Regio- and Stereoselectivity of Particulate Methane Monooxygenase from Methylococcus capsulatus (Bath)

The regiospecificity and stereoselectivity of alkane hydroxylation and alkene epoxidation by the particulate methane monooxygenase from Methylococcus capsulatus (Bath) was evaluated over a range of substrates. Oxidation products were identified by conventional GC analysis, and the stereoselectivity of oxidation was determined by a combination of chiral GC and HPLC methods, as well as 1 H NMR analysis of the corresponding ( R )-2-acetoxy-2-phenylethanoate ester derivatives in the case of alkanol products. Alkane hydroxylation was found to proceed favoring attack at the C-2 position in all cases, and the stereoselectivity for n -butane and n -pentane was characterized by an enantiomeric excess of 46% and 80%, respectively, with preference for the ( R )-alcohol noted for both substrates. Epoxides were formed with smaller stereoselectivities. Together, the regio- and stereoselectivity results suggest that an equilibrium of competing substrate binding modes exists. A simple substrate-binding model that incorporates preferential C-2 oxidation with the observed stereoselectivity of alkane hydroxylation is proposed, and hypotheses for the general mechanism are suggested and discussed.