The Citric Acid Cycle

1. The Citric Acid Cycle Chapter 16 (Page 601-614)

2. Glycolysis Review

3. 1. Summary of Glycolysis Glycolysis is the catabolic processing of glucose to extract energy. Glucose + 2 NAD+ + 2 ADP + 2 Pi 2 Pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O Payoff Phase Preparatory Phase 2 2

4. Glucose Carbons in GAP D- Glucose 1P : 3C

5. Glucose Carbons in GAP D- Glucose 1P : 3C

6. 1. Summary of Glycolysis Glycolysis is the catabolic processing of glucose to extract energy. Glucose + 2 NAD+ + 2 ADP + 2 Pi 2 Pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O Carbon Oxidation States: Payoff Phase Preparatory Phase +3 +1 0 +1 +2 2 2 0 0 -3 0 -1 0 -1 Net Carbon Oxidation States: 2 x +2 = +4 0 0

7. 1. Summary of Glycolysis Glycolysis is the catabolic processing of glucose to extract energy. Glucose + 2 NAD+ + 2 ADP + 2 Pi 2 Pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O Where do the four e¯ go? NAD+ + H+ + 2e- →NADH Energy is conserved in the formation of NADH.

8. 1. Summary of Glycolysis Glycolysis is the catabolic processing of glucose to extract energy. Glucose + 2 NAD+ + 2 ADP + 2 Pi 2 Pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O Standard free energy of the whole process: Glucose + 2 NAD+ 2 Pyruvate + 2 NADH + 2 H+ G1′ = –146 kJ/mol 2 ADP + 2 Pi 2 ATP + 2 H2O G2′ = +61.0 kJ/mol Glucose + 2 NAD+ + 2 ADP + 2 Pi 2 Pyruvate + 2 NADH + 2 H+ + 2 ATP + 2 H2O Gs′ = -85 kJ/mol Energy is conserved in the formation of ATP.

9. 2. Fates of Pyruvate

10. Glucose and Cellular Respiration

11. 1. Only a small amount of energy available in glucose is captured in glycolysis Glycolysis 2 G′ = –146 kJ/mol GLUCOSE Full oxidation (+ 6 O2) G′ = –2,840 kJ/mol 6 CO2 + 6 H2O The full oxidation of glucose is part of the process of cellular respiration.

12. 2. Cellular Respiration • Process in which cells consume O2 and produce CO2 • Provides more energy (ATP) from glucose than glycolysis • Also captures energy stored in lipids and amino acids • Evolutionary origin: developed about 2.5 billion years ago • Used by animals, plants, and many microorganisms • Occurs in three major stages: • acetyl CoA production • acetyl CoA oxidation • electron transfer and oxidative phosphorylation

13. 3A. Respiration: Stage 1 Acetyl-CoA Production Generates some: ATP, NADH, FADH2

14. 3B. Respiration: Stage 2 Acetyl-CoA oxidation Generates more NADH, FADH2,and one GTP

15. 3C. Respiration: Stage 3 Oxidative Phosphorylation Generates a lot of ATP

16. 4. Localization of events involved in glucose catabolism and cellular respiration • Cellular respiration involving glucose: • A. Glycolysis occurs in the cytoplasm • B. Citric acid cycle occurs in the mitochondrial matrix • Except the action of succinate dehydrogenase, which is located in the inner membrane • Mostly dealing with soluble proteins • C. Oxidative phosphorylation occurs in the inner membrane

17. 4A. Structure of a Mitochondrion Double membrane leads to four distinct compartments: • Outer Membrane: • Relatively porous membrane allows passage of metabolites • Intermembrane Space (IMS): • similar environment to cytosol • higher proton concentration (lower pH) • Inner Membrane • Relatively impermeable, with proton gradient across it • Location of electron transport chain complexes • Convolutions called Cristae serve to increase the surface area • Matrix • Location of the citric acid cycle and parts of lipid and amino acid metabolism • Lower proton concentration (higher pH)

18. 4A. Structure of a Mitochondrion Courtesy of Mariana Ruiz Villareal.

19. The Citric Acid Cycle

20. 1. Conversion of Pyruvate to Acetyl-CoA • Net Reaction: • Oxidative decarboxylation of pyruvate • First carbons of glucose to be fully oxidized

21. 1. Conversion of Pyruvate to Acetyl-CoA • Catalyzed by the pyruvate dehydrogenase complex • Requires 5 coenzymes • NAD+ and CoA-SH are co-substrates • TPP, lipoyllysine, and FAD are prosthetic groups

22. 1AI. Structure of Coenzyme A • Coenzymes are not a permanent part of the enzymes’ structure. • They associate, fulfill a function, and dissociate • The function of CoA is to accept and carry acetyl groups

23. 1AII. Structure of Lipoyllysine • Prosthetic groups are strongly bound to the protein • The lipoic acid is covalently linked to the enzyme via a lysine residue • An acetyl transporter

24. 1AIII. Structure of TPP • Thiamine pyrophosphate (TPP)

25. 1AIV. Structure of FAD • Flavin adenine dinucleotide (FAD) • - An electron carrier.

26. 1B. Pyruvate Dehydrogenase Complex (PDC) • PDC is a large (up to 10 MDa) multienzyme complex • pyruvate dehydrogenase (E1) • dihydrolipoyltransacetylase (E2) • dihydrolipoyl dehydrogenase (E3) • Advantages of multienzyme complexes: • short distance between catalytic sites allows channeling of substrates from one catalytic site to another • channeling minimizes side reactions • regulation of activity of one subunit affects the entire complex

27. 3D Reconstruction from Cryo-EM data • A cryoelectron micrograph inspired model of PDC shows a core (green) consisting of 60 molecules of E2 arranged in 20 trimers forming a pentagonal dodecahedron • (50 nm in diameter). • pyruvate dehydrogenase (E1) • dihydrolipoyltransacetylase (E2) • dihydrolipoyl dehydrogenase (E3)

28. 1C. Overall Reaction of PDC • Enzyme 1 • Step 1: Decarboxylation of pyruvate • Step 2: Oxidation of acetyl-TPP • -Electrons reduce lipoamide and form a thioester

29. 1C. Overall Reaction of PDC +1 • Enzyme 1 • Step 1: Decarboxylation of pyruvate • Step 2: Oxidation of acetyl-TPP • -Electrons reduce lipoamide and form a thioester

30. 1C. Overall Reaction of PDC +3 • Enzyme 1 • Step 1: Decarboxylation of pyruvate • Step 2: Oxidation of acetyl-TPP • -Electrons reduce lipoamide and form a thioester

31. 1C. Overall Reaction of PDC +3 +3 • Enzyme 2 • Step 3: Formation of acetyl-CoA (product 1)

32. 1C. Overall Reaction of PDC +3 +3 • Enzyme 3 • Step 4:Reoxidation of the lipoamide cofactor • Step 5: Regeneration of the oxidized FAD cofactor • Forming NADH (product 2)