重组二硫键异构酶在大肠杆菌中表达条件的统计优化  

Statistical Optimization of Protein Disulfide Isomerase Expression in E. coli

作　　者：罗曼[1] 关怡新[1] 姚善泾[1] 

机构地区：[1]浙江大学化学工程与生物工程学系,杭州310027

出　　处：《中国生物化学与分子生物学报》2005年第3期376-383,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金资助项目(No.20476093)~~

摘　　要：二硫键形成蛋白A (disulfidebondformationproteinA ,DsbA)是大肠杆菌周质胞腔中辅助多种含有二硫键的蛋白质正确折叠并具有生物学活性的一种二硫键异构酶.通过统计实验设计的方法将生产重组DsbA的发酵过程进行了优化.首先通过Plackett Burman设计挑选出了对DsbA表达量影响较大的四个因素,然后再利用杂合设计进行实验,并通过拟合得到了响应曲面函数,但该函数的驻点是鞍点,因此不具有全局的极值.最后通过约束优化得到了较佳的实验点,在该实验点下DsbA的表达量比基本培养条件下提高了5 0 .14 % .Disulfide bond formation protein A, DsbA, existing in the periplasm of E.coli, is a protein disulfide isomerase (PDI), which assists folding of the disulfide bond containing proteins to achieve biological activities. In this paper, statistical experimental designs were applied to optimize the fermentation process of recombinant DsbA in E.coli. Firstly, four most important parameters, which affect the DsbA expression, were screened among 17 parameters by Plackett-Burman design. Then hybrid design was applied to fit a response surface function, where stationary point was a saddle point with neither global maximum nor minimum. Finally, the optimal settings of fermentation parameters were calculated using the nonlinear constrained optimization method. Verification experiment followed by SDS-PAGE showed that the settings resulted in 50.14% higher expression after the optimization.

关 键 词：DsbA 统计优化 Plackett-Burman设计 杂合设计 响应面法 

分 类 号：Q78[生物学—分子生物学]