The human immunodeficiency virus type 1 gp120 V2 domain mediates gp41-independent intersubunit contacts
The human immunodeficiency virus type 1 gp120 V2 domain mediates gp41-independent intersubunit contacts
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DOI：
10.1128/jvi.74.10.4448-4455.2000
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发表时间：
2000-05-01
影响因子：
5.4
通讯作者：
Moss, B
中科院分区：
文献类型：
--
作者：
Center, RJ;Earl, PL;Moss, B
人类免疫缺陷病毒 1 型 HIV-1 的包膜蛋白在高尔基复合体中经历蛋白水解裂解，产生名为 gp120 和 gp41 的亚基，它们仍保持非共价关联。虽然 gp41 具有良好表征的寡聚结构，但独立于 gp41 的 gp120 亚基间接触的维持仍然是一个有争议的问题。使用重组牛痘病毒在哺乳动物细胞中实现 gp120 的高水平表达，并结合凝胶过滤分析，我们能够分离出 gp120 的离散寡聚形式。 gp120 的寡聚化在合成后 30 至 120 分钟之间在细胞内发生。通过沉降平衡分析明确地将寡聚体鉴定为二聚体。为了鉴定参与亚基间接触的结构域，我们表达了一系列缺乏各种结构域的 gp120 蛋白，并评估了突变对寡聚结构的影响。与野生型 gp120 相比，V1 或 V3 环的删除对单体和二聚体的相对量几乎没有影响。相反，删除全部或部分 V2 环会大大减少二聚体的形成，表明该结构域是亚基间接触形成所必需的。与此一致的是，二聚体的 V2 环比单体的 V2 环更难接触到特定的单克隆抗体。之前的研究表明，虽然 V2 环不是病毒进入的绝对必要条件，但该结构域的缺失会降低病毒对单克隆抗体或血清中和的抵抗力。我们认为 gp120 的四级结构可能通过限制保守表位的暴露而有助于抵抗中和。
The envelope protein of human immunodeficiency virus type 1 HIV-1 undergoes proteolytic cleavage in the Golgi complex to produce subunits designated gp120 and gp41, which remain noncovalently associated. While gp41 has a well-characterized oligomeric structure, the maintenance of gp41-independent gp120 intersubunit contacts remains a contentious issue. Using recombinant vaccinia virus to achieve high-level expression of gp120 in mammalian cells combined with gel filtration analysis, we were able to isolate a discrete oligomeric form of gp120. Oligomerization of gp120 occurred intracellularly between 30 and 120 min after synthesis. Analysis by sedimentation equilibrium unequivocally identified the oligomeric species as a dimer. In order to identify the domains involved in the intersubunit contact, we expressed a series of gp120 proteins lacking various domains and assessed the effects of mutation on oligomeric structure. Deletion of the V1 or V3 loops had little effect on the relative amounts of monomer and dimer in comparison to wild-type gp120. in contrast, deletion of either all or part of the V2 loop drastically reduced dimer formation, indicating that this domain is required for intersubunit contact formation. Consistent with this, the V2 loop of the dimer was less accessible than that of the monomer to a specific monoclonal antibody, Previous studies have shown that while the V2 loop is not an absolute requirement for viral entry, the absence of this domain reduces viral resistance to neutralization by monoclonal antibodies or sera. We propose that the quaternary structure of gp120 may contribute to resistance to neutralization by limiting the exposure of conserved epitopes.