Mitochondria, ATP, Membrane Potential and Cell Volume

ESSAY
GIVE AN ACCOUNT OF MITOCHONDRIAL STRUCTURE AND ATP

SYNTHETIC FUNCTION AND DESCRIBE THE IMPORTANCE OF ATP IN
THE MAINTENANCE OF MEMBRANE POTENTIAL AND CELL VOLUME.
Copyright ©Anna Hunter 2010, Trinity College, Dublin 2, Ireland. All rights reserved.
Introduction
This essay aims to describe the structure and overall function of mitochondria in cells and
their unique involvement in the synthesis of Adenosine Triphosphate (ATP). Mitochondria
are organelles found in the cytoplasm of each cell and function in cellular respiration. ATP is
an energy source produced during cellular respiration. ATP has many bearings on the overall
function of the cell including the membrane potential and cell volume.
This essay aims to describe in detail the synthesis of ATP through the complex processes of
glycolysis, the citric acid cycle and oxidative phosphorylation. Glycolysis occurs in the
cytoplasm and results in the production of a molecule called pyruvate which then moves into
the matrix of the mitochondria and begins a series of reactions called the citric acid cycle
resulting in the formation of ATP.
The goal will be to inform the reader of the complex reactions that occur to generate ATP.
The Structure of Mitochondria
Mitochondria are an essential organelle in all cells of the body, bar mature red blood cells, as
they function to produce most of the energy of the cell. Mitochondria vary in size and shape,
but are mostly elongated oval – shaped organelles. Mitochondria are mobile, thus circulating
the cell and accumulating at the main sites of energy requirement. For example, liver cells
can contain as many as 2000 mitochondria whereas inactive cells will contain much fewer 1
Although mitochondria vary considerably in their size and shape, their structure is the same,
each consisting of an outer membrane, an inner membrane, a narrow intermembranous space
and the matrix. The outer membrane is a smooth, relatively permeable membrane as it
contains porin, a pore – forming protein, therefore allowing small molecules to pass through.
Enzymes present in this outer membrane act on and convert particular lipid substrates into
forms that can be metabolised in the mitochondrion1. The inner membrane is comprised of a
series of many folds, called cristae, that project into the central cavity, the matrix. Aerobic
respiration occurs on this inner membrane, where the large surface area of the cristae
enhances the process2. It contains the cytochromes, the carrier molecules of the electron
transport chain, as well as enzymes that are involved in the production of ATP. The inner
cavity is called the matrix of the mitochondrion and aerobic respiration occurs here. It
contains enzymes for the Kreb’s cycle in the production of ATP. The matrix also contains
matrix granules, thought to be the binding sites for the storage of calcium1). The
intermembranous space contains enzymes for metabolism.
Within the matrix is found one or more circular double – stranded molecules of DNA as well
as ribosomes. It is known that 2 – 10 copies of mitochondrial genomes codes for 37 genes.
Thus mitochondria synthesise 37 of their own constituent proteins.
Mitochondria replicate independently. Their main functions, as mentioned above, are calcium
storage, apoptosis and aerobic respiration. It is for this reason that mitochondria may be
referred to as “factories of ATP”.
academic.brooklyn.cuny.edu/.../page/mito.htm
The Synthesis of ATP
Glycolysis
In cellular respiration, energy released by the catabolic reactions that break down glucose and
fatty acids is used to synthesise ATP. The breakdown of glucose for energy begins with a
metabolic process called glycolysis which occurs in the cytoplasm. It is a metabolic pathway
involving various enzymatically controlled steps.
Stage 1: Glucose enters the cell and is phosphorylated by ATP to form glucose 6 –phosphate.
This addition of a phosphoryl group means that the glucose molecule is now destabilised so
as to promote its further metabolism.3
http://www.sparknotes.com/biology/cellrespiration/glycolysis/section1.rhtml
glucose 6 – phosphate is then isomerised to fructose 6 – phosphate.4
This isomeration reaction is proceeded by the phosphorylation of fructose 6 – phosphate by

ATP to fructose 1,6 – bisphosphate.5
Stage 2: Fructose 1,6 – bisphospphate is split into glyceraldehyde 3 – phosphate and
dihydroxyacetone phosphate, both three – carbon units rather than six – carbon units.5
Stage 3: Glyceraldehyde 3 – phosphate is now converted into 1,3 – bisphosphoglycerate.6
1,3 – bisphosphoglycerate is an energy rich molecule with a greater phosphoryl –

transferpotential than that of ATP, therefore 1,3 – bisphosphoglycerate is used to facilitate the
production of ATP from ADP. The products are ATP and 3 – phosphoglycerate.7
The final steps of the glycolitic pathway sees the conversion of 3 – phosphoglycerate into
pyruvate and the production of a second ATP molecule.8
Each molecule of pyruvate contains three carbons, three oxygens and four hydrogens. The six
carbons and six oxygens in glucose (C6H12O6) are all accounted for between the two pyruvate
molecules but only eight hydrogens out of the twelve are accounted for. Four hydrogen atoms
are removed from the intermediates in glycolysis. Each of these hydrogen atoms is used to
reduce a molecule of nicotinamide adenine dinucleotide (NAD+). This process involves the
reduction of NAD. The reduced NAD (NADH) binds one proton from the hydrogen atoms
and thus leaves a proton. Glycolysis results in the production of 2 NADH molecules and 2
H+. Since glycolysis is an exergonic reaction some of its energy is used to drive the
phosphorylation of ADP to ATP: ADP + Pi ATP. So at the end of glycolysis there is a net
of 2ATP molecules per molecule of glucose. However, glucose must be “activated” at the
start of the glycolitic pathway before any energy may be gained, requiring the addition of two
phosphate groups, obtained from 2ATP molecules. This energy is initially consumed at the
beginning of glycolysis from ATP ADP + Pi.9
Further on in glycolysis 4 more molecules of ATP are formed and 2NADH molecules,
resulting in the gain of 2ATP molecules and 2NADH + H+ molecules per glucose molecule.
Two molecules of pyruvate are also formed. 10
Kreb’s Citric Acid Cycle
The pyruvic acid molecule then leaves the cell cytoplasm and enters the matrix of the
mitochondria where CO2 is removed from each three – carbon pyruvate to form a two –
carbon organic acid, called acetic acid. This acetic acid is the combined with a coenzyme,
called coenzymeA, resulting in the formation of the molecule acetyl coenzymeA.12
Since there are two molecules of pyruvate, two molecules of acetyl coenzymeA are formed
along with two molecules of CO2. On formation of acetyl coenzymeA, the acetic acid subunit
combines with oxaloacetic acid to form a molecule of citric acid, beginning a cyclic
metabolic pathway called the citric acid cycle or Kreb’s cycle. The proceeding reactions that
occur involve the elimination of two molecules of CO2 and removal of hydrogens. A
molecule of guanosine Triphosphate (GTP) is formed and this donates a phosphate group to
ADP to the production of 1ATP. During these reactions three molecules of NAD are reduced
to NADH and one molecule of flavin adenosine dinucleotide (FAD) is reduced to FADH2.13
The Kreb’s cycle does not generate a large amount of ATP but removes electrons from
acetyl coenzymeA and uses them to produce NADH and FADH2. Six electrons are
transferred to three molecules of NAD+ and two electrons i.e. a pair of hydrogen atoms are
transferred to one molecule of FAD. During the process of oxidative phosphorylation these
two electron carriers produce nine molecules of ATP when oxidised by oxygen.14
When pyruvate is transported into the mitochondria it is oxidatively decarboxylated by the

pyruvate dehydrogenase complex to form acetyl coenzymeA:
Pyruvate+CoA+NAD+ Acetyl CoA+CO2+NADH+H+ 15
The citric acid cycle commences with the condensation of a four – carbon unit, called
oxaloacetate, and acetyl CoA to produce citrate (citric acid), a six – carbon unit. Citrate is
then isomerised into isocitrate so as to enable this six – carbon unit to undergo oxidative
decarboxylation. Water is added to citrate to form cis – Aconitic acid and this is dehydrated
to form the isocitrate. Oxidative decarboxylation of isocitrate yields α – ketogluatrate:
Isocitrate+NAD+ α – ketogluatrate+CO2+NADH 16
Oxidative decarboxylation of α – ketogluatrate forms succinyl coenzymeA, with the release

of a molecule of CO2. 16
http://www.sparknotes.com/biology/cellrespiration/citricacidcycle/section2.rhtml
Succinyl coenzymeA is an energy rich thioester compound and this thioester bond is cleaved
by orthophosphate, producing succinate and generating GTP, a high - phosphoryl – transfer -
potential compound. Succinate is oxidised to fumerate with FAD being the proton acceptor.
Fumerate is then dehydrated to form malate, which is then oxidised, hence regenerating
oxaloacetate and initiating the cycle once again.17
For every one glucose molecule we get two Kreb’s cycles and for each turn we get 1ATP,
3NADH and 1FADH2. The net reaction of the citric acid cycle is:
Acetyl CoA+3NAD++FAD+GDP+Pi+2H2O 2CO2+3NADH+FADH2+GTP+2H++CoA
employees.csbsju.edu/.../ch112/CH112_OLSG.htm
So from one turn of the Kreb’s cycle we can see that only one molecule of ATP is produced.
However humans require about 83kg of ATP per day. We only possess about 250g of the
required ATP so this discrepancy is resolved by recycling ADP back to ATP, mainly by the
process of oxidative phosphorylation.
Oxidative Phosphorylation
Oxidative phosphorylation is the process in which ATP is formed as a result of the transfer of
electrons from NADH or FADH2 to O2 by a series of electron carriers. This process yields 26
of the 30 molecules of AT that are produced on the complete oxidation of glucose to CO2 and
water. Electrons are transferred from NADH to O2 via a chain of three protein complexes:
NADH – Q oxidoreductase, Q – cytochrome c oxidoreductase, and cytochrome oxidase,
complexes I, III and IV respectively. Electron flow within these complexes results in the
transport of protons across the inner mitochondrial membrane. A fourth protein complex
called succinate – Q reductase contains the succinate dehydrogenase which generates FADH2
in Kreb’s citric acid cycle. This is referred to as complex II.
Electrons from NADH enter the electron transport chain at NADH – Q oxidoreductase, an L
– shaped proton pump. The two electrons are transferred to flavin mononucleotide (FMN),
which is then reduced to FMNH2. NADH – Q oxidoreductase also contains Fe – S centres and
the electrons emerge in QH2, the reduced form of ubiquinone, an electron carrier.18
Complex II, succinate – Q reductase, does not transfer protons but contains succinate
dehydrogenase, which formed FADH2 in the citric acid cycle. Electrons from FADH2 are
transferred to ubiquinone to form QH2 for entry into the transfer chain. QH2 transfers
electrons to Q – cytochromes c oxidoreductase which reduces cytochromes c, donating its
protons to cytochrome c oxidase. From here electrons are transferred to O2, the final electron
acceptor, forming water.19
The flow of electrons through the proton pumps leads to the transfer of protons from one side
of the inner mitochondrial membrane (matrix side) to the other (cytoplasmic side), leading to
a proton – motive force consisting of a pH gradient (matrix side basic) and a voltage gradient
(or membrane potential). The matrix side is positively charged while the cytoplasmic side is
negatively charged. The flow of protons back to the matrix side through ATP synthase,an
enzyme complex in the inner membrane, drives the synthesis of ATP.20 ATP synthase is
composed of a rotating component (fo unit) and a stationary component (f1 unit).
www.biologie.uni-hamburg.de/b-online/e19/19d.htm
The fo unit contains a proton channel through which protons enter. The c subunit rotates,
promoting the rotation of the γ subunit, thus inducing structural changes in the β subunit,
resulting in the synthesis and release of ATP.
The γ subunit rotates 120° anticlockwise, converting the T – form site to an O – form site and
enabling the combination of ADP + Pi to form ATP. The same process occurs with the L –
form site. For every full turn, one molecule of ATP is generated.21
www.bio.miami.edu/.../255atp/ATPase/ATPase.htm
The total amount of ATP generated is 30 molecules per one glucose molecule. ATP functions
in many cellular reactions and other aspects of cell regulation. ATP is required, for example
on active transport, the movement of ions and molecules against their concentration gradients
from lower concentration to higher concentration.
Membrane Potential
Membrane potential is the electric potential existing across the two sides of a membrane
resulting from the unequal distribution of charge across the membrane.
Primary active transport is entirely ATP dependent. An example is the Na+/K+ pump, an
ATPase, which is controlled by the addition and removal of phosphate groups obtained from
ATP. These ATPases use ATP hydrolysis to drive the thermodynamically uphill transport of
ions. 22
Phosphate groups of ATP have a negative charge at the pH of the cytoplasm. These anions
are set within the cell, attracting inorganic positively charged ions from extracellular fluid
which pass through ion channels in the plasma membrane. Thus the fixed anions influence
the distribution of the inorganic cations.
The Na+/K+ pump breaks down ATP for energy. Cells contain a high K+ concentration and
low Na+ concentration relative to the external medium. The Na+/K+ pump uses the hydrolysis
of ATP to provide energy for the active transport of Na+ out o the cell and K+ into the cell,
generating the gradients. The Na+/K+ pump is an example of an ATPase which becomes
momentarily phosphorylated during the transport process.23
Na+ in the cytoplasm binds to the protein and stimulates phosphorylation by ATP, causing the
protein to alter its structure and expelling the Na+. The K+ then binds to the protein,
stimulating the release of a phosphate group, therefore restoring the protein’s original
conformation. K+ is released and the cycle recommences.
legacy.owensboro.kctcs.edu/GCaplan/anat/Notes...
More than a third of the ATP consumed is used to pump these ions. The Na+/K+ gradient also
regulates cell volume. Osmotic pressure of the intracellular fluid and extracellular fluid is
comparable to 280 mOsm/L. As mentioned above the hydrolysis of ATP drives the Na+/K+
pumps, thus maintaining osmotic balance.
Cell Volume
Cells contain a significant concentration of proteins and organic phosphates that cannot
escape from the cell. The high intracellular concentration of non – diffusible anions results in
a Gibbs – Donnan equilibrium across the cell membrane. At equilibrium there will be high
osmolality intracellularly, causing water to enter the cell and so the cell would swell, thus
upsetting the Gibbs – Donnan equilibrium. More solute particles (sodium) would enter the
cell and cause further swelling. The Na+/K+ pump prevents the entry of sodium and so the
sodium becomes the extracellular cation. A Gibbs – Donnan equilibrium is once again set up
and the Na+ is the impermeable charged species.24
Because the Na+/K+ pump is involved in the regulation of cell volume, cell volume is ATP
dependent.
Summary
The structure of the mitochondrion comprises four components: the inner membrane on
which aerobic respiration occurs, the outer membrane, the mitochondrial matrix consisting of
the cristae, and the intermembranous space. The processes of glycolysis, the citric acid cycle
and oxidative phosphorylation to generate ATP take place within the mitochondrion.
The generation of ATP in mitochondria is an extremely complicated chemical process due to

the many intermediates formed in both glycolysis and the citric acid cycle. ATP has a wide
range of functions within the cell. For example ATP is used to drive the Na+/K+ pump, setting
up a concentration gradient which is essential for the regulation of membrane potential and
cell volume. The Na+/K+ pump then utilises the energy obtained from the hydrolysis of ATP
to maintain cell volume and regulate ionic concentrations in the intracellular and extracellular
fluid of the cell.
Conclusion
The synthesis of ATP is a significant function within the cell. Energy obtained from ATP
serves as an integral driving force for many of the cells reactions as well as the maintenance
of its ionic concentrations.
References
1
Young B. et al. Wheater’s Functional Histology: A Text and Colour Atlas. Churchill
Livingstone Elsevier (2006) Ch.3 Pg.21
2
Fox S.I. Human Physiology McGraw – Hill Higher Education (2007) Ch.3 Pg.59
3
Berg J.M. Biochemistry (Stryer) W.H. Freeman and Company (2006) Ch.16 Pg.435
4
J.M. Biochemistry (Stryer) W.H. Freeman and Company (2006) Ch.16 Pg.437
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Fox S.I. Human Physiology McGraw – Hill Higher Education (2007) Ch.6 Pg142
23
24
www.anaesthesaimcq.com

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ESSAY

GIVE AN ACCOUNT OF MITOCHONDRIAL STRUCTURE AND ATP

The Structure of Mitochondria

The Synthesis of ATP

glucose 6 – phosphate is then isomerised to fructose 6 – phosphate.4

This isomeration reaction is proceeded by the phosphorylation of fructose 6 – phosphate by

Stage 3: Glyceraldehyde 3 – phosphate is now converted into 1,3 – bisphosphoglycerate.6

1,3 – bisphosphoglycerate is an energy rich molecule with a greater phosphoryl –

When pyruvate is transported into the mitochondria it is oxidatively decarboxylated by the

Pyruvate+CoA+NAD+ Acetyl CoA+CO2+NADH+H+ 15

Oxidative decarboxylation of α – ketogluatrate forms succinyl coenzymeA, with the release

The generation of ATP in mitochondria is an extremely complicated chemical process due to

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