Overexpression of cytochrome p450 125 in Mycobacterium: a rational strategy in the promotion of phytosterol biotransformation

• Published in: Journal of industrial microbiology & biotechnology 2018-10, Vol.45 (10), p.857-867
• Main Authors: Su, Liqiu, Shen, Yanbing, Xia, Menglei, Shang, Zhihua, Xu, Shuangping, An, Xingjuan, Wang, Min
• Format: Article
• Language: English
• Subjects: Biocatalysis - Original Paper; Biochemistry; Biodegradation; Bioinformatics; Biomedical and Life Sciences; Biotechnology; Biotransformation; Cytochrome P450; Cytochromes P450; Fermentation; Genetic Engineering; Inorganic Chemistry; Life Sciences; Metabolites; Microbiology; Monooxygenase; Mycobacterium; NAD; NADH; Nicotinamide adenine dinucleotide; Phytosterols; Production increases; Sterols; Strategy
• ISSN: 1367-5435; 1476-5535
• DOI: 10.1007/s10295-018-2063-z

Androst-4-ene-3, 17-dione (AD) and androst-1, 4-diene-3, 17-dione (ADD) are generally produced by the biotransformation of phytosterols in Mycobacterium . The AD (D) production increases when the strain has high NAD + /NADH ratio. To enhance the AD (D) production in Mycobacterium neoaurum TCCC 11978 ( MNR M3), a rational strategy was developed through overexpression of a gene involved in the phytosterol degradation pathway; NAD + was generated as well. Proteomic analysis of MNR cultured with and without phytosterols showed that the steroid C27-monooxygenase (Cyp125-3), which performs sequential oxidations of the sterol side chain at the C27 position and has the oxidative cofactor of NAD + generated, played an important role in the phytosterol biotransformation process of MNR M3. To improve the productivity of AD (D), the cyp125 -3 gene was overexpressed in MNR M3. The specific activity of Cyp125-3 in the recombinant strain MNR M3C3 was improved by 22% than that in MNR M3. The NAD + /NADH ratio in MNR M3C3 was 131% higher than that in the parent strain. During phytosterol biotransformation, the conversion of sterols increased from 84 to 96%, and the yield of AD (D) by MNR M3C3 was increased by approximately 18% for 96 h fermentation. This rational strain modification strategy may also be applied to develop strains with important application values for efficient production of cofactor-dependent metabolites.