影响A型流感病毒样颗粒包装及释放的关键结构蛋白研究  

Key Proteins Involving in the Assembly and Release of Influenza A Virus-Like Particles(VLPs)

作　　者：张树梅[1,2] 黄海碧[2] 颜丕熙[2] 滕巧泱[2] 闫丽萍[2] 徐大伟[2] 呼和巴特尔[1] 李泽君[2] 

机构地区：[1]内蒙古农业大学兽医学院,呼和浩特010018 [2]中国农业科学院上海兽医研究所/农业部寄生虫重点开放实验室,上海200241

出　　处：《中国农业科学》2011年第13期2823-2829,共7页Scientia Agricultura Sinica

基　　金：浦江人才计划项目(09PJ1411900);农业部动物转基因专项(2009ZX08010-022B);国家公益性行业(农业)专项经费(201003012)

摘　　要：【目的】流感病毒样颗粒在病毒学研究和新型疫苗开发方面发挥着巨大作用。本研究旨在建立一种简便的包装病毒样颗粒的方法来阐明影响流感病毒样颗粒包装及释放的关键结构蛋白。【方法】构建同时表达HA、NA和M1 3个蛋白,同时表达HA和NA两个蛋白以及只表达HA蛋白的真核表达质粒,然后转染293 T细胞,检测细胞培养上清血凝效价初步判断是否包装并释放出病毒样颗粒,并采用透射电子显微镜观察病毒样颗粒的形态。【结果】表达HA和NA或表达HA、NA和M1蛋白的真核表达质粒转染293 T细胞后,检测细胞上清血凝价均为24;而转染单独表达HA蛋白质粒的细胞上清没有血凝活性。电镜观察表明单独表达HA蛋白没有可见的病毒样颗粒,而表达上述两个或3个蛋白均可观察到病毒样颗粒。【结论】本研究采用同时表达多个结构蛋白的真核表达载体来研究病毒样颗粒包装及释放的必须蛋白,研究结果表明M1在病毒样颗粒包装及释放过程中是非必须蛋白,而NA和HA蛋白是影响流感病毒样颗粒形成及释放的关键蛋白。[Objective] Virus-like particles （VLPs） play a crucial role in research of virology and development of novel vaccines. In this study the authors focus on the major structural proteins which involves in the assembly and release of influenza A VLPs. [Method] A serial of eukaryotic expression plasmids, including the plasmid simultaneously expressing HA, NA and M1 proteins, the plasmid simultaneously expressing HA and NA proteins and the plasmid expressing HA protein of influenza virus, were developed. To determine the key proteins in assembly and release of VLPs, 293T cells were transfected with the plasmids mentioned above, and VLPs were detected by hemagglutinin test and transmission electron microscope observation. [Result] The result showed that the 293T cells simultaneously expressing HA, NA and M1 proteins, or HA and NA proteins could produce VLPs, while the 293T cells expressing only HA could not. [ Conclusion ] In this study, the eukaryotic expression plasmid which could express 3 proteins simultaneously was used to determine the key proteins in assembly and release of VLPs. The results indicated that both HA and NA were crucial in this process, while M 1 was not necessary.

关 键 词：HA NA 流感病毒样颗粒 包装 释放 

分 类 号：S852.65[农业科学—基础兽医学]