Structural and Mechanistic Insight into the Enantioselectivity of (R)-Selective Styrene Monooxygenases: A Tug-of-War between Proximal and Distal Residues

稿件作者:Z.-P. Li, L. Wang, Y. Liu, X.-Q. Pei, M. Q. Fatmi, Z. Shen, J. Zhao, H. Lin, J. Zhou, Z.-L. Wu
通讯作者:Z.-L. Wu
刊物名称:Angew. Chem. Int. Ed
发表年份:2025
卷:64
期:
页码:e202423117
影响因子:
文章摘要:

Group E flavoprotein monooxygenases (GEMs) are well-known for catalyzing enantioselective epoxidation reactions. However, engineering their enantioselectivity remains a significant challenge, largely due to a limited understanding of the underlying mechanisms. Among these enzymes, (R)-selective styrene monooxygenases ((R)-SMOs) stand out due to their unusual enantio-switch behavior when catalyzing the reactions of p-substituted styrenes. This unique property provides an exceptional opportunity to investigate the enantiocontrol mechanisms within GEMs. In this study, we resolved the first crystal structure of an (R)-SMO, SeStyA, derived from Streptomyces. By integrating this structural information with molecular docking and molecular dynamics (MD) simulations, we identified four key residues critical to enantiodivergency: two distal residues (S178 and A219) and two proximal residues (A59 and A312). Strikingly, a “tug-of-war” mechanism was revealed through saturation mutagenesis, wherein the side-chain sizes of proximal and distal residues exerted opposing influences on enantioselectivity at the C=C bond. Leveraging this mechanistic insight, we successfully engineered SMOs with excellent (R)- or (S)-enantioselectivity.