代谢工程改造多粘芽孢杆菌及合成光学纯(R,R)-2,3-丁二醇  

Metabolic Engineering Transforming Paenibacillus polymyxa and Producing Enantiomerically Pure(R,R)-2,3-Butanediol

作　　者：莫棋文 陈先锐 李燕婷 彭龙云 黄艳燕[1] 李检秀[1] 黄日波[1] Mo Qiwen;Chen Xianrui;Li Yanting;Peng Longyun;Huang Yanyan;Li Jianxiu;Huang Ribo(Guangxi Key Laboratory of Bio-refinery,Guangxi Biomass Engineering Technology Research Center,National Engineering Research Center for Non-food Biorefinery,State key Laboratory of Non-food Biomass and Enzyme Technology,Guangxi Academy of Sciences,Nanning,530007;College of Life Science and Technology,Guangxi University,Nanning,530004)

机构地区：[1]广西科学院非粮生物质酶解国家重点实验室,国家非粮生物质能源工程技术研究中心,广西生物质工程技术研究中心,广西生物炼制重点实验室,南宁530007 [2]广西大学生命科学与技术学院,南宁530004

出　　处：《基因组学与应用生物学》2021年第4期1643-1649,共7页Genomics and Applied Biology

基　　金：国家自然科学基金项目(21868007,21466007);广西重大科技创新基地建设项目(2018-15-Z03);广西自然科学基金项目(2018GXNSFBA281017)共同资助。

摘　　要：(R,R)-2,3-BD是一种重要的四碳平台化合物,在液晶材料、高附加值手性化合物,尤其是不对称合成光学纯药物等方面有天然优势。将来源于多粘芽孢杆菌(Paenibacillus polymyxa) DSM 365的α-乙酰乳酸合成酶(α-acetolactate synthase)基因alsS、α-乙酰乳酸脱羧酶(α-acetolactate decarboxylase)基因alsD和(R,R)-2,3-丁二醇脱氢酶(2,3-butanediol dehydrogenase)基因R,R-bdh与表达载体pMA5连接,导入多粘芽孢杆菌P. polymyxa DSM 365中加强(R,R)-2,3-丁二醇的主代谢途径,构建可高效合成(R,R)-2,3-丁二醇的多粘芽孢杆菌工程菌株DM-5。利用工程菌株DM-5补料分批发酵60 h,(R,R)-2,3-丁二醇产量达54.91 g/L,得率为0.52 g/g,生产强度为0.92 g·L^(-1)·h^(-1),与野生菌株相比(R,R)-2,3-丁二醇产量增加19.66%,且副产物甲醇浓度不变,乙醇、乙偶姻积累下降。本研究结果表明,在多粘芽孢杆菌中过量表达关键基因alsS、alsD和R,R-bdh能够显著提高(R,R)-2,3-丁二醇的产量和生产强度,为多粘芽孢杆菌的代谢工程改造和工业化生产(R,R)-2,3-丁二醇提供参考。(R,R)-2,3-butanediol is an important four-carbon platform compound with a variety of applications in liquid crystal materials, high value added chiral compounds, especially in asymmetric synthesis of optical active pharmaceuticals. α-acetolactate synthase gene alsS, α-acetolactate decarboxylase gene alsD and(R,R)-2,3-butanediol dehydrogenase gene R,R-bdh from Paenibacillus polymyxa DSM365 were connected to the expression vector pMA5. The main metabolic pathway of(R,R)-2,3-butanediol was enhanced by introducing P. polymyxa DSM-365 to construct an engineering P. polymyxa strain DM-5 which can efficiently synthesize(R,R)-2,3-butanediol. The production of(R,R)-2,3-butanediol via DM-5 was up to 54.91 g/L in fed-batch fermentation, with a yield of 0.52 g/g and a productivity of 0.92 g·L^(-1)·h^(-1) after 60 h. Comparing with the parent strain, the yield of(R,R)-2,3-butanediol increased by 19.66%, and the concentration of ethanol and acetoin decreased, methanol with little change. The results of this study showed that the overexpression of key genes alsS, alsD and R,R-bdh in P. polymyxa could significantly improve the yield and production intensity of(R,R)-2,3-butanediol, providing reference for metabolic engineering and industrial production of(R,R)-2,3-butanediol in P. polymyxa.

关 键 词：(R R)-2 3-丁二醇 代谢工程 补料分批发酵 多粘芽孢杆菌 

分 类 号：TQ923[轻工技术与工程—发酵工程]