文献类型：期刊文献

英文题名：Efficient Synthesis of 12-Oxochenodeoxycholic Acid Using a 12α-Hydroxysteroid Dehydrogenase from Rhodococcus ruber

作者：Shi, Shou-Cheng[1];You, Zhi-Neng[1];Zhou, Ke[1];Chen, Qi[1,2];Pan, Jiang[1,2];Qian, Xiao-Long[3];Xu, Jian-He[1,2];Li, Chun-Xiu[1,2]

机构：[1]East China Univ Sci & Technol, State Key Lab Bioreactor Engn, Lab Biocatalysis & Synthet Biotechnol, 130 Meilong Rd, Shanghai 200237, Peoples R China;[2]East China Univ Sci & Technol, Sch Biotechnol, Shanghai Collaborat Innovat Ctr Biomfg, 130 Meilong Rd, Shanghai 200237, Peoples R China;[3]Suzhou Bioforany EnzyTech Co Ltd, Econ Dev Zone, 8 Yanjiuyuan Rd, Changshu 215512, Jiangsu, Peoples R China

年份：2019

卷号：361

期号：20

起止页码：4661

外文期刊名：ADVANCED SYNTHESIS & CATALYSIS

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000484811500001)】；

基金：This work was financially supported by The National Key Research and Development Program of China (No. 2018YFC1706200), The National Natural Science Foundation of China (Nos. 21536004, 21871085, and 31500592), and the Fundamental Research Funds for the Central Universities (No. 22221818014). We are grateful to Dr. Yu-Cong Zheng and Dr. Chao Shou, both from East China University of Science and Technology, for their experimental assistance.

语种：英文

外文关键词：biocatalysis; 12 alpha-hydroxysteroid dehydrogenase; NAD(+)-dependence; structure-guided genome mining; 12-oxochenodeoxycholic acid

摘要：12 alpha-Hydroxysteroid dehydrogenase (12 alpha-HSDH) has the potential to convert cheap and readily available cholic acid (CA) to 12-oxochenodeoxycholic acid (12-oxo-CDCA), a key precursor for chemoenzymatic synthesis of the therapeutic bile acid ursodeoxycholic acid (UDCA). In this work, a native nicotinamide adenine dinucleotide (NAD(+))-dependent 12 alpha-hydroxysteroid dehydrogenase (Rr12 alpha-HSDH) from Rhodococcus ruber was identified using a structure-guided genome mining (SSGM) approach, which is based on the structure of cofactor pocket and the conserved nicotinamide cofactor binding motif alignment. Rr12 alpha-HSDH was heterologously overexpressed in Escherichia coli BL21 (DE3), purified and characterized. The purified Rr12 alpha-HSDH showed a high oxidative activity of 290 U mg(protein)(-1) toward CA, with a catalytic efficiency (k(cat)/K-M) of 5.10x10(3) mM(-1) s(-1). In a preparative biotransformation (100 mL), CA (200 mM, 80 g L-1) was efficiently converted to 12-oxo-CDCA in 1 h, with a 85% isolated yield and a space-time yield (STY) of up to 1632 g L-1 d(-1). Furthermore, Rr12 alpha-HSDH was shown to be able to catalyze the oxidation of other 12 alpha-hydroxysteroids at high substrate loads (up to 200 mM), giving the corresponding 12-oxo-hydroxysteroids in 71%-85% yields, indicating the great potential of Rr12 alpha-HSDH as a promising biocatalyst for the synthesis of various therapeutic bile acids.