1. Pyruvate Oxidation and the Citric Acid Cycle
Lecture 30
Pyruvate Oxidation and the Citric Acid Cycle
Title slide

2. Aerobic Fate of Pyruvate: Overview
Aerobic fate of acetylCoA - citric acid cycle

3. Mitochondria: Site of Oxidations
Both the citric acid cycle enzymes and the pyruvate dehydrogenase complex are found in the mitochondrial matrix.

4. Pyruvate Oxidation to Acetyl-CoA
Pyruvate dehydrogenase complexes uses 5 cofactors.

5. Pyruvate Dehydrogenase Complex
Complex enzyme
60 polypeptides of 3 kinds
In mitocondrion matrix
Regulated
Inhibited by NADH and GTP
Stimulated by insulin
Properties

6. Pyruvate Dehydrogenase Complex
Three enzymes and 5 coenzymes
The three components of the complex. I use E1, E2, & E3. This is the eukaryotic complex since there are regulatory proteins.
Total MW: 4.5 x 106!
Also present: regulatory protein kinase, phosphatase

7. Lipoic Acid: Acyl- and Redox Carrier
A swinging arm of some 14 angstroms, undergoes redox, carries acyls. Note that the acyl is on the S from C-8.

8. Thiamine Pyrophosphate (TPP)
TPP functions in the decarboxylation reaction. It loose a proton to yield a carbanion.

9. Pyruvate Dehydrogenase Mechanism
The carbanion attacks the alpha C and forms an adduct.

10. Pyruvate Dehydrogenase Mechanism
The carboxyl group is labilized and lost as carbon dioxide.

11. Pyruvate Dehydrogenase Mechanism
Stabilized by resonance (shift between the two forms). Shifty objects are harder to pin down.

12. Pyruvate Dehydrogenase Mechanism
Hydroxyethyl-TPP is an isolateable intermediate.

13. Pyruvate Dehydrogenase Mechanism
Now they put the acyl group on the wrong S.

14. Pyruvate Dehydrogenase Mechanism
The 8-acetyl lipoamide transfers the acetyl group to coenzyme A giving acetylCoA and producing dihydrolipoic acid.

15. Pyruvate Dehydrogenase Mechanism
E3 dihydrolipoamide dehydrogenase reoxidizes the lipoamide.

16. Pyruvate Dehydrogenase Mechanism
The special redox potential of this bound flavin allows it to transfer hydrogens to NAD+.

17. PDH: The Overall Reaction
Acetyl-CoA enter Krebs cycle
NADH passes e– to O2

18. Krebs Cycle Summary
The Krebs citric acid acid or tricarboxylic acid cycle - three names.

19. Importance of the Krebs Cycle
1. Central energy-yielding path
2. Point of convergence of catabolism of fats, CHO, protein
3. Source of precursors for biosynthesis
What it does.

20. Citrate Synthase: “Condensing Enzyme”
∆Gº= –32.2 kJ/mol
Enzyme # 1 citrate synthase.

21. Aconitase ∆Gº = +13.3 kJ/mol
Enzyme # 2 aconitase makes isocitrate. Functional Fe-S clusters.
∆Gº =
+13.3 kJ/mol

22. Isocitrate Dehydrogenase
Isocitrate dehydrogenase - an oxidative decarboxylation. The first carbon dioxide is lost.
∆Gº= –20.9 kJ/mol

23. -Ketoglutarate Dehydrogenase Complex
a-oxoglutarate dehydrogenase - similar to the pyruvate dehydrogenase. Same cofactors. Another oxidative decarboxylation where the second carbon dioxide is lost.
∆Gº= –33.5 kJ/mol

24. -Ketoglutarate Dehydrogenase Complex
Cofactors: FAD, NAD+, lipoate, TPP, CoASH
Mechanism  PDH complex
This enzyme complex does not involve a regulatory kinase and phosphatase.

25. Succinic Thiokinase ∆Gº= –2.9 kJ/mol
A substrate level phosphorylation.
∆Gº= –2.9 kJ/mol

26. Succinic Thiokinase Multistep reaction:
A phospho-histidine enzymeintermediate is formed.

27. Nucleoside Diphosphate Kinase
∆Gº= 0
Also uses
CTP
UTP
So NTPs are in equilibrium with each other
GTP and ATP are interconvertible.

28. Succinate Dehydrogenase
Complex II of the electron transport chain is succinate dehydrogenase.
∆Gº= 0
Enzyme is membrane bound

29. Fumarase (Fumarate Hydratase)
This is a stereospecific reaction.
∆Gº= –3.8 kJ/mol

30. Malate Dehydrogenase ∆Gº= +29.7 kJ/mol [OAA] normally < 10-6 M
Now to close the circle and come back to the OAA starting compound.
∆Gº= kJ/mol
[OAA] normally < 10-6 M

31. Overall Reaction of Krebs Cycle
The overall reactions.

32. Fate of NADH, FADH2ATP Each NADH3 ATP Each FADH22 ATP
The reduced coenzymes are oxidized via the electron transport chain and coupled to oxidative phosphorylation. The lipoamide is oxidized by E3 which produces NADH. 2.5 & 1.5 ATPs instead of the 3 & 2 shown here.
Each NADH3 ATP
Each FADH22 ATP

33. Energy Yield from Glucose Oxidation
The correct value is about 32.

34. Citric Acid Cycle: Metabolic Hub
A metabolism center.

35. Regulation of the Citric Acid Cycle
Factors that regulate the citric ac id cycle.
Regulation of the Citric Acid Cycle

36. Krebs Cycle in Motion Jon Maber
Dept of Biochemistry and Molecular Biology
The University of Leeds, UK