多酶一步法还原石胆酸制备熊去氧胆酸

Multi-Enzymatic One-Pot Reduction of Lithocholic Acid to Ursodeoxycholic Acid

摘要:

熊去氧胆酸是一种重要的治疗性胆汁酸，特别是在治疗胆囊炎和胆固醇型胆结石方面有重要应用。利用大肠杆菌全细胞系统中的P450单加氧酶CYP107D1（OleP），在石胆酸7β-位置区域进行选择性和立体选择性羟基化反应产生熊去氧胆酸。一步法生物合成熊去氧胆酸可以避免残酷的活熊胆汁提取和低效率的化学合成。通过对OleP 的突变体F84Q/S240A/V291G（OlePs）与氧化还原伴侣蛋白的共表达组合设计，构建了一种高效合成熊去氧胆酸的重组表达菌株Escherichia coli BL21/PDR-PDX-SD-AS-OlePs。基于重组菌株E. coli BL21/PDR-PDX-SD-AS-OlePs的全细胞催化反应，熊去氧胆酸产量显著提升，产率为88.28%，比未共表达氧化还原伴侣时的产率提升73.96%。该研究结果在一步法还原石胆酸制备熊去氧胆酸方面取得进展，为以生物废料石胆酸为底物制备熊去氧胆酸开辟了新的合成途径。

Abstract:

Ursodeoxycholic acid is an important therapeutic bile acid, especially in the treatment of cholecystitis and the prevention of cholesterol-type gallstones. The authors produced ursodeoxycholic acid through regioselective and stereoselective hydroxylation at the 7β-position of lithocholic acid using the P450 monooxygenase CYP107D1 (OleP) in E. coli whole-cell system. Ursodeoxycholic acid was biosynthesized using an efficient one-step method, avoiding the cruel practice of extracting bile from live bear and the inefficiencies of chemical synthesis. Through the design of the co-expression combination of the mutant F84Q/S240A/V291G (OlePs) of OleP and the redox chaperone, a recombinant expression strain Escherichia coli BL21/PDR-PDX-SD-AS-OlePs was developed. Utilizing this recombinant strain, the whole-cell catalytic reaction of E. coli BL21/PDR-PDX-SD-AS-OlePs led to a substantial increase in a significantly increased yield of ursodeoxycholic acid yield at 88.28%, which was 73.96% higher than that of without the co-expression of redox chaperone. The author has made progress in the preparation of ursodeoxycholic acid through one-step reduction of lithocholic acid, offering possibilities for the synthesis of ursodeoxycholic acid from biowaste lithocholic acid.