Topology of the C-terminal tail of HIV-1 gp41: differential exposure of the Kennedy epitope on cell and viral membranes.
Topology of the C-terminal tail of HIV-1 gp41: differential exposure of the Kennedy epitope on cell and viral membranes.
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DOI：
10.1371/journal.pone.0015261
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发表时间：
2010-12-07
期刊：
影响因子：
3.7
通讯作者：
Montelaro RC
中科院分区：
文献类型：
--
作者：
Steckbeck JD;Sun C;Sturgeon TJ;Montelaro RC
HIV-1 gp41包膜蛋白的C末端尾部(CTT)被认为是包膜结构和功能特性的重要决定因素，包括融合性和抗原性。虽然CTT通常被称为“胞浆内结构域”，基于膜脂双层的独有定位假设，但早期的抗原性研究和最近的生化分析已经为特定的CTT序列的表面暴露提供了可靠的证据，包括经典的gp41的“肯尼迪表位”(KE)，导致了具有多个跨膜结构域的gp41拓扑的另一种模型。目前的研究旨在通过表征细胞和病毒粒子相关环境中天然CTT序列和替换的VSV-G表位标签与参考单抗(MAb)的暴露来检验CTT拓扑结构的这些相互冲突的模型。对完整的表达Env的细胞的表面染色和FACS分析表明，KE可以被插入的VSV-G表位标签和天然KE序列的单抗结合。重要的是，VSV-G标签只有在插入KE时才是反应性的；在表达Env的细胞中没有观察到VSV-G标签插入LLP2结构域的反应性。与细胞表面表达的Env不同，免疫沉淀法和表面等离子共振谱均未观察到KE导向的单抗与完整病毒粒子表面的结合。这些数据表明与病毒粒子和细胞相关的环境物种明显不同的CTT拓扑结构，并增加了重新考虑比目前想象的更复杂的CTT拓扑结构的理由。
The C-terminal tail (CTT) of the HIV-1 gp41 envelope (Env) protein is increasingly recognized as an important determinant of Env structure and functional properties, including fusogenicity and antigenicity. While the CTT has been commonly referred to as the “intracytoplasmic domain” based on the assumption of an exclusive localization inside the membrane lipid bilayer, early antigenicity studies and recent biochemical analyses have produced a credible case for surface exposure of specific CTT sequences, including the classical “Kennedy epitope” (KE) of gp41, leading to an alternative model of gp41 topology with multiple membrane-spanning domains. The current study was designed to test these conflicting models of CTT topology by characterizing the exposure of native CTT sequences and substituted VSV-G epitope tags in cell- and virion-associated Env to reference monoclonal antibodies (MAbs). Surface staining and FACS analysis of intact, Env-expressing cells demonstrated that the KE is accessible to binding by MAbs directed to both an inserted VSV-G epitope tag and the native KE sequence. Importantly, the VSV-G tag was only reactive when inserted into the KE; no reactivity was observed in cells expressing Env with the VSV-G tag inserted into the LLP2 domain. In contrast to cell-surface expressed Env, no binding of KE-directed MAbs was observed to Env on the surface of intact virions using either immune precipitation or surface plasmon resonance spectroscopy. These data indicate apparently distinct CTT topologies for virion- and cell-associated Env species and add to the case for a reconsideration of CTT topology that is more complex than currently envisioned.
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影响因子：
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DOI：
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作者：
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