Part 4

Module 9: The Citric Acid Cycle

Overview

• Date: Friday, October 24th

• Reading assignment: Chapter 19

• Key Points:

  • 2 carbons in acetyl-CoA (from pyruvate dehydrogenase complex, PDHC) fully oxidized to CO₂.

  • Starting from pyruvate (produced in glycolysis), all three carbon atoms are now oxidized to CO₂, leading to significant energy release.

  • Most chemical energy is temporarily stored in NADH.

  • One GTP is produced from GDP plus inorganic phosphate.

  • The cycle consists of 8 reactions organized in a cyclic pathway.

• Adaptive Quiz: Chapter 19 due Monday, October 27th, 5:00 pm

Citric Acid Cycle Equation

• $C<em>6H</em>{12}O<em>6 + 6O</em>2 <br>ightarrow 6H<em>2O + 6CO</em>2$

• Terminal electron acceptor: O₂, which is required for full oxidation of carbon.

• Result: Fully oxidized carbon results in potential energy release.

Pyruvate Oxidation Overview

• Combines the actions of the pyruvate dehydrogenase complex and the citric acid cycle to yield 3 CO₂.

Stages of Pyruvate Oxidation

1. Stage 1: Pyruvate to Acetyl-CoA

   • Process: Pyruvate + Coenzyme A-SH → Acetyl-CoA

   • Outputs: 1 CO₂ released, 1 NADH produced, uses 5 coenzymes.

   • Enzyme: Pyruvate dehydrogenase complex (PDHC).

2. Stage 2: Citric Acid Cycle

   • Process: Acetyl-CoA → 2 CO₂ + Coenzyme A-SH

   • Outputs: 3 NADH, 1 FADH₂ produced, 1 ATP (or GTP).

   • The cycle is catalyzed by 8 different enzymes, regenerating oxaloacetate.

   • Starts with oxaloacetate, ends with oxaloacetate.

Steps of the Citric Acid Cycle

1. Step 1: Acetyl-CoA condenses with oxaloacetate (OAA).

   • Enzyme: Citrate synthase.

   • Mechanism: Hydrolysis of CoA thioester provides energy.

   • Type of reaction: Lyase - involves addition across a double bond to form a C-C bond without ATP.

   • Active site for hydrolysis of thioester does not form until citryl-CoA has formed.

2. Step 2: Conversion of citrate to isocitrate.

   • Enzyme: Aconitase.

   • Involves two lyase reactions: dehydration followed by hydration.

   • Reaction type resembles isomerase but is completed in two steps by lyases.

3. Step 3: Isocitrate to α-ketoglutarate.

   • Enzyme: Isocitrate dehydrogenase.

   • Key reaction: Hydroxyl group oxidized to ketone, generating β-keto acid, followed by decarboxylation.

   • Results: Production of α-ketoglutarate and CO₂; classified as oxidation-reduction and then decarboxylation.

   • Note: Incorrectly labeled as α-ketoacid in textbook page 332.

4. Step 4: α-ketoglutarate to succinyl-CoA.

   • Enzyme: α-ketoglutarate dehydrogenase complex.

   • Functional similarity to PDHC, differing only by acyl group (succinate vs acetyl).

5. Step 5: Succinyl-CoA to succinate.

   • Enzyme: Succinyl-CoA synthase.

   • Mechanism: Conversion of thioester to carboxylic acid; free energy released used for ATP synthesis (substrate-level phosphorylation).

   • The mechanism resembles another important glycolysis reaction (glyceraldehyde-3-P dehydrogenase).

6. Steps 6-8: Regeneration of oxaloacetate.

   • Step 6: Succinate to fumarate, carried out by succinate dehydrogenase (a redox reaction).

   • FAD reduced to FADH₂; enzyme-bound, does not leave the enzyme.

   • Step 7: Fumarate to malate, catalyzed by fumarase (lyase reaction, hydration).

   • Step 8: Malate to oxaloacetate, catalyzed by malate dehydrogenase (redox reaction), reducing NAD⁺ to NADH.

Completed Cycle Reaction

• Total reaction for the citric acid cycle:

  • $Acetyl-CoA + 3 NAD^+ + FAD + ADP + P<em>i + 2 H</em>2O <br>ightarrow 2 CO<em>2 + 3 NADH + 3 H^+ + FADH</em>2 + ATP + CoA-SH$

• The coenzyme that entered at PDH complex is regenerated in the cycle.

Energy Accounting and Storage

• Summary of energy production from the cycles and pathways.

  • GTP: 1

  • NADH: 3

  • FADH₂: 1

• Total energy yield from the pyruvate dehydrogenase complex and the citric acid cycle:

  • 1 ATP + 4 NADH + 1 FADH₂.

• Reaction Classes in the Citric Acid Cycle:

1. Acetyl-CoA + oxaloacetate + H₂O → citrate (by citrate synthase).

2. Citrate → cis-aconitate → isocitrate (by aconitase).

3. Isocitrate + NAD⁺ → α-ketoglutarate + CO₂ + NADH (by isocitrate dehydrogenase).

4. α-Ketoglutarate + NAD⁺ + CoA → succinyl-CoA + CO₂ + NADH (by α-ketoglutarate dehydrogenase).

5. Succinyl-CoA + GDP + P_i → succinate + GTP + CoA (by succinyl-CoA synthetase).

6. Succinate + FAD → fumarate + FADH₂ (by succinate dehydrogenase).

7. Fumarate + H₂O → malate (by fumarase).

8. L-Malate + NAD⁺ → oxaloacetate + NADH (by malate dehydrogenase).

Multifunctionality of the Citric Acid Cycle

• The citric acid cycle serves not just to capture energy in NADH and FADH₂ but also:

  • Interconverts carbon atoms among various biomolecules, supporting gluconeogenesis.

  • Converts α-ketoglutarate to glutamic acid, thus playing roles in both catabolism and anabolism.

  • These pathways are termed amphibolic due to their dual roles.

Anaplerotic Reactions

• Continuous replenishment of citric acid cycle intermediates is crucial.

  • Intermediates drawn off must be restored, referred to as anaplerotic reactions.

  • Example: Pyruvate + CO₂ → OAA, catalyzed by pyruvate carboxylase, which does not yield CO₂ during the process and requires ATP (also serves as a step in gluconeogenesis).

The Glyoxylate Cycle

• This alternative pathway converts two acetyl-CoA to one oxaloacetate (OAA), allowing for gluconeogenesis and amino acid synthesis.

  • The citric acid cycle cannot convert acetyl-CoA into oxaloacetate (releases 2 CO₂).

  • The glyoxylate cycle bypasses the decarboxylation steps and uses alternative enzymes.

Summary of the Glyoxylate Cycle

• Inputs: Two acetyl-CoA and one oxaloacetate (OAA).

• Outputs: Two oxaloacetate (OAA).

• The cycle is present in plants and some microbes, unlike mammals who cannot utilize a lipid-only diet.

• Important for plants during germination of oil seeds.

Conclusion

• The citric acid cycle and its associated pathways play crucial roles in cellular respiration, energy production, and metabolic flexibility, supporting both energy generation and the biosynthesis of key biomolecules.