Kinetic study of the reaction between cytochrome c peroxidase and hydrogen peroxide. Dependence on pH and ionic strength

Abstract

The rate of the reaction between cytochrome c peroxidase and hydrogen peroxide was investigated using the stopped-flow technique. The apparent bimolecular rate constant was determined between pH 3.3 and pH 11 as a function of ionic strength. The pH dependence of the apparent bimolecular rate constant can be explained by assuming that two ionizable groups on the enzyme strongly influence the rate of the reaction. At 0.1 M ionic strength, a group with a pKa of 5.5 must be unprotonated and a group with a pKa of 9.8 must be protonated for the enzyme to react rapidly with hydrogen peroxide. The apparent acid dissociation constants depend upon the ionic strength. The true bimolecular rate constant has a value of (4.5 +/- 0.3) X 10(7) M-1 sec-1 and is independent of ionic strength.