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

• Published in: Molecular catalysis 2023-04, Vol.540, p.113055, Article 113055
• Main Authors: Chen, Qiang, Xiao, Hu, Li, Zhi-Pu, Pei, Xiao-Qiong, Yang, Wen, Liu, Yan, Wu, Zhong-Liu
• Format: Article
• Language: English
• Subjects: Consensus; Epoxidation; Monooxygenase; Streptomyces; Tasimelteon
• ISSN: 2468-8231; 2468-8231
• DOI: 10.1016/j.mcat.2023.113055

•Thermostabilizing mutations of SeStyA were identified via the consensus approach.•Half-life of combinatory mutants at 40 °C increased up to 25-fold of the wild-type.•Both (R)- and (S)-epoxide were achieved from 4-vinyl-2,3-dihydrobenzofuran.•Enantio-purity of (R)-epoxide was increased from 78% ee to 99% ee.•Inversion of enantioselectivity afforded (S)-epoxide with 99% ee. 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. [Display omitted]