BST-2和Vpu相互作用抑制剂筛选模型的建立  

Establishment of a High-throughput Screening Assay for Interaction Inhibitor Between BST-2 and Vpu

作　　者：庞晓静[1] 胡斯奇[1] 张悦[1] 岑山[2] 金奇[1] 郭斐[1] 

机构地区：[1]中国医学科学院北京协和医学院病原生物学研究所,北京100730 [2]中国医学科学院北京协和医学院医药生物技术研究所免疫生物室,北京100050

出　　处：《病毒学报》2012年第6期633-638,共6页Chinese Journal of Virology

摘　　要：骨髓基质细胞抗原2(Bone marrow stromal cell antigen 2,BST-2,又称Tetherin、CD317、HM1.24)是宿主先天性免疫应答的组成部分,可抑制人类免疫缺陷病毒1(Human immunodeficiency virus 1,HIV-1)的释放,HIV-1的辅助蛋白Vpu可通过跨膜区与BST-2的跨膜区产生相互作用,进而将其降解,下调其在细胞表面的数量,拮抗BST-2的抗病毒功能。本研究将海肾荧光素酶(Renilla luciferase,Rluc)与BST-2的N端连接,增强型黄色荧光蛋白(Enhanced yellow fluorescent protein,EYFP)与Vpu的C端连接,分别构建质粒RB和VE,使两种融合蛋白在细胞内共表达,产生生物发光共振能量转移(Bioluninescence resonance energy transfer,BRET)信号,进而建立稳定双表达细胞系,以BST-2和Vpu的跨膜区相互作用为靶点,应用BRET技术,建立两种蛋白相互作用抑制剂的筛选模型,以期通过BRET信号变化筛选出相互作用抑制剂,发展新型的艾滋病治疗手段。BST-2 plays an important role in host innate immune response via inhibiting the release of HIV-1.HIV-1 accessory protein Vpu can interact with BST-2 through its transmembrane domains,degrade BST-2,and decrease BST-2 that are transported to the cell surface,thus anti-virus function of BST-2 is antagonized.In our study,we constructed plasmid RB connecting Rluc to the N-termimal of BST-2,and plasmid VE connecting EYFP to the C-terminal of Vpu.The two fusion proteins were co-expressed in 293 cells,and the interaction between the two proteins was detected via BRET method.And we further established a stable 293 cell line of dual-expression.By using BRET method,and the interaction between BST-2 and Vpu transmembrane domain as the target,a high-throughput screening assay was created that was expected to seek novel interaction inhibitors.

关 键 词：生物发光共振能量转移 BST-2 VPU 筛选模型 

分 类 号：R373.9[医药卫生—病原生物学] Q78[医药卫生—基础医学]