摘要
目的:采用分子伴侣pTf16共表达的形式促进截短型人乳头瘤病毒(HPV)58亚型L1蛋白在大肠杆菌中的可溶性表达。方法:将重组表达质粒pGEX-6P-HPV58-L1转入含有分子伴侣pTf16的大肠杆菌BL21感受态细胞中,在2.0 mg/mL L-阿拉伯糖诱导表达20 min之后加入1.0 mmol/L的IPTG,18℃诱导表达24 h,使目的蛋白与伴侣蛋白共表达;重组蛋白经GST亲和纯化后去除GST标签免疫昆明鼠,ELISA检测免疫血清效价。结果:分子伴侣pTf16能显著提高目的蛋白的可溶性表达,经纯化及去除GST标签,用SDS-PAGE与Western印迹检测,约在相对分子质量50×103处出现特异性反应条带,该条带与截短型HPV58 L1蛋白预期大小相符,免疫动物后经ELISA检测小鼠血清的抗体效价为1∶6400。结论:获得了截短型HPV58 L1蛋白,动物免疫试验证实该蛋白具有较好的免疫原性,为进一步研制HPV58预防型疫苗奠定了基础。
Objective： To improve the soluble expression efficiency of truncated human papillomavirus（HPV） type 58 L1 protein in E.coli through the co-expression with molecular chaperone pTf16.Methods： The recombinant expression plasmid, pGEX-6P-HPV58-L1, was transferred into the competence of E.coli BL21 containing pTf16, to construct prokaryotic co-expression vector. The best inducing conditions of recombinant protein were, at 18℃ for 24 h with 2.0 mg/mL L-arabinose and 1.0 mmol/L IPTG. The recombinant protein GST-HPV58-L1 was obtained and purified with GST affinity column using one-step protocol. After cleavage of GST-Tag, the truncated HPV58 L1 proteins were used to immunize Kunming-mice to acquire antiserum, and antiserum was analysed by ELISA.Results： Molecular chaperone pTf16 can significantly improve the soluble expression efficiency of truncated HPV58 L1 protein in E.coli. The truncated HPV58 L1 protein was identified by SDS-PAGE and Western blot and showed a specific band at about 50 kD. The results of ELISA showed that the antiserum had the high titer（1∶6400）.Conclusion： The truncated HPV58 L1 was successfully obtained and with high immunogenicity, which laid the foundation for further development of HPV58 preventive vaccine.
出处
《生物技术通讯》
CAS
2017年第3期295-300,共6页
Letters in Biotechnology
基金
河南省高校科技创新团队和创新人才支持计划(14HASTIT027)
