Optimizing expression of recombinant human monocyte chemoattractant protein-1 in E.coli
MIAO Hong, GUO Bao-Yu, YANG Xu, YUAN Peng-Qun (Department of Biochemical Pharmacy, College of Pharmacy,Second Military Medical University, Shanghai 200433,China)
Objective: To optimize the expression of recombinant human monocyte chemoattractant protein (rhuMCP-1) in E.coli DE3. Methods:With NBS-MICROS 15 L T.DR fermentor, pGEX-IN/huMCP-1 was constructed by our laboratory. Four parameters including pH,temperature,agitation rate and concentration of IPTG were studied by orthogonal experimental design. Results: It was found that the expression level was greatly affected by the amount of dissolved oxygen. This indicated that the agitation rate and ventilation amount were the most important parameters during fermentation. Examined by SDS-PAGE and gel scanning, the expression level of total protein was over 40% when agitation rate was 300 r/min and ventilation amount was 10 L/min. Conclusion: A method for high-level expression of huMCP-1 on pilot-scale is established, and it will be useful for large-scale industrial production of target protein.