HPV 58 L1改造基因的表达及其VLP产物抗血清制备  

Expression of Genetically Modified Human Papillomavirus Type 58 L1 Gene and Production of Antisera Against Resultant VLP

作　　者：范东升[1] 张婷[1] 彭清林[1] 杨丽君[1] 许于飞[1] 张然[1] 许雪梅[1] 

机构地区：[1]中国医学科学院基础医学研究所北京协和医学院基础学院生物物理与结构生物学系,北京100005

出　　处：《中国生物化学与分子生物学报》2009年第7期677-682,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家高技术研究发展计划(863计划;No.2007AA02Z475);国家自然科学基金项目(No.30772514)资助~~

摘　　要：高危型人乳头瘤病毒(human papillomavirus,HPV)慢性持续性感染是诱发宫颈癌的主要病因.体外表达的HPV主要衣壳蛋白(L1)可自组装成病毒样颗粒(virus-like particle,VLP),免疫后可诱导产生型别特异性中和抗体,有效保护机体免受同型病毒的感染,因此可望预防病毒感染及感染相关的宫颈癌等病变.HPV58是诱发我国妇女宫颈癌的主要高危型病毒之一,目前尚无针对HPV58的疫苗问世.本研究联合采用多种策略对HPV 58 L1野生型基因进行改造,获得HPV 58 L1改造基因,命名为HPV 58 mL1,用杆状病毒-昆虫细胞表达系统进行HPV 58 mL1的表达,CsCl密度梯度离心法纯化获得HPV58 mL1重组蛋白,电镜分析结果显示,重组蛋白形成直径约55 nm的VLP.皮下免疫新西兰兔和豚鼠,ELISA检测显示,免疫动物产生高滴度针对HPV 58 mL1 VLP的抗血清,免疫斑点印迹检测显示,抗血清是针对VLP表面表位的.本研究表达了均一性好的HPV 58 mL1 VLP,并获得两个种属的HPV 58 mL1 VLP抗血清,为进一步研究有效预防HPV 58感染的疫苗打下基础.Persistent infection with high-risk types of human papillomavirus has been proved as an important etiological factor for cervical cancer. HPV major capsid protein （L1） can serf-assemble into VLPs when expressed in a recombinant expression system. VLP can induce high levets of type-specific neutralizing antibodies to efficiently prevent the infection of the same genotype HPV and eventually prevent cervical cancer and other infection related diseases. HPV type 58 is one of the most common high-risk HPVs in Chinese cervical cancer patients. However, there is no effective vaccine to prevent HPV 58 infection so far. The HPV 58 L1 wt gene was modified by several strategies. The modified gene, named as HPV 58 mL1, was expressed by Baculovirus Expression System, and the recombinant protein was purified by CsC1 uhraeentrifugation. The transmission electron microscopy analysis showed that the recombinant HPV 58 mL1 protein could self-assemble into regular VLP with a diameter of about 55 nm. New Zealand white rabbit and guinea pigs were subcutaneously immunized with HPV 58 mL1 VLP. VLP ELISA showed that HPV 58 VLP could induce high titers of HPV 58 mL1 VLP specific antibodies. Dot blot assay indicated that the sera antibodies were against the surface epitopes of VLP. Taken together, HPV 58 mL1 VLP was produced by Baculovirus Expression System and antisera were obtained from two different species, which were important for further developing effective HPV 58 VLP based prophylactic vaccines.

关 键 词：人乳头瘤病毒58型 病毒样颗粒 基因改造 抗血清 

分 类 号：R373.9[医药卫生—病原生物学] Q786[医药卫生—基础医学]