Metabolism — citric acid cycle (step 2)
Aconitase: citrate to isocitrate isomerization
How the Aconitase works, in 3 steps. Each step shows the structures and the curved arrows that move the electrons.
In short. A 3-step mechanism. Typical reagents: Citrate, [4Fe-4S] cluster (Fe₄ binds OH), enzyme base (Ser) and acid. Conditions: Mitochondrial matrix, pH ≈ 7.4, 37 °C. Bonds formed: C=C (cis-aconitate); then C–O at the other carbon; bonds broken: C–O (tertiary); C–H; then C=C π. Stereochemistry: Anti elimination; the intermediate flips so water adds to the opposite carbon. Rate and selectivity: Near-equilibrium; citrate:isocitrate ≈ 90:10 at equilibrium.
At a glance
Step 1: Fe–S polarizes the OH, E1cB-type loss of water
An iron-sulfur cluster in the enzyme's active site polarizes citrate's hydroxyl, helping it leave as water in an E1cb-type dehydration — the same elimination logic covered throughout this library's aldol condensation and Grob fragmentation entries, just enzyme-facilitated.
Step 2: cis-Aconitate stays bound
This forms a bound intermediate, cis-aconitate, still held in the active site rather than released into solution.
Step 3: Water re-adds at the other carbon
Water re-adds across the double bond at a different carbon than it left from — a formal dehydration/rehydration isomerization that relocates the hydroxyl group, converting the tertiary alcohol of citrate (which can't be directly oxidized) into the secondary alcohol of isocitrate (which can) — setting up the very next step's oxidation.
Abbreviations and terms
- H2
- Hydrogen gas. Reductant for catalytic hydrogenation.
- E1cB
- Elimination, unimolecular conjugate base. Deprotonation first to a carbanion, then loss of the leaving group.
- RE
- Reductive elimination. Two groups on a metal couple and leave, lowering the oxidation state by two.
- Ser
- Serine. Amino acid with a nucleophilic –OH.
How it compares with related reactions
| Aconitase | Citrate synthase | Fumarase | |
|---|---|---|---|
| Reagents | Citrate, [4Fe-4S] cluster (Fe₄ binds OH), enzyme base (Ser) and acid | Acetyl-CoA, oxaloacetate, citrate synthase (His274, His320, Asp375) | Fumarate, H₂O, active-site base (Ser/His) and acid |
| Conditions | Mitochondrial matrix, pH ≈ 7.4, 37 °C | Mitochondrial matrix, pH ≈ 7.8, 37 °C | Mitochondrial matrix, pH ≈ 7.4 |
| Stereochemistry | Anti elimination; the intermediate flips so water adds to the opposite carbon | Enolate attacks the si face of oxaloacetate; pro-S citrate | Anti addition on one face gives L-malate (S) only |
| Rate / selectivity | Near-equilibrium; citrate:isocitrate ≈ 90:10 at equilibrium | Hydrolysis of citryl-CoA (ΔG°′ ≈ −32 kJ/mol overall) makes the step irreversible | Near diffusion-limited; reversible |
Test yourself
Try to answer each question before opening it.
Which reagents and conditions are typical for the Aconitase?
Citrate, [4Fe-4S] cluster (Fe₄ binds OH), enzyme base (Ser) and acid. Conditions: Mitochondrial matrix, pH ≈ 7.4, 37 °C.
Which bonds are formed and which are broken?
Formed: C=C (cis-aconitate); then C–O at the other carbon. Broken: C–O (tertiary); C–H; then C=C π.
What is the stereochemical outcome?
Anti elimination; the intermediate flips so water adds to the opposite carbon
What controls the rate or the selectivity?
Near-equilibrium; citrate:isocitrate ≈ 90:10 at equilibrium
Study guides that help with this reaction
- How to learn reaction mechanisms (without memorising everything)
- Curved-arrow notation: rules and the mistakes students make