人参皂苷葡萄糖苷酶基因在大肠杆菌中的表达及包涵体的变复性

Expression of ginsenoside glucosidase gene in E.coli and renaturation of inclusion body

将人参皂苷葡萄糖苷酶的基因亚克隆到大肠杆菌表达载体pET32a(+)中,构建重组质粒,并转化至大肠杆菌BL21(DE3)中。对该基因进行诱导表达,并对包涵体进行变性和复性。结果显示,经终浓度为0.5mmol/L IPTG在30℃下诱导表达4h,进行SDS-PAGE分析,目的蛋白主要以包涵体形式存在并确定其分子质量约为75ku,与理论值基本相符,表明该基因得到了表达。包涵体经过变性和稀释复性后,经TLC活性检测,能够水解底物Rb1,表明具有人参皂苷葡萄糖苷酶的活性。

Ginsenoside glucosidase gene was subcloned into expression vector of pET32a(+) to construct the recombinant plasmid.The recombinant plasmid was then transformed into E.coli BL21(DE3) and induced by IPTG.The expressed products were solubilized and refolded to be an active form.The result indicated that the purpose protein of inclusion body expression was obtained under the condition of 0.5 mmol/L IPTG at 30 ℃ for 4 h.And SDS-PAGE analysis suggested its molecular weight is about 75 ku.TLC analysis showed that the target protein has activity of ginsenoside glucosidase.