作　　者： ; ; ; ; ; ;

机构地区： 中国人民解放军第一军医大学

出　　处： 《第一军医大学学报》 2003年第3期230-232,共3页

摘　　要： 目的表达和纯化福氏志贺菌毒力蛋白IpaC,研究福氏志贺菌的致病机制。方法将含有ipaC基因的pET32a-i-paC表达质粒载体转化大肠杆菌BL21(λDE3),在异丙基硫代-β-D半乳糖苷(IPTG)诱导下表达,对诱导后的表达产物进行SDS-PAGE鉴定,并采用QIA expressionistTM蛋白纯化系统来纯化目的蛋白。结果诱导后的表达产物经SDS-PAGE发现有一相对分子质量约为63 000的条带,其含量约占总蛋白量的11%,以350 mmol/L咪唑洗脱液洗脱,目的蛋白纯度可达90%以上。结论pET32a-ipaC重组表达质粒转入大肠杆菌后,可稳定、高效地表达目的蛋白,QIA-expressionistTM蛋白纯化系统是一种简便、高效的纯化系统,可获得高纯度的目的蛋白。 Objective To induce the expression of and purify invasion plasmid antigen C (IpaC) of Shigella flexneri for study-ing the pathogenesis of Shigella flexneri. Methods Prokaryotic expression plasmid pET32a-ipaC was constructed and incorpo-rated into E.coli BL21 (λDE3). The engineered bacteria were induced by isopropyl-β-D-thiogalactopyranoside (IPTG) to ex-press IpaC, which was identified by SDS-PAGE and purified by QIA expressionistTM system. Results SDS-PAGE presented a band for the fusion protein with the relative molecular mass of approximately 63 000, whose expression reached up to 11% of the total protein of E.coli BL21(λDE3). After proper purification, a purity of the target fusion protein of over 90% was achieved when the concentration of imidazole for elution was 350 mmol/L. Conclusion The recombinant plasmid pET32a-i-paC has been stably and efficiently expressed in E.coli BL21 (λDE3), and QIA expressionistTM purification system proves to be simple and highly efficient.

关 键 词： 福氏志贺菌 毒力蛋白 原核表达 蛋白纯化

领　　域： [生物学] [生物学]