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Enzyme from engineering strain and application in 1, 3-propanediol coupling-enzymatic catalysis

PENG Yiqiang1,ZHUANG Yuan1,LIN Zhiqiang1,HUANG Wenai1,FANG Baishang1,2   

  1. 1Provincial Key Laboratory of Industrial Biotechnology,Hua Qiao University,Xiamen 361021,Fujian,China;2College of Chemistry and Chemical Engineering,Xiamen University,Xiamen 361005,Fujian,China
  • Online:2010-11-05 Published:2010-11-05

工程菌发酵产酶及其在1, 3-丙二醇耦合酶催化制备中的应用

彭益强1,庄 园1,林志强1,黄文爱1,方柏山1,2   

  1. 1华侨大学厦门校区省工业生物技术实验室,福建 厦门 361021;2 厦门大学化学化工学院,福建 厦门 361005

Abstract:

Engineering strain was acquired by transforming directly evolved plasmid from the incubated conservation bacterium E. coli BL21DE3pLysS/PET-15b-dhaT’-24 to the host cell E. coli BL21DE3pLysS. The lactose induced engineering strain was fermented to acquire 1,3-propanediol oxidoreductasePDORwith 182 U/mL activity. The optimal reaction pH was 10 and the pH stabile range was 7.09.0. The optimal reaction temperature was 55 and stabile temperature range was 3045 . 3-Hydroxypropionaldehyde3-HPAwas catalysed by the PDOR to produce 1,3-propanediol1,3-PD. The reaction was coupled with another reaction of glycerol dehydrogenaseGDHacquired from another engineering strainto realize NADH regeneration. Thus1,3-PD coupling enzymatic catalysis was constructed. Due to the two enzymes from engineering strains showed suitable characteristicsthe reaction was continued for 34 hours and 63.4% translation rate of 3-HPA64.6% 1,3-PD production rate were acquired.

摘要:

培养定向进化后的质粒保藏菌E. coli BL21DE3pLysS /PET-15b-dhaT’-24 并进行质粒抽提,将抽提的质粒转化入感受态宿主细胞E. coli BL21DE3pLysS中得产1,3-丙二醇氧化还原酶的工程菌。工程菌经乳糖诱导后进行发酵培养获得酶活为182 U/mL1,3-丙二醇氧化还原酶,最适反应pH值为10pH值稳定范围为7.09.0,最适反应温度为55 ,温度稳定范围为3045 。利用工程菌产的1,3-丙二醇氧化还原酶进行转化3-羟基丙醛为1,3-丙二醇的反应,同时偶联甘油脱氢酶(由另一工程菌制备)转化甘油的反应进行辅酶NADH的再生,实现了1,3-丙二醇的双酶耦合的连续反应。由于来源于工程菌的双酶酶学性质相适应,反应连续进行34 h后,底物3-羟基丙醛的转化率达63.4%,产物1,3-丙二醇的产率达64.6%

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