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Studies on the 3-Ketosteriod-1-Dehydrogenation of Steroid Hormone by Cellular lysates of Mycobacterium |
QIN Meng-fei, SUN Hong, SONG Hao |
School of Chemical Engineering and Technology, Key Laboratory of Systems Bioengineering, Ministry of Education, SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering, Tianjin University, Tianjin 300072, China |
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Abstract 9β, 11β-Epoxypregn-4-ene-17α, 21-diol-3, 20-dione 21-acetate (Ⅰ) is a substrate for the production of 9β, 11β-Epoxypregn-1, 4-diene-17α, 21-diol-3, 20-dione (Ⅳ), which is a key precursor for the production of many 9-fluoro-substituted corticosteroid hormones. By comparing whole cells catalysis and cellular lysates conversion, it was found that whole cells of Mycobacterium sp. MS136 could only convertⅠto 9β, 11β-Epoxypregn-4-ene-17α, 21-diol-3, 20-dione (Ⅱ), and Ⅰ can be effectively converted toⅣ by cellular lysates.The reaction order is that Ⅰ is spontaneously hydrolyzed to Ⅱ and Ⅱundergoes C1, 2-dehydrogenation reaction toⅣ.In order to improve the productivity of Ⅳ, the key genes kstD, kstD3 and kstDM encoding C1, 2-dehydrogenase (KSTD)were overexpressed in Mycobacterium sp. MS136 to enhance the C1, 2-dehydrogenation reaction rate, and the results showed that 1 g/L substrate Ⅰ can be converted by recombinant strain MS136-kstDM cellular lysates at pH 7.0, the productivity of Ⅳreached 78.4% after 45 h, which is 38.9% higher than original strain.The reaction rate is enhanced by optimizing the pH, and the results showed that 1 g/L substrate (Ⅰ) can be converted by recombinant strain MS136-kstDM cellular lysates at pH 7.5, the productivity of Ⅳreached 92.8% after 45 h, which was 63.4% higher than original strain.
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Received: 23 February 2017
Published: 25 August 2017
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