Cascade biocatalysis for production of enantiopure(S)-2-hydroxybutyric acid using recombinant Escherichia coli with a tunable multi-enzyme-coordinate expression system  

作　　者：Lingzhi Tian Junping Zhou Taowei Yang Xian Zhang Meijuan Xu Zhiming Rao 

机构地区：[1]The Key Laboratory of Industrial Biotechnology,Ministry of Education,School of Biotechnology,Jiangnan University,Wuxi 214122,Jiangsu Province,China

出　　处：《Systems Microbiology and Biomanufacturing》2021年第2期234-244,共11页系统微生物学与生物制造（英文）

基　　金：This work was funded by the National Key Research and Development Program of China(2018YFA0900300);the National Natural Science Foundation of China(31770058,32070035);Natural Science Foundation of Jiangsu Province(BK20181205);the Key Research and Development Program of Ningxia Hui Autonomous Region(No.2019BCH01002);the national first-class discipline program of Light Industry Technology and Engineering(LITE2018-06);the 111 Project(111-2-06).

摘　　要：Racemize 2-hydroxybutyric acid is usually synthesized by organic methods and needs additional deracemization to obtain optically pure enantiomers for industrial application.Here we present a cascade biocatalysis system in Escherichia coli BL21 which employed L-threonine deaminase(TD),NAD-dependent L-lactate dehydrogenase(LDH)and alcohol dehydrogenase(ADH)for producing optically pure(S)-2-hydroxybutyric acid((S)-2-HBA)from bulk chemical L-threonine.To solve the mismatch in the conversion rate and the consumption rate of intermediate 2-oxobutyric acid(2-OBA)formed in the multi-enzyme catalysis reaction,ribosome binding site regulation strategy was explored to control TD expression levels,achieving an eightfold alteration in the conversion rate of 2-OBA.With the optimized activity ratio of the three enzymes and using ADH for NADH regeneration,the recombinant strain ADH-r53 showed increased production of(S)-2-HBA with the highest titer of 129 g/L and molar yield of 93%within 24 h,which is approximately 1.65 times that of the highest yield reported so far.Moreover,(S)-2-HBA could easily be purified by distillation,making it have great potential for industrial application.Additionally,our results indicated that constructing a tunable multi-enzyme-coordinate expression system in single cell had great significance in biocatalysis of hydroxyl acids.

关 键 词：Cascade biocatalysis (S)-2-hydroxybutyric acid Multi-enzyme-coordinate expression system NADH regeneration Ribosome binding site strength 

分 类 号：O62[理学—有机化学]