猪脑心肌炎病毒2B蛋白与RACK1蛋白相互作用的鉴定  

Identification of the interaction between 2B protein of encephalomyocarditis virus and RACK1

作　　者：王岩[1,2] 黄丽[2] 崔尚金[2] 李江南[2] 杨春梅[2] 于会彬[2] 郭东伟[2] 张元峰 夏雪山[1] 翁长江[2] 

机构地区：[1]昆明理工大学生命科学与技术学院,云南昆明650500 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/动物病原监测与流行病学研究创新团队,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2014年第5期339-342,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(31300139);中国农业科学院基本科研业务费预算增量项目(2013ZL034)

摘　　要：为筛选与猪脑心肌炎病毒(EMCV)2B蛋白相互作用的宿主蛋白,本实验利用酵母双杂交方法筛选BHK-21细胞表达文库制备与EMCV 2B蛋白相互作用的宿主蛋白RACK1蛋白。酵母共转化试验和免疫共沉淀试验表明两者可以特异性结合,激光共聚焦试验表明两者共定位于细胞质中。为定位RACK1与2B蛋白相互作用区域,本研究构建了RACK1截短表达重组质粒,并将其与pCAGGS-Flag-2B共转染HEK293细胞,利用免疫共沉淀试验验证其相互作用区域。结果显示2B蛋白与宿主细胞RACK1蛋白存在相互作用,并且证明相互作用区域位于RACK1蛋白的N端。To screen the host proteins interacted with the 2B protein of encephalomyocarditis virus （EMCV）, we scanned the BHK-21 expression library using EMCV 2B protein as bait protein by yeast-two-hybrid, and found a potential interacting partner protein of RACK1. The interaction of the 2B and RACK1 proteins was further confirmed by co-transformation in the yeast and Co-IP assay in co-transfected HEK293 cells. In addition, they were co-localized in the cytoplasma of co-transfected HEK293 cells examined by laser confocal assays. To map the interacting domain of RACK1, a series of RACK1 truncated eukaryotic expression recombinant plasmids were construct and co-transfected with pCAGGS-Flag-2B into HEK293 cells, respectively, which were subjected to Co-IP and detected by western blot. The results suggested that EMCV 2B interacted with N terminal of RACK1.

关 键 词：猪脑心肌炎病毒 酵母双杂交 2B蛋白 RACK1蛋白 相互作用 

分 类 号：S852.65[农业科学—基础兽医学]