人乳头状瘤病毒6型类病毒颗粒的制备及其中和抗体的检测  

Production of human papillomavirus type 6 virus-like particles and detection of its neutralizing anti- bodies

作　　者：潘晖榕[1] 李少伟[1] 刘波[1] 朱子恒[1] 曹欢欢[1] 顾颖[1] 程通[1] 王颖彬[1] 张军[1] 夏宁邵[1] 

机构地区：[1]厦门大学国家传染病诊断试剂与疫苗工程技术研究中心生命科学学院细胞生物学与肿瘤细胞工程教育部重点实验室,361005

出　　处：《中华微生物学和免疫学杂志》2009年第1期71-76,共6页Chinese Journal of Microbiology and Immunology

基　　金：基金项目：国家工程中心技术研究建设项目（2005DC105006）;863计划重点项目（2006AA020905）;教育部新世纪优秀人才培养计划（NCET-05-0567）

摘　　要：目的利用大肠杆菌表达系统制备人乳头状瘤病毒6型（human papillomavims type6，HPV-6）类病毒颗粒（virus-like particles，VfJP）并研究其免疫原性。方法在大肠杆菌ER2566中表达HPV-6L1蛋白，并以硫酸铵沉淀、离子交换色谱、疏水相互作用色谱等手段对其进行纯化。纯化后的HPV-6L1经体外组装形成VLP后以动态光散射，透射电镜对其形态进行检测，并以假病毒中和实验评价HPV-6L1VLP在实验动物体内所诱导的抗HPV-6／11中和抗体水平。结果HPV-6L1蛋白在大肠杆菌中以可溶形式表达，经过纯化后的HPV-6L1蛋白可以在体外组装为半径25rim左右的VLP。该VLP可以在山羊及兔体内诱导高滴度的HPV-6／11中和抗体。结论大肠杆菌表达系统可以简便高效制备具有免疫原性的HPV-6VLP，可以用于HPV-6疫苗的研究。Objective To produce human papillomavirus type 6 （HPV-6）virus-like particles with Escherichia coli expression system and study its immunogenicity. Methods HPV-6 L1 gene was inserted in- to prokaryotic expression vector pTO-T7 and then expressed in Escherichia coli ER2566. The HPV-6 L1 protein was purified by ammonium sulfate precipitation, ion-exchange chromatography, and hydrophobic interac- tion chromatography. Then the purified HPV-6 L1 self-assembled into virus-like particle after removing 1,4- dithiothreitol （DTF）. The morphology of the virus-like particles was investigated with dynamic light scatter and transmission electron microscopy, and the immunogenicity was determined with in vitro pseudovirions neutralization assay. Results HPV-6 L1 was expressed in soluble form in Escherichia coll. Following the removal of DTI＇, purified HPV-6 L1 protein could assemble into virus-like particles as 25 nm in the radius. And the animal immunization test showed HPV-6 virus-like particles can elite high titer neutralizing antibod- ies. Conclusion The bacterially expressed HPV-6 L1 VLP is highly immunogenicity and easy to produce. And it can be good candidate of HPV-6 vaccine.

关 键 词：人乳头状瘤病毒6型 大肠杆菌表达系统 纯化 类病毒颗粒 免疫原性 

分 类 号：R3[医药卫生—基础医学]