Abstract
The kinetic theory of the substrate reaction during modification of enzyme activity previously described [Tsou (1988) Adv. Enzymol. Relat. Areas Mol. Biol. 61, 381-436] has been applied to a study on the kinetics of the course of inactivation of aminoacylase by 1,10-phenanthroline. Upon dilution of the enzyme that had been incubated with 1,10-phenanthroline into the reaction mixture, the activity of the inhibited enzyme gradually increased, indicating dissociation of a reversible enzyme--1,10-phenanthroline complex. The kinetics of the substrate reaction with different concentrations of the substrate chloroacetyl-L-alanine and the inactivator suggest a complexing mechanism for inactivation by, and substrate competition with, 1,10-phenanthroline at the active site. The inactivation kinetics are single phasic, showing that the initial formation of an enzyme-Zn(2+)-1,10-phenanthroline complex is a relatively rapid reaction, followed by a slow inactivation step that probably involves a conformational change of the enzyme. The presence of Zn2+ apparently stabilizes an active-site conformation required for enzyme activity.