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Efficient biosynthesis of antitumor ganoderic acid HLDOA using a dual tunable system for optimizing the expression of CYP5150L8 and a Ganoderma P450 reductase |
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| Authors: | Xiaoting Lan Wei Yuan Meng Wang Han Xiao |
| Affiliation: | 1. State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, and Laboratory of Molecular Biochemical Engineering, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China Xiaoting Lan and Wei Yuan are co-first authors and contributed equally to this work.;2. College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, China Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, China Xiaoting Lan and Wei Yuan are co-first authors and contributed equally to this work.;3. College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China;4. State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, and Laboratory of Molecular Biochemical Engineering, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China |
| Abstract: | Ganoderic acid 3-hydroxy-lanosta-8,24-dien-26-oic acid (GA-HLDOA), an antitumor triterpenoid from the traditional Chinese medicinal higher fungus Ganoderma lucidum, is considered as a key precursor for biosynthesizing other ganoderic acids (GAs) with superior antitumor activities. Our previous study identified CYP5150L8 from G. lucidum as a lanosterol oxidase, and achieved heterologous biosynthesis of GA-HLDOA in Saccharomyces cerevisiae. However, low production of GA-HLDOA in either G. lucidum or heterologous host hindered its further investigation and application. In this study, we constructed a dual tunable system for balancing the expression of CYP5150L8 and a Ganoderma P450 reductase iGLCPR, and performed a comprehensive optimization of CYP5150L8 expression, iGLCPR expression, and glycerol usage. Then, we investigated the fermentation behavior of the best strain in optimized condition in flask and achieved 154.45 mg/L GA-HLDOA production, which was 10.7-fold higher compared with previous report. This study may facilitate the wide-spread application of GA-HLDOA and the discovery of unknown cytochrome P450s in downstream GAs biosynthesis. |
| Keywords: | cytochrome P450 reductase cytochrome P450s ganoderic acid HLDOA metabolic engineering Saccharomyces cerevisiae |