Stereo-complementary epoxidation of 4-vinyl-2,3-dihydrobenzofuran using mutants of SeStyA with enhanced stability and enantioselectivity

Abstract

The asymmetric epoxidation of unfunctionalized terminal olefins remains a challenge. Styrene monooxygenase SeStyA from Streptomyces exfoliatus could catalyze the enantioselective epoxidation of terminal olefins. To improve the robustness of SeStyA, a consensus-based approach was applied, which led to the successful identification of several beneficial mutations with enhanced thermostability or enantioselectivity in the epoxidation of 4-vinyl-2,3-dihydrobenzofuran (1a), a Tasimelteon intermediate. After sequential combinatory mutagenesis, mutants M6R and M5S with complementary enantioselectivity were selected to catalyze respectively the (R)- and (S)-epoxidation of 1a, generating both enantiomers with 99% ee. The inactivation half-life of M6R and M5S at 40 °C was increased to 25.4 and 5.9-fold of the wild-type, respectively. Both can catalyze the complete epoxidation of 20 mM substrate within 2 to 3 h, a significant improvement from the 20% conversion of the wild-type. The results were interpreted in the context of the model structure of SeStyA.