Biochemistry Lecture: Pyruvate Decarboxylation and Citric Acid Cycle

Overview

• Focus: Linking glycolysis with the citric acid cycle via pyruvate decarboxylation.
• Context: Pyruvate formed from glycolysis moves into the mitochondrial matrix under aerobic conditions.
• Objective: Discussing Step One of the Citric Acid Cycle in detail.

Pyruvate Decarboxylation

• Location: Mitochondrial matrix.
• Process: Activation of pyruvate by removing CO₂ to form an acetyl group.
• Carrier Molecule: Co-enzyme A (CoA).
• Product: Acetyl-CoA complex, which enters the Citric Acid Cycle.
• Significance: Links glycolysis to aerobic cellular respiration.

First Step of the Citric Acid Cycle

• Reactants: Acetyl-CoA (2 carbon) and Oxaloacetate (4 carbon).
• Product: Citrate (6 carbon), conjugate base of citric acid (tricarboxylic acid, TCA).
• Catalyst: Citrate Synthase enzyme.

Citrate Synthase and Multi-Step Process

• Enzyme Type: Dimer with two identical subunits.
• Domains: Three domains per subunit, with active sites near domain boundaries.
• Sequence of Events:
  1. Oxaloacetate Binding: Binds to an active site, inducing conformational changes in the enzyme.
  2. Acetyl-CoA Binding: Conformational change creates a binding site for Acetyl-CoA.
  3. Aldol Condensation: Forms high-energy citryl-CoA intermediate.
  4. Hydrolysis: Water cleaves the high-energy thioester bond, forming citrate and regenerating Co-enzyme A.

Detailed Mechanism

• Residues Involved: Histidine 274, Histidine 320, Aspartate 375.
• Step-by-Step:
  1. Formation of Enol Intermediate:
     • Histidine 274 donates H+ to carbonyl oxygen.
     • Aspartate 375 removes H+ from the Acetyl-CoA methyl group, forming an enol.
  2. Electrophile Activation:
     • Histidine 320 donates H+ to carbonyl oxygen of oxaloacetate, making it a better electrophile.
  3. Nucleophilic Attack: Enol's pi-bond attacks oxaloacetate's carbonyl carbon to form citryl-CoA.
  4. Conformational Change: Seals off the active site, preventing competing reactions.

Summary

• Goal: Secure formation of citryl-CoA leading to citrate formation.
• Final Result: Complete closure of the enzyme's active site to facilitate efficient and specific reaction conditions.