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Enhancing Expression of 3-Ketosteroid-9α-Hydroxylase Oxygenase, an Enzyme with Broad Substrate Range and High Hydroxylation Ability, in Mycobacterium sp. LY-1

Applied Biochemistry and Biotechnology, ISSN: 1559-0291, Vol: 187, Issue: 4, Page: 1238-1254
2019
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Article Description

3-Ketosteroid-9α-hydroxylase (KSH) consists of two protein systems, KshA and KshB, and is a key enzyme in microbial degradation pathway of natural sterols. 9α-Hydroxy-4-androstene-3,17-dione (9α-OH-AD) is a valuable steroid pharmaceutical intermediate. The expression of a 3-ketosteroid-9α-hydroxylase oxygenase (KshA1) with a broad substrate range and high hydroxylation ability was enhanced in Mycobacterium sp. LY-1 to improve the yield of 9α-OH-AD. Through whole-genome sequence mining and homologous comparison, the putative genes (kshA1 and kshB) in wild strain LY-1 were firstly identified. Then they were heterogeneously co-expressed in Escherichia coli BL21. The transformation results of recombinant BL21-KshA1/B demonstrated KshA1/B had high hydroxylation ability to AD. Moreover, substrate preference analysis suggested that KshA1 had a broad substrate range. After enhancing expression of kshA1 and kshB in the strain LY-1, the maximum productivity of 9α-OH-AD in recombinant LY-1-KshA1/B reached 0.064 g/L/h in a 5-L stirred fermenter.

Bibliographic Details

Li, Hui; Wang, Xiangdong; Zhou, Longfei; Ma, Yang; Yuan, Wanjuan; Zhang, Xiaomei; Shi, Jinsong; Xu, Zhenghong

Springer Science and Business Media LLC

Biochemistry, Genetics and Molecular Biology; Chemical Engineering; Immunology and Microbiology

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