Glycolysis

Chapter 18
Not section 18.8

Metabolic Pathways comprise of Anabolic and Catabolic Processes

Usually exergonic and release energy

Overview of anabolic pathways

Large molecules are synthesized from smaller ones by adding carbon (usually derived from CO2) and nitrogen (usually as NH3+)

Do not memorize

Overview of catabolic pathways

Amino acids, nucleotides, monosaccharides, and fatty acids are formed by enzymatic hydrolysis of their respective polymers
Do not memorize

 The ATP cycle in cells

Hydrolysis of ATP

(1) ADP and inorganic phosphate (Pi) and (2) AMP and inorganic pyrophosphate (PPi)
ATP + H2O p ADP + Pi (G = 32 kJ/mol

Metabolic Pathways comprise of Anabolic and Catabolic Processes

Usually exergonic and release energy

NADPH Provides the Reducing Power for Anabolic Processes

Figure 17.12 Transfer of reducing equivalents from catabolism to anabolism via the NADPH cycle.

The Substrates of Catabolism Contain Relatively Reduced Forms of Carbon-usually sugars

Figure 17.10 Comparison of the state of reduction of carbon atoms in biomolecules. Chains of CH2- groups are the most practical form of reduced carbon in the biosphere. Carbon dioxide is the final product of catabolism and the most oxidized form of carbon in the biosphere.

NAD+ Collects Electrons Released in Catabolism

Figure 17.11 Hydrogen and electrons released in the course of oxidative catabolism are transferred as hydride ions to the pyridine nucleotide, NAD+, to form NADH + H+ in dehydrogenase reactions.

ATP
ATP is the universal currency of chemical energy ATP is made from ADP and P through a process called chemiosmosis The energy that drives this process is from the proton-motive force Proton concentration gradient and membrane electric potential

12

 The substrates of catabolism are good sources of chemical energy because the carbon atoms are relatively reduced
Oxidative reactions often form hydride ions, H:- (reducing equivalents) The ultimate electron acceptor (oxidizing agent) is O2 which is reduced to water

13

NAD+ Collects Electrons Released in Catabolism

The substrates of catabolism proteins, carbohydrates, and lipids are good sources of chemical energy because their carbon is reduced The oxidative reactions of catabolism release reducing equivalents from these substrates, often in the form of hydride ions These hydrides are transferred to NAD+ molecules, reducing them to NADH NADH in turn passes these reducing equivalents to other acceptors The ultimate oxidizing agent, O2, is the final acceptor of electrons, becoming reduced to H2O

 NADPH provides the reducing power for anabolic processes

In the reduction of substrate, NADPH is oxidized to NADP+

15

Glycolysis
Nearly all living cells carry out a catabolic process known as Glycolysis (stepwise degradation of glucose) Carried out in the cytosol of cells (therefore it is an anaerobic process)

1. How does Glycolysis work?

2. What is the chemical basis and logic for this central pathway of metabolism?

Glycolysis (The Embden-Meyerhof Pathway)

Glycolysis means sugar splitting
Meyerhof

Conversion of ONE molecule of glucose to TWO molecules of pyruvate It is a primitive metabolic pathway as it operates in even the simplest and archaic cells and does not require oxygen Involves 9 intermediate compounds and 10 enzymes; the enzymes are located in the cytoplasm of the cell
The balanced chemical equation for glycolysis is: C6H12O6 + 2ADP + 2NAD+ + 2Pi p 2C3H4O3 + 2ATP + 2NADH + 2H+ + 2H2O

 In eukaryotes, glucose degradation begins in the cytosol and ends in the mitochondria Glucose is completely oxidized to CO2 by O2 The stepwise oxidation of glucose:

19

 Food molecules are broken down in three stages

Digestion in the intestines or lysosome
Reduces large molecules into subunits
Proteins/amino acids Polysaccharides/sugars Fats/fatty acids and glycerol

Small molecules enter the cytosol of the cell

Glycolysis - glucose is converted to pyruvate and then acetyl CoA Citric Acid Cycle - acetyl CoA is completely oxidized to H2O and CO2
20

21

Glycolysis and the Citric Acid Cycle The energy released from the hydrolysis of phosphoanhydride bonds (ATP) powers otherwise energetically unfavorable reactions
Transport of molecules against a concentration gradient Movement of cilia Muscle contraction Synthesis of nucleic acids and proteins
22

 ATP is the universal currency of chemical energy ATP is made from ADP and P through a process called chemiosmosis The energy that drives this process is from the proton-motive force
Proton concentration gradient and membrane electric potential

23

The Mechanism of Glycolysis

We will divide glycolysis into two phases: 1) The preparatory phase the first five steps of glycolysis: 2 molecules of ATP are consumed to activate the glucose molecule for its cleavage into two three-carbon pieces ATP raises the free-energy content of the intermediates and the carbon chains of all the metabolized hexoses which are converted into a common product: glyceraldehyde 3phosphate 2) The payoff phase the last five steps of glycolysis involve oxidative conversion of two molecules of glyceraldehyde to form pyruvate

Glycolysis - glucose is converted to two threecarbon molecules of pyruvate

The chemical reactions take place in the cytosol No oxygen is required Highly regulated - just enough glucose to meet the cell s need for ATP is transported into the cell All of the metabolic intermediates are phosphorylated compounds Four molecules of ATP are made and two molecules are consumed for a net production of 2 ATP

25

The Energetics of Glycolysis

We can resolve the equation for glycolysis into two processes: 1) The conversion of glucose to pyruvate, which is exergonic (release of energy) Glucose + 2NAD+ p 2 pyruvate + 2NADH + 2H+ (G'r = 146 kJ/mol

2) The formation of ATP from ADP and Pi, which is endergonic (absorbing energy) 2ADP + 2 Pi p 2ATP + 2H2O (G'r = 2(30.5 kJ/mol) = 61 kJ/mol

The sum of the two processes gives the overall standard free-energy change of glycolysis:
(G'r = 146 kJ/mol + 61 kJ/mol = 85 kJ/mol Glycolysis releases only a small fraction of the total available energy of the glucose molecule. When glucose is oxidized completely to CO2 and H2O, the total standard free-energy change is 2,840 kJ/mol Glycolytic degradation of glucose to two molecules of pyruvate yields only ~5.2% of the total energy that can be released from glucose by complete oxidation. The remainder of the energy will be extracted by the citric acid cycle and oxidative phosphorylation

Overview of Glycolysis:
glycolysis can be divided into two phases: the preparatory or energy investment phase and the energy payoff phase.

 The first phase of glycolysis: Glucose converted to Pyruvate

The phosphorylation of glucose - priming reaction Requires energy from ATP (30.5 kJ/mol) Phosphorylation of glucose costs 13.8 kJ/mol Liberates 16.7 kJ/mol First site of regulation

29

The Importance of Glucose Phosphorylation

1) Phosphorylation keeps glucose in the cell. Glucose is a neutral molecule and could diffuse across the cell membrane (at a low rate), but phosphorylation confers a negative charge on glucose, and the plasma membrane is impermeable to glucose-6-phosphate. 2) Rapid conversion of glucose to G-6-P keeps the intracellular concentration of glucose low, favoring movement of glucose into the cell. 3) Regulatory control can be imposed only on reactions not at equilibrium, the favorable thermodynamics of this first glycolytic reaction makes it an important site for regulation.

Substrate-Induced Conformational Change in Hexokinase

Glucose molecule in purple

The two lobes that form the active site cleft swing together in hexokinase (8 angstroms) to capture the glucose molecule. This movement places the ATP in close proximity to the C6H2OH group of glucose and excludes water from the active site. Water must be excluded otherwise the thermodynamically favored transfer of the phosphoryl group to water would dominate.

Isozymes of Hexokinase
The conversion of glucose to glucose-6-phosphate is also performed by the enzyme glucokinase. Hexokinase and glucokinase are isozymes (enzymes that catalyze the same reaction) but are encoded by different genes. Glucokinase is found in the liver but unlike hexokinase, it is highly specific for D-glucose, higher KM for glucose (~ 10 mM), and is not product-inhibited. With such a high KM for glucose, glucokinase becomes important metabolically only when liver glucose levels are high (for example, when an individual has consumed large amounts of sugar). When glucose levels are low, hexokinase is primarily responsible for phosphorylating glucose. Glucokinase is an inducible enzyme; the amount present in the liver is controlled by insulin.

Reaction 2: Phosphoglucosimerase catalyzes the isomerization of Glucose-6-Phosphate

Isomerization of glucose-6-phosphate

33

Reaction 3: Phosphofructokinase phosphorylates

Second priming reaction Requires energy from ATP (30.5 kJ/mol) Phosphorylation of fructose-6-phosphate costs 16.3 kJ/mol Liberates 14.2 kJ/mol

Site of regulation ATP is an allosteric inhibitor Citrate is an allosteric inhibitor - coupled to citric acid cycle

34

Fig. 18-1, p. 536

 The second phase of glycolysis

Substrate-level phosphorylation

Step 6

Pyruvate kinase

ADP has been phosphorylated to form ATP at the expense of a substrate

36

Fig. 18-1, p. 536

Last step
Pyruvate kinase is activated by AMP and fructose1,6-bisphosphate and inhibited by ATP, acetyl-CoA, and alanine Third site of regulation (large negative (G)

38

 Glycolysis generates 2H+ and 4e- that are transferred to two molecules of the oxidized form of the electron carrier nicotinamide adenine dinucleotide, NAD+

39

 Other sugars can enter the pathway

40

 Thermodynamics of Glycolysis Couple an energetically unfavorable reaction to an energetically favorable one

41

42

Coupling of the reactions in steps 6 and 7 of glycolysis allows the energetically unfavorable formation of a high energy phosphate bond

43

The Glycolysis Balance Sheet

Control Points in Glycolysis

Read about the various isozymes of phosphofructokinase, and about how this enzyme is regulated

The End of Glycolysis: Possible Fates of Pyruvate

 Some eukaryotic and prokaryotic cells metabolize (break down) glucose anaerobically Glucose is not converted to CO2 but to one or more twoor three-carbon molecules and sometimes CO2 Much less ATP is produced per molecule of glucose The products are lactic acid and ethanol Anaerobic fermentation - beer and wine industry

47

 Some eukaryotic and prokaryotic cells metabolize (break down) glucose anaerobically Glucose is not converted to CO2 but to one or more twoor three-carbon molecules and sometimes CO2 Much less ATP is produced per molecule of glucose The products are lactic acid and ethanol Anaerobic fermentation - beer and wine industry

48

The 4 Major Fates of Pyruvate

The fates of pyruvate produced during glycolysis depends on the cell type or conditions
1) In aerobic organisms or tissues, under aerobic conditions, glycolysis is only the first stage in the complete degradation of glucose; the pyruvate derivative, acetyl CoA is oxidized completely to CO2 by the citric acid (TCA) cycle. 2) In vigorously contracting skeletal muscle, pyruvate can be reduced to lactate via lactic acid fermentation; this occurs under low oxygen conditions (hypoxia); pyruvate accepts electrons from NADH and thereby regenerates the NAD+ necessary for glycolysis to continue. 3) In a process termed alcohol fermentation, microorganisms can convert pyruvate to ethanol and CO2.

51

52

Citric Acid Cycle

Chapter 19 (NOT Sections 19.8 and 19.10)

 Following glycolysis
pyruvate is transported to the mitochondria where it is oxidized by O2 to CO2 Sugars and fats are both degraded to acetyl CoA These oxidation reactions generate the bulk of the ATP produced

54

Fatty acids are also oxidized to acetyl CoA

55

 Citric Acid Cycle (Tricarboxylic acid cycle or Krebs cycle) Oxidizes acetyl group to CO2 Reduces NAD and FAD to NADH and FADH2

56

The Citric Acid Cycle (aka, The Krebs Cycle or The Tricarboxylic Acid (TCA) Cycle)
Krebs

The citric acid cycle is a series of biochemical reactions that take place in the mitochondrial matrix and release the chemical energy stored in acetyl CoA (or Coenyzme A). Acetyl CoA is a product synthesized from pyruvate produced as the end product of glycolysis. The two-carbon acetyl CoA enters the cycle to combine with oxaloacetate to from citric acid. Two carbons eventually emerge from the cycle as CO2. The energy released by the oxidations reactions of the cycle is conserved in the reduction of 3 NAD+ and one FAD and the production of one ATP (or GTP).

The overall consumption of one molecule of acetyl CoA in the citric acid cycle is exergonic: (G'r = 60 kJ/mol.

Essential Questions
Recall The glycolytic pathway converts glucose to pyruvate producing 2 molecules of ATP (net). This represents only 5.2% of the potential energy available from glucose Anaerobic conditions Pyruvate is reduced to lactate in animals and ethanol in yeast Aerobic conditions CITRIC ACID CYCLE How is pyruvate oxidized under aerobic conditions? What is the chemical logic that dictates this process?

Pyruvate to Acetyl-CoA
The Step Connecting Glycolysis to the TCA Cycle

A large enzyme complex called the Pyruvate Dehydrogenase Complex catalyzes the conversion of pyruvate to acetyl CoA in the matrix of the mitochondrion. This reaction involves three steps: 1) pyruvate s carboxyl group is removed and given off as CO2; 2) acetate is formed from the remaining two-carbon fragment via oxidation in which two electrons are passed to NAD+ to form NADH; 3) coenzyme A, a derivative of vitamin B, is attached to acetate by an unstable bond making it a reactive compound.

Translocation of Pyruvate from the Cytosol to the Mitochondrial Matrix

Pyruvate diffuses through the outer mitochondrial membrane via aqueous channels. Pyruvate translocase then transports pyruvate and H+ into the mitochondrial matrix, where pyruvate is converted to acetyl-CoA by the pyruvate dehydrogenase enzyme complex.

Aerobic pathways permit production of 30-38 molecules of ATP per glucose oxidized -2 ATP molecules from glycolysis and 2 from Citric acid cycle, most from oxidative phosphorylation

62

Chapter 18 Glycolysis Questions 1,3,4,10,16

(NOT Sections 18.8)