应用分子伴侣共表达系统表达结核分枝杆菌编码蛋白

Co-expression of protein of M. tuberculosis with molecular chaperone

目的研究共同表达分子伴侣能否提高结核分枝杆菌编码基因的表达效率及被表达蛋白的生物活性。方法将携带有结核分枝杆菌编码基因的表达质粒Rv3790::pET16b、Rv3791::pET16b与能够同时表达3个分子伴侣DnaK、DnaJ和GrpE的质粒pkJE7共同在大肠埃希菌BL21(DE3)表达,之后对所获得的蛋白通过聚丙烯酰胺凝胶电泳和免疫印记分析(Western Blot)检测蛋白产量,并分析蛋白的活性。以Rv3790::pET16b、Rv3791::pET16b单独在BL21(DE3)内的表达作为对照。结果与分子伴侣共表达的结核分枝杆菌编码基因相对于目的基因单独表达时获得较多的可溶性蛋白,较少的包涵体和蛋白降解;共表达的蛋白也具有相对较好的活性。结论共同表达分子伴侣能够提高某些结核分枝杆菌编码蛋白的表达效率和生物活性。

Objective To elucidate the effect of chaperones co-expression on increasing the expression of protein from M. tuberculosis encoding genes, and on enhancing biology activity of the protein expressed. Methods Co-expressing the plasmids Rv3790：：pET16b and Rv3791：：pET16b which harbor protein encoding gene Rv3790 and Rv3791 of Mycobacterium tuberculosis with chaperones plasmid pkJE7 which can express 3 chaperones, DnaK, DnaJ, GrpE at the same time in E. coll. BL21 （DE3）. The yield of the expression was then checked by SDS-PAGE and Western Bloting. The biology activity of the expressed candidate proteins was analyzed by related activity assay. The expression system, which expressed the candidate genes alone in E. coll. BL21（DE3）, was used as control. Results Compared with the control, the co-expression system could produce more soluble protein, less inclusion body and less degradation of protein. When same amount of protein was used in the activity assay, the protein from the chaperone co-expression system had higher activity than that from the non-chaperone co-expression system. Conclusion Co-expression with chaperone could increase protein expression and protein activity of M. tuberculosis.