强化NADPH再生途径驱动手性β-羟基酯的高效合成  

Efficient synthesis of chiralβ-hydroxyesters drived by enhanced regeneration pathway of NADPH

作　　者：杜惠君 高晨 左振宇 李凌凌 杨忠华 Du Huijun;Gao Chen;Zuo Zhenyu;Li Lingling;Yang Zhonghua(College of Chemistry and Chemical Engineering,Wuhan University of Science and Technology,Wuhan 430081,China)

机构地区：[1]武汉科技大学化学与化工学院,湖北武汉430081

出　　处：《武汉科技大学学报》2022年第5期356-362,共7页Journal of Wuhan University of Science and Technology

基　　金：国家自然科学基金资助项目(21376184);湖北省教育厅科研计划资助项目(D20121108).

摘　　要：通过代谢调控大肠杆菌EMP途径中的NADP;依赖性甘油醛-3-磷酸脱氢酶和过表达NAD激酶,构建了羰基还原酶和NADPH再生偶联体系E.coli BL21(DE3)/pETDueT-1-gapB-bcCR&pET-28a-yfjB,以强化NADPH的内源性合成,为β-酮酯的不对称还原过程提供足够推动力。结果表明,通过EMP调控与引入NAD激酶可使胞内NADPH合成能力提高2.19倍;以4-氯乙酰乙酸乙酯为模型底物,改造后的工程菌不对称合成(R)-4-氯-3-羟基丁酸乙酯的转化效率提高了1.69倍,目标手性醇的产率高达98.18%,对映体过量值超过99%。Based on metabolic regulation of NADP;-dependent glyceraldehyde-3-phosphate dehydrogenase and overexpression of NAD kinase in EMP pathway of Escherichia coli,the engineered E.coli BL21(DE3)/pETDueT-1-gapB-bcCR&pET-28 a-yfjB which coupled carbonyl reductase with NADPH regeneration was constructed to enhance the endogenous synthesis of NADPH and provide sufficient driving force for the asymmetric reduction process ofβ-ketoesters.The results show that the intracellular NADPH synthesis capacity can be increased at 2.19-fold through the regulation of EMP and the overexpression of NAD kinase.By using 4-chloroacetoacetate as a model substrate,the conversion efficiency of the engineered E.coli asymmetrically synthesized ethyl(R)-4-chloro-3-hydroxybutyrate is increased at 1.69-fold,the yield of the(R)-CHBE is as high as 98.18%,and the enantiomeric excess value exceeds 99%.

关 键 词：NADPH再生 全细胞催化 β-羟基酯 不对称还原 对映体过量值 

分 类 号：TH122[机械工程—机械设计及理论]