Identification and engineering of cholesterol oxidases involved in the initial step of sterols catabolism in Mycobacterium neoaurum |
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| Institution: | 1. Molecular and Cellular Pharmacology Program, Stony Brook University, Stony Brook, NY 11794-8651, United States;2. Department of Chemistry, Stony Brook University, Stony Brook, NY 11794-3400, United States;1. Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Key Laboratory of Industrial Microbiology, College of Biotechnology, Tianjin University of Science & Technology, Tianjin 300457, China;2. College of Life Science, Liaocheng University, Liaocheng, Shandong 252059, China;3. Tianjin Engineering Research Center of Microbial Metabolism and Fermentation Process Control, 300457 Tianjin, China;1. Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China;2. State Key Laboratory of Chemical Engineering, Tianjin University, Tianjin, 300072, PR China;3. Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin, 300072, PR China;4. Tianjin Key Laboratory of Membrane Science and Desalination Technology, Tianjin University, Tianjin, 300072, PR China;5. Ever-Sky Bioscience (Tianjin) Co., Ltd., PR China;6. Biosyn Healthy Pharma Co., Ltd, PR China;1. School of Pharmaceutical Sciences, Jiangnan University, Wuxi, 214122, China;2. The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China;1. School of Life Science, Division of Life Sciences and Medicine, University of Science and Technology of China, No.443, Huangshan Road, Shushan District, Hefei, Anhui, 230027, P.R. China;2. Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, 32 West 7th Avenue, Tianjin Airport Economic Area, Tianjin, 300308, P.R. China;3. Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, P.R. China;4. National Innovation Center for Synthetic Biotechnology, Tianjin, 300308, P.R. China |
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| Abstract: | Mycobacteria have been modified to transform sterols to produce valuable steroids. Here, we demonstrated that the oxidation of sterols to sterones is a rate-limiting step in the catabolic pathway of sterols in Mycobacterium neoaurum. Two cholesterol oxidases ChoM1 and ChoM2 involved in the step were identified in M. neoaurum and the ChoM2 shared up to 45% identity with other cholesterol oxidases. We demonstrated that the combination of ChoM1 and ChoM2 plays a significant role in this step. Accordingly, we developed a strategy to overcome this rate-limiting step by augmenting the activity of cholesterol oxidases in M. neoaurum strains to enhance their transformation productivity of sterols to valuable steroids. Our results indicated that the augmentation of ChoM2 achieved 5.57 g/l androst-1,4-diene-3,17-dione in M. neoaurum NwIB-01MS and 6.85 g/l androst-4-ene-3,17-dione in M. neoaurum NwIB-R10, greatly higher than the original yield, 3.87 g/l androst-1,4-diene-3,17-dione and 4.53 g/l androst-4-ene-3,17-dione, respectively. |
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