The effects of chemical penetrators on NADP biosynthesis by recombinant E. coli BL21 (DE3) pET30α (+) -Nmnat were investigated. The single factor screening showed that four kinds of permeable agents such as Triton X-100, Tween 80, dimethyl sulfoxide and cetyltrimethyl ammonium bromide (CTAB) all could increase the catalytic yield of the whole cell, however, the difference was not significant. Then, the factorial design was used to investigate the composite effects of the four penetrators on the whole cell catalysis. Except that the interactions of Triton X-100 with Tween 80 and Triton X-100 with CTAB (P> 0.05), were not significant. All the other interaction items (P < 0.05) showed significant difference, and the amount of CTAB was only increased in the composite penetrator. The synergy of the composite system would be reduced, and the restriction would be increased, so it was not suitable for compound formulation. Finally, the central composite design was applied to obtain the best ratio of the penetrators. The results showed that when 1.64％Triton X-100, 2.53％ Tween 80 and 1.06％ dimethyl sulfoxide were combined, the predicted NADP yield could reach the maximum 77.46％ in accordance with the verified yield, which revealed that the experimental optimization model had good predictability. The result of optimization was 47％ higher than that of no penetrator. It indicated that optimized formula for composite penetrators was beneficial to improving the NADP biosynthesis by recombinant bacteria.%本文以重组大肠杆菌E.coli BL21(DE3)p ET30α(+)-Nmnat为研究对象,考察化学通透剂处理重组菌对其催化合成NADP的影响.实验显示,在单因素筛选实验中,四种通透剂曲拉通X-100、吐温80、二甲基亚砜和十六烷基三甲基溴化铵(CTAB)对细胞进行通透化处理均可提高全细胞催化产率,且提高幅度差距并不显著.进一步采用析因设计考察四种通透剂的复合效应,除了曲拉通X-100与吐温80和曲拉通X-100与CTAB交互作用的P大于0.05为不显著,其余所有交互作用项P值均小于0.05,呈现显著性,且复合通透剂中,仅单一增加CTAB量,复合体系的协同作用将减少,制约作用增加,不适宜用于复合配方.最后采用中心组合设计进一步对复配效果进行优化,通过拟合得到最佳通透剂配比为曲拉通X-100(1.64％),吐温80(2.53％)以及二甲基亚砜(1.06％),此时NADP预测产率达到77.46％,与实际验证产率均值77.05％相差甚微,说明实验优化模型具有良好预测性,优化结果与未添加通透剂相比提升近47％,表明优化复合通透剂配方更有利于提高重组菌催化合成NADP.
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