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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。
Bone marrow stromal cell antigen 2 (BST-2, also known as Tetherin, CD317, HM1.24) is part of the host’s innate immune response and inhibits human immunodeficiency virus 1 HIV-1), Vp1, an accessory protein of HIV-1, can interact with the transmembrane region of BST-2 through the transmembrane region to further degrade it, down-regulate its number on the cell surface and antagonize the resistance of BST-2 Virus function. In this study, renilla luciferase (Rluc) was ligated to the N terminus of BST-2, and enhanced yellow fluorescent protein (EYFP) was ligated to the C terminus of Vpu to construct plasmid RB and VE, The two fusion proteins were co-expressed in the cells to generate a bioluminescence resonance energy transfer (BRET) signal, and then a stable double-expression cell line was established to target the interaction between BST-2 and the transmembrane region of Vpu , The application of BRET technology to establish a screening model of two protein interaction inhibitors, in order to screen out inhibitors of interaction by BRET signal changes, the development of new types of AIDS treatment.