Subject Area物理化学
Optimization of Recombinant Cytochrome P450 2C9 Protein Production in Escherichia Coli DH5α by Statistically-based Experimental Design
Hao DC(郝大程); Zhu PH(朱盼鸿); Yang SL(杨胜利); Yang L(杨凌)
Conference NameThe 1st asia Pacific ISSX Meeting
Conference Date2006-5-24
Conference Place韩国
AbstractObjective: To optimize Cytochrome P450 (CYP) 2C9 recombinant protein production in E. coli DH5α. Methods: Taguchi design and desirability function were first used to investigate the effect of medium components (glycerol, δ-ALA, IPTG, ampicillin, chloramphenicol, inoculum density, peptone, thiamine, trace element, NH4Cl, and MgSO4) on recombinant human CYP2C9 production by E. coli DH5α. An L12 (211) orthogonal array was used to design the experiments to screen out the most influential factors. The CYP concentration and the specific content of CYP were considered as two product quality variables. A desirability function was applied to combine these two qualities as a single objective function. Amp, IPTG, Cm, and peptone were found to be four significant factors. Optimization via the central composite design (CCD) was then undertaken to yield the best performance. Results: The location of optimum was determined to be Amp 169.5 μg/ml, Cm 1.92 μg/ml, IPTG 1.22 mM, and peptone 1.84 g/l obtained by the differentiation of the quadratic model. The confirmation experiments indicated that the expression performance under the optimized conditions was better than those obtained under other conditions. A compromise between conflicting goals, such as achievement of good yield of recombinant CYP2C9 and facility of the following purification, was found by means of the desirability function D. Conclusion: This is the first report that combined Taguchi design and CCD, and performed experiments in a multiresponse framework to optimize the human CYP production in a recombinant E. coli strain. (Supported by the 973 program 2003CB716006 of the Ministry of Science & Technology of China)
Document Type会议论文
Corresponding AuthorYang L(杨凌)
Recommended Citation
GB/T 7714
Hao DC,Zhu PH,Yang SL,et al. Optimization of Recombinant Cytochrome P450 2C9 Protein Production in Escherichia Coli DH5α by Statistically-based Experimental Design[C],2006:230/1.
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