On the mechanism of action of cytochrome P-450. Evaluation of homolytic and heterolytic mechanisms of oxygen-oxygen bond cleavage during substrate hydroxylation by peroxides.

Abstract

somal cytochrome P-450 (P-450~~) with aromatic hydroperoxides and peracids two transient spectral intermediates are produced reversibly: Complex C, which is directly involved in the hydroxylation pathway, and Complex D, which is formed in a nonproductive side reaction. In the present study the broad substrate specificity of P-450~~ was exploited to examine the effect of structural changes on the reaction Toluene + cumene hydroperoxide -j benzyl alcohol + cumyl alcohol Determination of Complexes C and D by stopped flow spectrophotometry and measurement of the initial rate of benzyl alcohol formation in the presence of 3-acetylpyridine adenine dinucleotide and alcohol dehydrogenase indicated that a number of m- and p-substituted toluenes and cumene hydroperoxides could be used in place of the parent compounds. The rate-limiting decay of Complex C was found to be dependent on the structure of both the peroxy compound and the hydroxylatable substrate. These results are compatible with a homolytic mechanism of oxygen-oxygen bond cleavage but not with the heterolytic formation of a common iron-oxo intermediate from the various peroxides.