 2T, Access Denied, AdamRetchless, Adenosine, Adrian J. 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Rgocs, Riana, Ribrob, Richard001, Rida97, Rmky87, Rod57, Ronz, Rosieredfield, RyanGerbil10, S73v3n, SIMONCOHEN, SU Linguist, Sakkura, Sameerbau, Saurabh523, Scresawn, Seans Potato Business, Segabud, Shanel, Sjakkalle, Sl, Smartse, Smelissali, Sophos II, Squidonius, Sriram sh, Srlasky, Staffwaterboy, Steinsky, Sydney3803, Tedtoal, Teethies563, Teply, TestPilot, The Anome, The Flying Spaghetti Monster, The undertow, Thingg, This lousy T-shirt, Tide rolls, TimVickers, Timemutt, Tmh, Tomjc, Totophe64, Transitiveinstance, Tycho, Useight, Uthbrian, Vanished user 39948282, Vidric, Vikky2904, Vojtech.dostal, WAS 4.250, WAvegetarian, Wayne Slam, WikHead, Woohookitty, Wouterstomp, Yerpo, Yided, Yvorez1274, Z.E.R.O., Zamftb, Zashaw, Zephyris, , , 635 anonymous edits Regulation of gene expression Source: http://en.wikipedia.org/w/index.php?oldid=463581999 Contributors: Aarre, Active contributor, Agathman, Ale jrb, AndreasJS, Annatyler23, Arcadian, Arjayay, AxelBoldt, Ayush girish nagar, 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Pupunwiki, RDBrown, Rasmusw, Rjwilmsi, Rolando, Saippuakauppias, Salt Yeung, Sanbioinfo, Secundus Zephyrus, Shadowelve11, SimonP, Sjdk13, SomethingCatchy, Sord halo, Squidonius, Stroppolo, Superdoggy, Sverdrup, Taoster, TenOfAllTrades, The Anome, Thisismikesother, Timemutt, Timwi, TransControl, Trinit, Uncle-P, Unfortunate, Uraza, Vossman, Wavelength, Wickey-nl, Xiahou, YaacovCR, Yashgaroth, Yasingam, Yontally, ZayFoTT, Zephyris, 269 anonymous edits Posttranslational modification Source: http://en.wikipedia.org/w/index.php?oldid=452718420 Contributors: A876, Alon Gabbay, Amikake3, Aquaphobic, Arcadian, Arminius, Banus, Bdekker, Bensaccount, Bromomir, Bryan Derksen, Bsadowski1, C31004, Ceyockey, Clicketyclack, Dcirovic, Dimethylformamide, Edgar181, Epingchris, GAThrawn22, Geobacter, Glane23, Graft, Hodja Nasreddin, Jfdwolff, Jni, Jolivio, Jonkerz, Julesd, Kaarel, Kkmurray, Kosigrim, KuduIO, Kupirijo, La goutte de pluie, Lemchesvej, Madeleine Price Ball, Magnus Manske, MarXidad, Mike Serfas, Minnsurfur2, PFHLai, Pgan002, Reinoutr, Rich Farmbrough, Rjwilmsi, Shuvaev, Silly shrimp, SimonP, Social tamarisk, Stewartadcock, Stillnotelf, Thewildtype, Tuomas Htinen, Vermiculus, WillowW, Wmpearl, Xwu, Yyadam7, Yyocean, Zephyris, 61 anonymous edits Proteolysis Source: http://en.wikipedia.org/w/index.php?oldid=433412130 Contributors: 217.228.14.xxx, Actarux, Algont, Almazi, Arcadian, Bryan Derksen, Champ0815, Clicketyclack, Conversion script, Edward, EdwardLane, Habj, Hirokun, JohnOwens, Jotomicron, Jwinius, Kosigrim, Marj Tiefert, Martious, Maxamegalon2000, Michael Hardy, Nina Gerlach, Nono64, PaddyM, PhD Dre, Rcej, Roadnottaken, SimonP, Stepa, Svartkell, WillowW, 21 anonymous edits Proteasome Source: http://en.wikipedia.org/w/index.php?oldid=461878424 Contributors: Aa77zz, AdamRetchless, Alai, Alansohn, Anthonyhcole, Ashanda, BCvek, Barticus88, Bender235, Berkay0652, Biochemza, Biogvk, BloodIce, Bobblewik, Bobmarley1987, BostonBiochem, Brighterorange, Bryan Derksen, CBM, 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Image:Sucrose-inkscape.svg Source: http://en.wikipedia.org/w/index.php?title=File:Sucrose-inkscape.svg License: GNU Free Documentation License Contributors: Don A. 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Image:Mars Hubble.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Mars_Hubble.jpg License: Public domain Contributors: NASA and The Hubble Heritage Team (STScI/AURA) Image:Heart of Steel (Hemoglobin).jpg Source: http://en.wikipedia.org/w/index.php?title=File:Heart_of_Steel_(Hemoglobin).jpg License: GNU Free Documentation License Contributors: Photographer: Julian Voss-Andreae; Uploaded to en-wiki by en:User:Julianva; Uploaded to Commons by User:Brainmachine Image:Enzyme catalysis delta delta G.png Source: http://en.wikipedia.org/w/index.php?title=File:Enzyme_catalysis_delta_delta_G.png License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Original uploader was Zephyris at en.wikipedia Image:Induced fit diagram.svg Source: http://en.wikipedia.org/w/index.php?title=File:Induced_fit_diagram.svg License: Public Domain Contributors: Created by TimVickers, w:vector graphicsvectorized by Fvasconcellos Image:Enzyme catalysis uniform+differential binding.png Source: http://en.wikipedia.org/w/index.php?title=File:Enzyme_catalysis_uniform+differential_binding.png License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Original uploader was Zephyris at en.wikipedia Image:Lysozyme transition state.png Source: http://en.wikipedia.org/w/index.php?title=File:Lysozyme_transition_state.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia Image:Inter vs intramolecular reaction rates.png Source: http://en.wikipedia.org/w/index.php?title=File:Inter_vs_intramolecular_reaction_rates.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia Image:Serine protease catalysis.png Source: http://en.wikipedia.org/w/index.php?title=File:Serine_protease_catalysis.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia Image:Carboxypeptidase catalysis.png Source: http://en.wikipedia.org/w/index.php?title=File:Carboxypeptidase_catalysis.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia Image:EcDHFR raytraced.png Source: http://en.wikipedia.org/w/index.php?title=File:EcDHFR_raytraced.png License: Public Domain Contributors: TimVickers Image:KinEnzymo(en).svg Source: http://en.wikipedia.org/w/index.php?title=File:KinEnzymo(en).svg License: Public Domain Contributors: TimVickers, YassineMrabet, Image:Enzyme progress curve.svg Source: http://en.wikipedia.org/w/index.php?title=File:Enzyme_progress_curve.svg License: Copyrighted free use Contributors: en:User:Poccil, Based on public domain JPG by TimVickers. 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Image:Reversible inhibition.svg Source: http://en.wikipedia.org/w/index.php?title=File:Reversible_inhibition.svg License: Public Domain Contributors: User Poccil on en.wikipedia Image:Activation2 updated.svg Source: http://en.wikipedia.org/w/index.php?title=File:Activation2_updated.svg License: GNU Free Documentation License Contributors: Originally uploaded by Jerry Crimson Mann, vectorized by Tutmosis, corrected by Fvasconcellos File:Common lipids lmaps.png Source: http://en.wikipedia.org/w/index.php?title=File:Common_lipids_lmaps.png License: GNU Free Documentation License Contributors: Eoin Fahy Original uploader was Lmaps at en.wikipedia File:Phosphatidyl-Ethanolamine.png Source: http://en.wikipedia.org/w/index.php?title=File:Phosphatidyl-Ethanolamine.png License: Public Domain Contributors: Original uploader was Jag123 at en.wikipedia File:Sphingomyelin-horizontal-2D-skeletal.png Source: http://en.wikipedia.org/w/index.php?title=File:Sphingomyelin-horizontal-2D-skeletal.png License: Public Domain Contributors: Benjah-bmm27, 1 anonymous edits File:Kdo2-lipidA.png Source: http://en.wikipedia.org/w/index.php?title=File:Kdo2-lipidA.png License: Creative Commons Attribution-Sharealike 3.0 Contributors: en:User:Lmaps and User:TimVickers File:Phospholipids aqueous solution structures.svg Source: http://en.wikipedia.org/w/index.php?title=File:Phospholipids_aqueous_solution_structures.svg License: Public Domain Contributors: Mariana Ruiz Villarreal ,LadyofHats Image:2r9r opm.gif Source: http://en.wikipedia.org/w/index.php?title=File:2r9r_opm.gif License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Andrei Lomize Image:Cell membrane detailed diagram 4.svg Source: http://en.wikipedia.org/w/index.php?title=File:Cell_membrane_detailed_diagram_4.svg License: Creative Commons Attribution-Sharealike 3.0 Contributors: derivative work: Dhatfield (talk) Cell_membrane_detailed_diagram_3.svg: *derivative work: Dhatfield (talk) Cell_membrane_detailed_diagram.svg: LadyofHats Mariana Ruiz Image:Fluid Mosaic.svg Source: http://en.wikipedia.org/w/index.php?title=File:Fluid_Mosaic.svg License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Jerome Walker Image:Alpha Intercalated Cell Cartoon.svg Source: http://en.wikipedia.org/w/index.php?title=File:Alpha_Intercalated_Cell_Cartoon.svg License: Creative Commons Attribution 3.0 Contributors: Rswarbrick Image:Membrane lipids.png Source: http://en.wikipedia.org/w/index.php?title=File:Membrane_lipids.png License: Public Domain Contributors: BorisTM, Gwernol, WOSlinker, 2 anonymous edits Image:Lactose.svg Source: http://en.wikipedia.org/w/index.php?title=File:Lactose.svg License: Public Domain Contributors: Calvero. 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File:1GZX Haemoglobin.png Source: http://en.wikipedia.org/w/index.php?title=File:1GZX_Haemoglobin.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia File:Catabolism schematic.svg Source: http://en.wikipedia.org/w/index.php?title=File:Catabolism_schematic.svg License: Public Domain Contributors: Tim Vickers, vectorized by File:Plagiomnium affine laminazellen.jpeg Source: http://en.wikipedia.org/w/index.php?title=File:Plagiomnium_affine_laminazellen.jpeg License: GNU Free Documentation License Contributors: Kristian Peters -- Fabelfroh File:Sterol synthesis.svg Source: http://en.wikipedia.org/w/index.php?title=File:Sterol_synthesis.svg License: Public Domain Contributors: , original by Tim Vickers File:Insulin glucose metabolism ZP.svg Source: http://en.wikipedia.org/w/index.php?title=File:Insulin_glucose_metabolism_ZP.svg License: Public Domain Contributors: Original uploader was XcepticZP at en.wikipedia File:Tree of life int.svg Source: http://en.wikipedia.org/w/index.php?title=File:Tree_of_life_int.svg License: Creative Commons Attribution-Sharealike 3.0,2.5,2.0,1.0 Contributors: myself, based on TimVickers's work. 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Image:ATPsyn.gif Source: http://en.wikipedia.org/w/index.php?title=File:ATPsyn.gif License: Public Domain Contributors: Original uploader was TimVickers at en.wikipedia File:Seawifs global biosphere.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Seawifs_global_biosphere.jpg License: unknown Contributors: Luis Fernndez Garca, Tano4595, TheDJ, Trelio, Yikrazuul, 1 anonymous edits File:Photosynthesis equation.svg Source: http://en.wikipedia.org/w/index.php?title=File:Photosynthesis_equation.svg License: Public Domain Contributors: ZooFari File:Photosynthesis.gif Source: http://en.wikipedia.org/w/index.php?title=File:Photosynthesis.gif License: Creative Commons Attribution-Sharealike 3.0 Contributors: User:At09kg File:Simple photosynthesis overview.svg Source: http://en.wikipedia.org/w/index.php?title=File:Simple_photosynthesis_overview.svg License: Creative Commons Attribution-Share Alike Contributors: Daniel Mayer (mav) - original imageVector version by Yerpo 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File:Main symptoms of diabetes.png Source: http://en.wikipedia.org/w/index.php?title=File:Main_symptoms_of_diabetes.png License: Public Domain Contributors: Mikael Hggstrm Image:Suckale08 fig3 glucose insulin day.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Suckale08_fig3_glucose_insulin_day.jpg License: Creative Commons Attribution-Sharealike 3.0 Contributors: Jakob Suckale, Michele Solimena Image:Glucose-insulin-release.png Source: http://en.wikipedia.org/w/index.php?title=File:Glucose-insulin-release.png License: GNU Free Documentation License Contributors: Original uploader was Prisonblues at en.wikipedia File:Diabetes world map - 2000.svg Source: http://en.wikipedia.org/w/index.php?title=File:Diabetes_world_map_-_2000.svg License: Creative Commons Attribution-Sharealike 2.5 Contributors: Lokal_Profil Image:Diabetes mellitus world map - DALY - WHO2004.svg Source: http://en.wikipedia.org/w/index.php?title=File:Diabetes_mellitus_world_map_-_DALY_-_WHO2004.svg License: Creative Commons Attribution-Sharealike 2.5 Contributors: Lokal_Profil File:Loudspeaker.svg Source: http://en.wikipedia.org/w/index.php?title=File:Loudspeaker.svg License: Public Domain Contributors: Bayo, Gmaxwell, Husky, Iamunknown, Mirithing, Myself488, Nethac DIU, Omegatron, Rocket000, The Evil IP address, Wouterhagens, 16 anonymous edits Image:DNA replication split.svg Source: http://en.wikipedia.org/w/index.php?title=File:DNA_replication_split.svg License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Madprime File:DNA replication.theora.ogv Source: http://en.wikipedia.org/w/index.php?title=File:DNA_replication.theora.ogv License: Creative Commons Attribution-Sharealike 3.0 Contributors: Cookatoo.ergo.ZooM Image:DNA polymerase.svg Source: http://en.wikipedia.org/w/index.php?title=File:DNA_polymerase.svg License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Madprime Image:DNA replication en.svg Source: http://en.wikipedia.org/w/index.php?title=File:DNA_replication_en.svg License: Public Domain Contributors: LadyofHats Mariana Ruiz Image:1axc tricolor.png Source: http://en.wikipedia.org/w/index.php?title=File:1axc_tricolor.png License: Creative Commons Attribution-ShareAlike 3.0 Unported Contributors: Opabinia regalis Image:Cell Cycle 2.png Source: http://en.wikipedia.org/w/index.php?title=File:Cell_Cycle_2.png License: GNU Free Documentation License Contributors: Original uploader was Zephyris at en.wikipedia Image:brokechromo.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Brokechromo.jpg License: GNU Free Documentation License Contributors: Multichill, Square87, Wickey, 2 anonymous edits Image:Ssvsds.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Ssvsds.jpg License: GNU Free Documentation License Contributors: Bestiasonica, Bkell, Square87 Image:Uracil base glycosidase.jpg Source: http://en.wikipedia.org/w/index.php?title=File:Uracil_base_glycosidase.jpg License: Public Domain Contributors: Original uploader was TimVickers at en.wikipedia Image:DNA Repair.jpg Source: http://en.wikipedia.org/w/index.php?title=File:DNA_Repair.jpg License: Public Domain Contributors: Courtesy of Tom Ellenberger, Washington University School of Medicine in St. Louis. 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