1. Chapter 9
Chem 341
Suroviec Fall 2016

2. I. Citric Cycle Overview
8 reactions
Oxidizes acetyl group of Acetyl CoA to 2 CO2

3. I. General Features
Circular Pathway oxidizes acetyl groups from many sources
Net reaction
In eukaryotes all enzymes of CAC are located in mitochondria
1 CO2 produced in 1 round of the cycle
Oxidation of acetyl groups to 2 CO2 requires transfer of 4 pair of electrons

4. II. Synthesis of Acetyl CoA
Pyruvate Dehydrogenase: Multienzyme Complex
Group of non covalently associated enzymes that catalyze 2+ sequential steps in metabolic pathway
Formed from pyruvate through oxidative decarboxylation
Pyruvate dehydrogenase (E1)
Dihydrolipoyl transacetylase (E2)
Dihydrolipoyl dehydrogenase (E3)
24 E2 proteins associated as trimers at the corners of cube
24 E1 proteins form dimers that associate with E2 core along the 12 edges.
The 12 E3 proteins form dimers that attach to the 6 faces of E2 cube
c) Combining a) and b) forms a 60 subunit complex

5. Section 9.2: Citric Acid Cycle
Decarboxylation
Action of lipoic acid
Figure 9.10 Reactions Catalyzed by the Pyruvate Dehydrogenase Complex
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

6. B. Pyruvate Dehydrogenase
Overall Reaction
Pyruvate dehydrogenase (E1)
A TPP requiring enzyme
TPP acts as electron sink in the reaction

7. 2. Lipoamide
Hydroxyethyl group transferred to E2
Lipoamide uses lysine
Cyclic disulfide reversibly reduced

8. 3. E2 transesterification
Yields acetyl CoA and dihydrolipoamide-E2
4. Regenerate E2
E3 becomes reduced
Regenerates E2
Disulfide interchange reaction

9. 5. Reoxidize E3
Reoxidize E3

10. Section 9.2: Citric Acid Cycle
Figure 9.15 Citrate Metabolism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

11. Section 9.2: Citric Acid Cycle
Figure 9.15 Citrate Metabolism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

12. III. Enzymes of CAC
Citrate synthase
Aconitase
Isocitrate dehydrogenase
a-ketoglutarate dehydrogenase:
Succinyl CoA synthetase
Succinate dehydrogenase
Fumerase
Malate dehydrogenase

13. III. Enzymes of the CAC
Citrate Synthase
Catalyzes the condensation of acetyl-CoA and oxaloacetate
Free enzyme is a dimer
Active site closes when oxaloacetate binds
Conformational changes seals oxaloacetate in binding site and shuts out the solvent

14. C. NAD+ Dependant Isocitrate Dehydrogenase
B. Aconitase
Catalyzes reversible isomerization of citrate to isocitrate
C. NAD+ Dependant Isocitrate Dehydrogenase
Catalyzes oxidative decarboxylation of isocitrate to a-ketogluterate

15. D. a-ketoglutarate dehydrogenase
Catalyzes oxidative decarboxylation of a-ketogluterate
Is a multienzyme complex
E1: a-ketoglutarate dehydrogenase
E2: dihydrolipoyl transsuccinylase
E3: dihydrolipoyl dehydrogenase

16. E. Succinyl-CoA Synthetase
Cleaves “high-energy” succinyl-CoA to synthesis of GTP
Reaction almost energy neutral.

17. F. Succinate Dehydrogenase
Catalyzes stereospecific dehydrogenation of succinate to fumerate
Inhibited by malonate

18. H. Malate dehydrogenase
G. Fumerase
Catalyzes the hydration of double bond of fumarate to form malate
H. Malate dehydrogenase
Regeneration of oxaloacetate

19. Availibity of substrates Need for CAC intermediates Demand for ATP
IV. Regulation of CAC
Availibity of substrates
Need for CAC intermediates
Demand for ATP

20. A. Regulation of pyruvate decarboxylation
Product inhibition by NADH and acetyl-CoA
NADH, acetyl-CoA compete with NAD+ and CoA for binding sites
Drive with E2 and E3
Covalent modification by phosphorylation/dephosphorylation of E1

21. B. Rate-Controlling Enzymes
Flux of metabolites through the CAC is proportional to the rate of cellular oxygen consumption
3 main mechanisms
Substrate availability
Product inhibition
Competitive feedback inhibition

22. B. Rate-Controlling Enzymes
Regulators are acetyl-CoA, oxaloacetate, NADH
Flux varies with substrate concentration

23. V. Pathways that use CAC intermediates
Glucose biosynthesis
Fatty acid synthesis
Amino Acid synthesis