Inactivation of citrate lyase by oxaloacetate and its structural analogues

Abstract

• 1.

  1. The inactivation of citrate lyase (citrate-oxaloacetate lyase, EC 4.1.3.6) by oxaloacetate has been investigated.

• 2.

  2. Studies of the pH profiles of inactivation at pH 7–9 with varying total oxaloacetate and magnesium concentrations show that the magnesium complexes of enolic oxaloacetate are involved.

• 3.

  3. The inhibitory effects of the following structural analogues of the keto and enol forms of oxaloacetate were examined: l-malate, α,α-dimethyloxaloacetate, tartronate, α,α-difluorooxaloacetate, pyruvate, ketomalonate, and isomalate. These results, and the effect of other divalent metal cations, indicate that the site of the inactivation is identical with the active site for citrate cleavage.

• 4.

  4. The inactivation is irreversible and is time and concentration dependent. Free oxaloacetate does not inactivate the enzyme.

• 5.

  5. The inactivation phenomenon has high structural specificity, requiring a straight-chain, four-carbon dicarboxylic acid with an ionisable α-hydroxy group, and the presence of a divalent metal cation.