Improved 2α-Hydroxylation Efficiency of Steroids by CYP154C2 Using Structure-Guided Rational Design
Qilin Gao, Bingbing Ma, Qianwen Wang, Hao Zhang, Shinya Fushinobu, Jian Yang, Susu Lin, Keke Sun, Bingnan Han, Lian‐Hua Xu
Abstract
Hydroxylated derivatives of steroids play essential roles in medicine. Cytochrome P450 enzymes selectively hydroxylate methyne groups on steroids, which can dramatically change their polarity, biological activity and toxicity. There is a paucity of reports on the 2α-hydroxylation of steroids, and documented 2α-hydroxylate P450s show extremely low conversion efficiency and/or low regio- and stereoselectivity. This study conducted crystal structure analysis and structure-guided rational engineering of CYP154C2 and efficiently enhanced the conversion efficiency of TES and ASD with high regio- and stereoselectivity. Our results provide an effective strategy and theoretical basis for the 2α-hydroxylation of steroids, and the structure-guided rational design of P450s should facilitate P450 applications in the biosynthesis of steroid drugs.