Efficient production of 14α‑OH‑AD by engineered Mycolicibacterium neoaurum via coupled cofactor and reconstructed electron transport system  

作　　者：Chenwei Zhang Yanbing Shen Yuanyuan Gao Zehui Zan Min Wang 

机构地区：[1]Key Laboratory of Industrial Fermentation Microbiology,Ministry of Education,Tianjin Key Laboratory of Industrial Microbiology,Tianjin University of Science and Technology,Tianjin,China [2]Engineering Research Center of Microbial Metabolism and Fermentation Process Control,College of Biotechnology,Tianjin University of Science and Technology,Tianjin 300457,People’s Republic of China

出　　处：《Systems Microbiology and Biomanufacturing》2023年第2期358-369,共12页系统微生物学与生物制造（英文）

基　　金：supported by the National Key R&D Program of China,Synthetic Biology Research(no.2019YFA0905300);the National Natural Science Foundation of China(21978221);the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-KJGG-001-08);the Innovative Research Team of Tianjin Municipal Education Commission(TD13-5013);the Tianjin Municipal Science and Technology Commission(21ZYJDJC00030).

摘　　要：14α-hydroxy-androst-4-ene-3,17-dione(14α-OH-AD)is an important precursor for the synthesis of steroid drugs with anticancer and carcinolytic activity.Initially,14α-OH-AD was mostly synthesized by whole-cell fermentation of mold fungi using androstenedione(AD)as a substrate,which had difficulties in product isolation and purification as well as problems of high production cost.In this study,the source of the 14α-hydroxylase gene was expanded.And 14α-hydroxylase genes were heterologously expressed in Mycolicibacterium neoaurum(MNR)M3ΔksdD,which enabled the one-step biotransformation from the cheap substrate phytosterols(PS)to 14α-OH-AD,reducing the difficulty of product purification and production cost.What is more,to alleviate the problem of poor activity of 14α-hydroxylase,the 14α-hydroxylase gene was co-expressed with the electron transport chain element genes and the coenzyme regeneration genes,and a superior engineered strain MNR M3ΔksdD/pMV261-14α-G6PDH was obtained.Finally,the transformation conditions were optimized for the transformation of PS by the engineered strain.The molar yield of 14α-OH-AD reached to 60.4±2.3%(about 0.22 g/L productivity).This study investigated for the first time the effects of the tandem electron transport chain element genes and the tandem coenzyme regeneration genes on the 14α-hydroxylation reaction,providing a theoretical basis for the industrial production of 14α-OH-AD.

关 键 词：14α-hydroxy-androst-4-ene-3 17-dione Mycolicibacterium neoaurum 14α-hydroxylase Electron transportation Coenzyme regeneration 

分 类 号：Q93[生物学—微生物学]