第一作者简介：彭益强（1973—），男，博士研究生。联系人：方柏
山，教授，博导，从事生物化工研究。E-mail [email protected]。[中文摘要]培养定向进化后的质粒保藏菌E.coli BL21(DE3)pLysS/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 BL21(DE3)pLysS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/mL的1,3-丙二醇氧化还原酶,最适反应pH值为10,pH值稳定范围为7.0～9.0,最适反应温度为55℃,温度稳定范围为30～45℃。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%。
[英文摘要]Engineering strain was acquired by transforming directly evolved plasmid from the
incubated conservation bacterium E. coli BL21（DE3）pLysS/PET-15b-dhaT’-24 to the host cell E. coli
BL21（DE3）pLysS. The lactose induced engineering strain was fermented to acquire 1,3-propanediol
oxidoreductase（PDOR）with 182 U/mL activity. The optimal reaction pH was 10 and the pH stabile
range was 7.0—9.0. The optimal reaction temperature was 55 ℃ and stabile temperature range was
30 — 45 ℃ . 3-Hydroxypropionaldehyde （ 3-HPA ） was catalysed by the PDOR to produce
1,3-propanediol（1,3-PD）. The reaction was coupled with another reaction of glycerol dehydrogenase
（GDH，acquired from another engineering strain）to realize NADH regeneration. Thus，1,3-PD
coupling enzymatic catalysis was constructed. Due to the two enzymes from engineering strains
showed suitable characteristics，the reaction was continued for 34 hours and 63.4% translation rate of
3-HPA，64.6% 1,3-PD production rate were acquired.国家863计划子课题(2006AA020103)，国家自然科学基金资助项目(20676048
