Competing modes of peroxyacid flux through cytochrome P-450.
In the presence of peroxyphenylacetic acid and a hydroxylatable substrate, cytochrome P-450LM2 catalyzes two reactions which proceed concurrently, decarboxylation of the peroxyacid and hydroxylation of substrate. While the first process is definitely the result of homolytic cleavage of the peroxide O--O bond, the second may involve a different mechanistic pathway. We have undertaken to determine the relationship between these two processes through a kinetic analysis of the system. Seven different mechanistic schemes are advanced to account for the two processes. The two reactions were found to have different apparent Michaelis constants for peroxyacid and different inhibition constants for cyanide. Since the ratio of the two products is saturable at high substrate concentrations, the decarboxylation reaction can proceed from both substrate-bound and substrate-free enzyme. By appropriate manipulation of the rate equations it is possible to derive expressions for the ratio of hydrogen abstraction rate constants for a series of p-substituted toluene substrates compared to toluene. The nature of the correlation of these expressions with delta in a Hammett plot allowed some of the schemes to be eliminated. After consideration of all the data, we concluded that the processes of decarboxylation and hydroxylation occur in separate pathways and do not involve any common intermediate beyond the ferric resting state of the enzyme.
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006900:Hydroxylation; D007700:Kinetics; D008433:Mathematics; D008862:Microsomes, Liver / Q000378:metabolism; D010648:Phenylacetates / Q000378:metabolism*; D011817:Rabbits; D013379:Substrate Specificity
D010648:Phenylacetates; C027720:peroxyphenylacetic acid; D003577:Cytochrome P-450 Enzyme System