title: The binding of propionyl-CoA and carboxymethyl-CoA to Escherichia coli citrate synthase.
abstract: The interaction of propionyl-CoA and acetyl-CoA with E. coli citrate synthase has been studied in order to gain insight into the structural requirements for substrate binding by this enzyme. In contrast to the enzyme from pig heart, the E. coli enzyme was unable to catalyse significant exchange of the methylene protons of propionyl-CoA while overall activity was very low with this enzyme. Carboxymethyl-CoA is a presumptive transition state analogue of acetyl-CoA using pig heart citrate synthase. The effect of carboxymethyl-CoA on both the native enzyme from E. coli and a catalytically active aspartate mutant (D362E) was investigated. Whereas the native enzyme was inhibited by carboxymethyl-CoA, the mutant enzyme (D362E) shows either no inhibition or minimal inhibition depending on the assay conditions. The binding of acetyl-CoA is not inhibited as a result of the mutation. The results with propionyl-CoA and carboxymethyl-CoA suggest that the active site of the E. coli enzyme is more restricted as compared with the enzyme from pig heart and, in the case of propionyl-CoA, this restriction prevents the formation of a catalytically productive enzyme-substrate complex.
authors.affiliation: Department of Biochemistry, University of Southampton, Bassett Crescent East, UK.
authors.forename: W J
authors.initials: WJ
authors.lastname: Man
authors.forename: Y
authors.initials: Y
authors.lastname: Li
authors.forename: C D
authors.initials: CD
authors.lastname: O'Connor
authors.forename: D C
authors.initials: DC
authors.lastname: Wilton
mesh_terms: D000214:Acyl Coenzyme A / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D002950:Citrate (si)-Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D009206:Myocardium / Q000201:enzymology; D013552:Swine
chemical_list: D000214:Acyl Coenzyme A; C009061:propionyl-coenzyme A; C064063:carboxymethyl-coenzyme A; D002950:Citrate (si)-Synthase