Engineering P450 Peroxygenase to Catalyze Highly Enantioselective Epoxidation of cis-β-Methylstyrenes.

P450 119 peroxygenase and its site-directed mutants are discovered to catalyze the enantioselective epoxidation of methyl-substituted styrenes. Two new site-directed P450 119 mutants, namely T213Y and T213M, which were designed to improve the enantioselectivity and activity for the epoxidation of styrene and its methyl substituted derivatives, were studied. The T213M mutant is found to be the first engineered P450 peroxygenase that shows highly enantioselective epoxidation of cis-β-methylstyrenes, with up to 91 % ee. Molecular modeling studies provide insights into the different catalytic activity of the T213M mutant and the T213Y mutant in the epoxidation of cis-β-methylstyrene. The results of the calculations also contribute to a better understanding of the substrate specificity and configuration control for the regio- and stereoselective peroxygenation catalyzed by the T213M mutant.