Title:
METHOD FOR ENZYMATICALLY CONVERTING AND PRODUCING 9α-HYDROXYANDROST-4-ENE-3,17-DIONE
Document Type and Number:
WIPO Patent Application WO/2019/114157
Kind Code:
A1
Abstract:
Provided is a method for enzymatically converting and producing 9α-hydroxyandrost-4-ene-3,17-dione, the method comprising the steps of: expressing, in E. coli BL21, an oxidative subunit KshA, a reductive subunit KshB, and a subunit KshC, which has an unknown activity, of 3-ketosteroid-9α-hydroxylase derived from Mycobacterium sp. strain VKMAc-1817D, and identifying KshC as an oxidative subunit with a much higher enzymatic activity than KshA; expressing a formate dehydrogenase FDH by using BL21/pET-28a(+)-fdh constructed in the present laboratory, by using KSH (KshB+KshC) and the crude enzyme solution of FDH engineering strains as a biocatalyst, by using a steroid AD as a substrate and at a reaction temperature of 30℃ and a pH of 7.0; and converting AD under optimal conditions to generate the product 9-OH-AD, wherein within 20 hours, the yield of 9-OH-AD is 4.7 g/L and the molar conversion rate reaches 96.7%.
Inventors:
RAO ZHIMING (CN)
SHA ZONGYAN (CN)
ZHANG XIAN (CN)
YANG TAOWEI (CN)
XU MEIJUAN (CN)
SHA ZONGYAN (CN)
ZHANG XIAN (CN)
YANG TAOWEI (CN)
XU MEIJUAN (CN)
Application Number:
PCT/CN2018/081871
Publication Date:
June 20, 2019
Filing Date:
April 04, 2018
Export Citation:
Assignee:
UNIV JIANGNAN (CN)
International Classes:
C12P33/06; C12N9/02; C12N15/70; C12R1/19
Domestic Patent References:
| WO2001031050A1 | 2001-05-03 |
Foreign References:
| CN104232722A | 2014-12-24 | |||
| CN106119180A | 2016-11-16 |
Other References:
WANG, GUIE ET ET AL.: "Enhancement Expression of 3-Phytosterone-9a-Hydroxylase in Mycobacterium", CHINESE JOURNAL OF PHARMACEUTICAL BIOTECHNOLOGY, vol. 23, no. 5, 15 October 2016 (2016-10-15), ISSN: 1005-8915
Attorney, Agent or Firm:
HARBIN SHINEIP INTELLECTUAL PROPERTY AGENCY CO., LTD. (CN)
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