Chen Shen1, Wanli Zhao1, Xuming Liu2, Jihua Liu3,4. 1. Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing, 211198, Jiangsu, China. 2. School of Life Science and Technology, China Pharmaceutical University, Nanjing, 211198, Jiangsu, China. liuxm@cpu.edu.cn. 3. Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing, 211198, Jiangsu, China. liujihua@cpu.edu.cn. 4. State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing, 210009, Jiangsu, China. liujihua@cpu.edu.cn.
Abstract
OBJECTIVES: To investigate the regio-selective demethylation of papaverine by CYP105D1 and develop a whole-cell biocatalytic system for the preparative synthesis of 6-O-demethyl-papaverine. RESULTS: CYP105D1 from Streptomyces griseus ATCC 13273 was used for the regioselective demethylation of papaverine at C-6 using putidaredoxin reductase (PDR) and putidaredoxin (Pdx) as the electron transport system. The Km value of CYP105D1 towards papaverine was estimated to be 92.24 μM. Furthermore, a CYP105D1-based whole-cell system was established in E. coli BL21(DE3). The whole cell biotransformation condition was optimized as 25 °C, pH 7.5, 8 g (cell dry weight) L-1 whole cell biomass and 3% (v/v) PEG-200 as cosolvent. Under the optimal condition, the conversion yield of papaverine reached to 61.15% within 24 h. CONCLUSIONS: The selective demethylation of papaverine by CYP105D1 was accomplished. The CYP105D1-based whole-cell biocatalyst has a potential used for the efficient synthesis of 6-O-demethyl-papaverine.
OBJECTIVES: To investigate the regio-selective demethylation of papaverine by CYP105D1 and develop a whole-cell biocatalytic system for the preparative synthesis of 6-O-demethyl-papaverine. RESULTS: CYP105D1 from Streptomyces griseus ATCC 13273 was used for the regioselective demethylation of papaverine at C-6 using putidaredoxin reductase (PDR) and putidaredoxin (Pdx) as the electron transport system. The Km value of CYP105D1 towards papaverine was estimated to be 92.24 μM. Furthermore, a CYP105D1-based whole-cell system was established in E. coli BL21(DE3). The whole cell biotransformation condition was optimized as 25 °C, pH 7.5, 8 g (cell dry weight) L-1 whole cell biomass and 3% (v/v) PEG-200 as cosolvent. Under the optimal condition, the conversion yield of papaverine reached to 61.15% within 24 h. CONCLUSIONS: The selective demethylation of papaverine by CYP105D1 was accomplished. The CYP105D1-based whole-cell biocatalyst has a potential used for the efficient synthesis of 6-O-demethyl-papaverine.