Inactivation of yeast alpha-isopropylmalate synthase by CoA. Antagonism between CoA and adenylates and the mechanism of CoA inactivation.
Yeast alpha-isopropylmalate synthase (EC 4.1.3.12) is inactivated by micromolar concentrations of CoA in the presence of Zn2+. We report here that rapid reactivation of inactivated enzyme (full recovery in less than 10 min) occurred in the presence of millimolar concentrations of ATP or ADP, using permeabilized cells. With purified, CoA-inactivated enzyme, ATP had only a weak reactivating effect which increased drastically, however, when a chelator was added at a concentration (0.1 mM) which by itself had little effect. Higher concentrations of chelator (1 mM) caused rapid reactivation even in the absence of ATP. Reactivation was also possible by removing CoA from equilibrium with oxidized glutathione, with acetyl phosphate in the presence of phosphotransacetylase, or by dialysis; however, these processes were very slow. Protection against CoA inactivation of alpha-isopropylmalate synthase was provided by high concentrations of ATP and, to a much lesser extent, ADP, by a high adenylate energy charge, by chelators, and by 3'-dephospho-CoA. Enzyme which had been inactivated with [3H]CoA did not retain any radioactivity (above control) when extracted with phenol. This result, together with other observations, is interpreted to mean that inactivation does not involve covalent modification, but is more likely the result of the formation of an enzyme.CoA.zinc complex held together by noncovalent forces. The physiological significance of the CoA effect is discussed.
D015079:2-Isopropylmalate Synthase / Q000037:antagonists & inhibitors*; D000244:Adenosine Diphosphate / Q000494:pharmacology*; D000249:Adenosine Monophosphate / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D003065:Coenzyme A / Q000494:pharmacology*; D007700:Kinetics; D007652:Oxo-Acid-Lyases / Q000037:antagonists & inhibitors*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D015032:Zinc / Q000494:pharmacology
D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D015079:2-Isopropylmalate Synthase; D007652:Oxo-Acid-Lyases; D015032:Zinc; D003065:Coenzyme A