P450DA Monooxygenase-Catalyzed Chemoselective and Enantiodivergent Epoxidation of Unactivated Alkenes

Abstract

While enzymatic epoxidation of activated olefins by P450s has been well-established, chemo- and enantioselective epoxidation of unactivated olefins remains a formidable challenge, mainly due to the presence of competing hydroxylation of allylic C–H bonds. In addition, P450 monooxygenase-catalyzed epoxidation of olefins generally provides S-configured products with high enantiopurity, and examples of P450 enzymes demonstrating high R-enantioselectivity in epoxidation reactions remain rare. Herein, we report a chemoselective and enantiodivergent epoxidation of unactivated alkenes using engineered P450DA monooxygenases. The P450DA variants, obtained through structure-guided directed evolution based on the X-ray of P450DA-WT and P450DA-M3, switch the reactivity from the native hydroxylation of the allylic C–H bonds to epoxidation of C═C bonds and exhibit superior chemoselectivity (up to 99% epoxidation selectivity) and enantioselectivity (up to >99:1 er), delivering a wide variety of versatile and enantioenriched epoxides. Notably, an enantiodivergent synthesis was achieved simply by employing different P450DA variants, leading to both enantiomers of the epoxide products. Various transformations of the products were carried out, illustrating the synthetic utilities of the methods. Furthermore, molecular dockings and molecular dynamics simulations reveal the origin of high epoxidation selectivity and complementary stereoselectivity of the mutants.