Efficient stereoselective hydroxylation of deoxycholic acid by the robust whole-cell cytochrome P450 CYP107D1 biocatalyst
Chixiang Sun, Baodong Hu, Yanchun Li, Zhimeng Wu, Jingwen Zhou, Jianghua Li, Jian Chen, Guocheng Du, Xinrui Zhao
Abstract
Deoxycholic acid (DCA) has been authorized by the Federal Drug Agency for cosmetic reduction of redundant submental fat. The hydroxylated product (6β-OH DCA) was developed to improve the solubility and pharmaceutic properties of DCA for further applications. Herein, a combinatorial catalytic strategy was applied to construct a powerful Cytochrome P450 biocatalyst (CYP107D1, OleP) to convert DCA to 6β-OH DCA. Firstly, the weak expression of OleP was significantly improved using pRSFDuet-1 plasmid in the E. coli C41 (DE3) strain. Next, the supply of heme was enhanced by the moderate overexpression of crucial genes in the heme biosynthetic pathway. In addition, a new biosensor was developed to select the appropriate redox partner. Furthermore, a cost-effective whole-cell catalytic system was constructed, resulting in the highest reported conversion rate of 6β-OH DCA (from 4.8 % to 99.1 %). The combinatorial catalytic strategies applied in this study provide an efficient method to synthesize high-value-added hydroxylated compounds by P450s.