1. Oxidative decarboxylation of pyruvate and Krebs cycle

2. OXIDATIVE DECARBOXYLATION OF PYRUVATE
Matrix of the mitochondria contains pyruvate dehydrogenase complex

3. The fate of glucose molecule in the cell
Synthesis of glycogen
Glucose
Pentose phosphate pathway
Glycogen
Glucose-6-phosphate
Ribose, NADPH
Degradation of glycogen
Gluconeogenesis
Glycolysis
Ethanol
Pyruvate
Lactate
Acetyl Co A

4. OXIDATIVE DECARBOXYLATION OF PYRUVATE
Only about 7 % of the total potential energy present in glucose is released in glycolysis.
Glycolysis is preliminary phase, preparing glucose for entry into aerobic metabolism.
Pyruvate formed in the aerobic conditions undergoes conversion to acetyl CoA by pyruvate dehydrogenase complex.

5. OXIDATIVE DECARBOXYLATION OF PYRUVATE
Pyruvate dehydrogenase complex is a bridge between glycolysis and aerobic metabolism – citric acid cycle
Pyruvate dehydrogenase complex and enzymes of cytric acid cycle are located in the matrix of mitochondria 5

6. Entry of Pyruvate into the Mitochondrion
Pyruvate freely diffuses through the outer membrane of mitochon-dria through the channels formed by transmembrane proteins porins.
Pyruvate translocase, protein embedded into the inner membrane, transports pyruvate from the intermembrane space into the matrix in symport with H+ and exchange (antiport) for OH-.

7. Conversion of Pyruvate to Acetyl CoA
Pyruvate dehydrogenase complex (PDH complex) is a multienzyme complex containing 3 enzymes, 5 coenzymes and other proteins.
Pyruvate dehydrogenase complex is giant, with molecular mass ranging from 4 to 10 million daltons.
Electron micrograph of the pyruvate dehydrogenase complex from E. coli.

8. Pyruvate dehydrogenase complex
Enzymes:
E1 = pyruvate dehydrogenase
E2 = dihydrolipoyl acetyltransferase
E3 = dihydrolipoyl dehydrogenase

9. Pyruvate dehydrogenase complex
Coenzymes: TPP (thiamine pyrophosphate), lipoamide, HS-CoA, FAD+, NAD+.
TPP is a prosthetic group of E1;
lipoamide is a prosthetic group of E2; and
FAD is a prosthetic group of E3.
The building block of TPP is vitamin B1 (thiamin); NAD – vitamin B5 (nicotinamide); FAD – vitamin B2 (riboflavin), HS-CoA – vitamin B3 (pantothenic acid), lipoamide – lipoic acid 9

10. Overall reaction of pyruvate dehydrogenase complex
Pyruvate dehydrogenase complex is a classic example of multienzyme complex
Overall reaction of pyruvate dehydrogenase complex
The oxidative decarboxylation of pyruvate catalized by pyruvate dehydrogenase complex occurs in five steps.

11. The Citric
Acid Cycle
Aerobic cells use a metabolic wheel – the citric acid cycle – to generate energy by acetyl CoA oxidation

12. Glucose Fatty Acids Amino Acids
Synthesis of glycogen
Glucose
Pentose phosphate pathway
Glycogen
Glucose-6-phosphate
Ribose, NADPH
Degradation of glycogen
Gluconeogenesis
Glycolysis
Ethanol
Pyruvate
Lactate
Fatty Acids
Amino Acids
Acetyl Co A
The citric acid cycle is the final common pathway for the oxidation of fuel molecules — amino acids, fatty acids, and carbohydrates.
Most fuel molecules enter the cycle as acetyl coenzyme A.

13. Names: The Citric Acid Cycle Tricarboxylic Acid Cycle Krebs Cycle
Hans Adolf Krebs. Biochemist; born in Germany. Worked in Britain. His discovery in 1937 of the ‘Krebs cycle’ of chemical reactions was critical to the understanding of cell metabolism and earned him the 1953 Nobel Prize for Physiology or Medicine.
In eukaryotes the reactions of the citric acid cycle take place inside mitochondria

14. An Overview of the Citric Acid Cycle
A four-carbon oxaloacetate condenses with a two-carbon acetyl unit to yield a six-carbon citrate.
An isomer of citrate is oxidatively decarboxylated and five-carbon -ketoglutarate is formed.
-ketoglutarate is oxidatively decarboxylated to yield a four-carbon succinate.
Oxaloacetate is then regenerated from succinate.

15. An Overview of the Citric Acid Cycle
Two carbon atoms (acetyl CoA) enter the cycle and two carbon atoms leave the cycle in the form of two molecules of carbon dioxide.
Three hydride ions (six electrons) are transferred to three molecules of NAD+, one pair of hydrogen atoms (two electrons) is transferred to one molecule of FAD.
The function of the citric acid cycle is the harvesting of high-energy electrons from acetyl CoA. 15

16. 1. Citrate Synthase Citrate formed from acetyl CoA and oxaloacetate
Only cycle reaction with C-C bond formation
Addition of C2 unit (acetyl) to the keto double bond of C4 acid, oxaloacetate, to produce C6 compound, citrate
citrate synthase

17. 2. Aconitase
Elimination of H2O from citrate to form C=C bond of cis-aconitate
Stereospecific addition of H2O to cis-aconitate to form isocitrate
aconitase

18. 3. Isocitrate Dehydrogenase
Oxidative decarboxylation of isocitrate to a-ketoglutarate (a metabolically irreversible reaction)
One of four oxidation-reduction reactions of the cycle
Hydride ion from the C-2 of isocitrate is transferred to NAD+ to form NADH
Oxalosuccinate is decarboxylated to a-ketoglutarate
isocitrate dehydrogenase

19. 4. The -Ketoglutarate Dehydrogenase Complex
Similar to pyruvate dehydrogenase complex
Same coenzymes, identical mechanisms
E1 - a-ketoglutarate dehydrogenase (with TPP) E2 – dihydrolipoyl succinyltransferase (with flexible lipoamide prosthetic group) E3 - dihydrolipoyl dehydrogenase (with FAD)
a-ketoglutarate dehydrogenase

20. 5. Succinyl-CoA Synthetase
Free energy in thioester bond of succinyl CoA is conserved as GTP or ATP in higher animals (or ATP in plants, some bacteria)
Substrate level phosphorylation reaction
HS- +
GTP + ADP GDP + ATP
Succinyl-CoA Synthetase

21. 6. The Succinate Dehydrogenase Complex
Complex of several polypeptides, an FAD prosthetic group and iron-sulfur clusters
Embedded in the inner mitochondrial membrane
Electrons are transferred from succinate to FAD and then to ubiquinone (Q) in electron transport chain
Dehydrogenation is stereospecific; only the trans isomer is formed
Succinate Dehydrogenase

22. 7. Fumarase
Stereospecific trans addition of water to the double bond of fumarate to form L-malate
Only the L isomer of malate is formed
Fumarase

23. 8. Malate Dehydrogenase Malate is oxidized to form oxaloacetate.

24. Stoichiometry of the Citric Acid Cycle
Two carbon atoms enter the cycle in the form of acetyl CoA.
Two carbon atoms leave the cycle in the form of CO2 .
Four pairs of hydrogen atoms leave the cycle in four oxidation reactions (three molecules of NAD+ one molecule of FAD are reduced).
One molecule of GTP, is formed.
Two molecules of water are consumed.

25. Stoichiometry of the Citric Acid Cycle
9 ATP (2.5 ATP per NADH, and 1.5 ATP per FADH2) are produced during oxidative phosphorylation
1 ATP is directly formed in the citric acid cycle
1 acetyl CoA generates approximately 10 molecules of ATP 25

26. Functions of the Citric Acid Cycle
Integration of metabolism. The citric acid cycle is amphibolic (both catabolic and anabolic).
The cycle is involved in the aerobic catabolism of carbohydrates, lipids and amino acids.
Intermediates of the cycle are starting points for many anabolic reactions.
Yields energy in the form of GTP (ATP).
Yields reducing power in the form of NADH2 and FADH2.

27. Regulation of the Citric Acid Cycle
Pathway controlled by:
(1) Allosteric modulators
(2) Covalent modification of cycle enzymes
(3) Supply of acetyl CoA (pyruvate dehydrogenase complex)

28. Regulation of the Citric Acid Cycle
Three enzymes have regulatory properties
citrate synthase (is allosterically inhibited by NADH, ATP, succinyl CoA, citrate – feedback inhibition)
isocitrate dehydrogenase (allosteric effectors: (+) ADP; (-) NADH, ATP. Bacterial ICDH can be covalently modified by kinase/phosphatase)
-ketoglutarate dehydrogenase complex (inhibition by ATP, succinyl CoA and NADH 28

29. Regulation of the citric acid cycle
NADH, ATP, succinyl CoA, citrate -

30. Krebs Cycle is a Source of Biosynthetic Precursors
Phosphoenol- pyruvate
Glucose
The citric acid cycle provides intermediates for biosyntheses

31. Role of the citric cycle in anabolism