Engineering Regioselectivity of P450 BM3 Enables the Biosynthesis of Murideoxycholic Acid by 6β-Hydroxylation of Lithocholic Acid.

Autor: Deng F; MOE International Joint Research Laboratory on Synthetic Biology and Medicines, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, P. R. China., Zhou Z; MOE International Joint Research Laboratory on Synthetic Biology and Medicines, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, P. R. China., Du Z; MOE International Joint Research Laboratory on Synthetic Biology and Medicines, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, P. R. China., Mohany M; Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia., Wu Q; NMPA Key Laboratory for Quality Control of Blood Products, Guangdong Institute For Drug Control, Guangzhou, P. R. China., Liang W; NMPA Key Laboratory for Quality Control of Blood Products, Guangdong Institute For Drug Control, Guangzhou, P. R. China., Zhang L; MOE International Joint Research Laboratory on Synthetic Biology and Medicines, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, P. R. China., Li S; MOE International Joint Research Laboratory on Synthetic Biology and Medicines, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, P. R. China.
Jazyk: angličtina
Zdroj: Biotechnology journal [Biotechnol J] 2024 Nov; Vol. 19 (11), pp. e202400518.
DOI: 10.1002/biot.202400518
Abstrakt: Murideoxycholic acid (MDCA), as a significant secondary bile acid derived from the metabolism of α/β-muricholic acid in rodents, is an important component in maintaining the bile acid homeostasis. However, the biosynthesis of MDCA remains a challenging task. Here, we present the development of cytochrome P450 monooxygenase CYP102A1 (P450 BM3) from Bacillus megaterium, employing semi-rational protein engineering technique. Following three rounds of mutagenesis, a triple variant (T260G/G328A/L82V) has been discovered that proficiently catalyzes the 6β-hydroxylation of lithocholic acid (LCA), thereby generating MDCA with an impressive 8.5-fold increase in yield compared to the template P450 BM3 mutant. The MDCA selectivity has been also promoted from 62.0% to 96.3%. This biocatalyst introduces a novel approach for the biosynthesis of MDCA from LCA. Furthermore, molecular docking and dynamics simulations have been employed to unravel the molecular mechanisms underlying the enhanced LCA conversion and MDCA selectivity.
(© 2024 Wiley‐VCH GmbH.)
Databáze: MEDLINE
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