Steady-State Kinetics of Enzyme Reactions in the Presence of Added Nucleophiles

Abstract

Many enzyme-catalyzed reactions, such as hydrolyses, give rise to two products P1 and P2 which are formed in different reaction steps. The second product P2 is frequently formed by hydrolysis of an intermediate such as an acyl-enzyme or a phosphoryl-enzyme. An alternative nucleophile N introduced into the system forms an additional product P3. The present paper is concerned with the kinetics of formation of P1, P2, and P3 in the presence of added nucleophiles. A number of alternative mechanisms are considered, and equations are derived for the rates of formation of the three products, and the Michaelis constant, as functions of nucleophile concentration. Graphs are presented showing the variations of these parameters with the concentration of N, for a variety of special cases. Special attention is given to the possibility of specific binding sites for the water and the nucleophile molecules.The data for a number of enzyme systems are discussed with reference to the treatment. For reactions catalyzed by alkaline phosphatase it is concluded that only one mechanism (mechanism VI) is consistent with the results.