HPV31和52L2融合蛋白的原核高效表达及免疫效果评价

The high expression of HPV31 and 52 L2 fusion protein in E. coli and detection of its immunogenicity

目的高效表达HPV31和52型L2融合蛋白疫苗，并评价其免疫效果。方法根据GenBank上公布的HPV31、52型L2蛋白11-200位氨基酸序列，设计合成两个型别L2该区域对应的优化密码子基因融合序列，并将其克隆至原核表达载体中。利用原核表达系统表达HPV31和52L2融合蛋白，纯化目的蛋白后免疫小鼠，用血清抗体检测及HPV假病毒体外中和试验评价融合蛋白疫苗的免疫效果。结果HPV31和52L2融合蛋白在原核表达体系中呈高效表达，表达量约占全菌20％。融合蛋白加铝佐剂免疫小鼠后，血清中能检测到高滴度的总抗体及一定水平的中和抗体和交叉中和抗体。结论HPV31和52L2融合蛋白能够刺激机体产生广谱中和抗体，为高危型广谱HPVL2蛋白疫苗研发提供了实验室依据。

Objective To express HPV31 and 52 L2 fusion protein and detect its immunogenicity. Methods According to the amino acid sequences of HPV31 and 52 L2 ll-200AA published in the GenBank database, weartificially synthesized the HPV31 and 52 L2 fusion gene which was optimized according to Escherichia coli codon usage and encodes 11-200 amino acid of HPV31 and HPV52 L2, then cloned it into pET-9a vector. The HPV31 and 52 L2 fusion protein was expressed in Prokaryotic expression system and the mice were immunized with the fusion protein after purification. The immunogenicity was characterized in vaccinated mice. Results HPV31 and 52 L2 fusion protein was highly expressed in E. coli, the amount of fusion protein is nearly 20% of the total bacterial protein. The purified fusion protein with aluminum adjuvant could induce specific high titer of IgG antibodies detected by ELISA, and also induce the neutralizing antibodies against pseudovirus of HPV31 and HPV52 and cross-neutralizing antibodies against pseudovirus of HPV45,58,16,18. Conclusion HPV31 and 52 L2 fusion protein could induce neutralizing and cross-neutralizing antibodies against HPV pseudovirus. It provides laboratory basis for development of HPV L2 protein vaccine.