INTERMEDIATE COMPOUND FORMATION WITH PEROXIDASE AND STRONG OXIDIZING AGENTS

Abstract

The discovery that the second intermediate compound formed by horseradish or cytochrome c peroxidase and hydrogen peroxide is reducible to the ferric form of the enzyme in a 1 equivalent reaction showed that these compounds are not “enzyme-substrate complexes,” for such compounds would possess 2 oxidizing equivalents in relation to ferriperoxidase, like HzOn itself or its anion OeH(1). These compounds can best be regarded as containing the iron with an effective oxidation number of +4, and several alternative chemical structures accounting for this were suggested. If peroxide is not a component part of the structure, there is a possibility that other oxidizing agents could also form the same compounds. In this paper the action of a number of strong oxidizing agents on both peroxidases is reported. Spectrophotometric measurements show that intermediate compounds are formed in some cases and determinations of enzyme activity show that several oxidizing agents can replace H,Oz in the well known peroxidase reaction in which an organic compound is oxidized. The first problem discussed is whether these compounds are identical with those formed by HzOz. It is also possible that the action of some or all of these oxidizing agents depends on an initial react.ion between the oxidizing agent and some group in the protein, or impurity present, which leads to the formation of HzOz responsible for the intermediate compound formation observed. The likelihood of this occurring is examined in the light of the quantitative data. A preliminary account of these experiments has already been published (1, 2). Horseradish peroxidase and cytochrome c peroxidase are referred to throughout by the abbreviations HRP and CcP.