Reactions of the Peroxo Intermediate of Soluble Methane Monooxygenase Hydroxylase with Ethers

Soluble methane monooxygenase (sMMO) isolated from Methylococcus capsulatus (Bath) utilizes a carboxylate-bridged diiron center and dioxygen to catalyze the conversion of methane to methanol. Previous studies revealed that a di(μ-oxo)diiron(IV) intermediate termed Q is responsible for the catalytic activity with hydrocarbons. In addition, the peroxodiiron(III) intermediate (Hperoxo) that precedes Q formation in the catalytic cycle has been demonstrated to react with propylene, but its reactivity has not been extensively investigated. Given the burgeoning interest in the existence of multiple oxidants in metalloenzymes, a more exhaustive study of the reactivity of Hperoxo was undertaken. The kinetics of single turnover reactions of the two intermediates with ethyl vinyl ether and diethyl ether were monitored by single- and double-mixing stopped-flow optical spectroscopy. For both substrates, the rate constants for reaction with Hperoxo are greater than those for Q. An analytical model for explaining the transient kinetics is described and used successfully to fit the observed data. Activation parameters were determined through temperature-dependent studies, and the kinetic isotope effects for the reactions with diethyl ether were measured. The rate constants indicate that Hperoxo is a more electrophilic oxidant than Q. We propose that Hperoxo reacts via two-electron transfer mechanisms, and that Q reacts by single-electron transfer steps.