Conformational changes in HIV-1 gp41 in the course of HIV-1 envelope glycoprotein-mediated fusion and inactivation
Conformational changes in HIV-1 gp41 in the course of HIV-1 envelope glycoprotein-mediated fusion and inactivation
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DOI：
10.1021/bi051092d
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发表时间：
2005-09-20
期刊：
影响因子：
2.9
通讯作者：
Blumenthal, R
中科院分区：
文献类型：
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作者：
Dimitrov, AS;Louis, JM;Blumenthal, R
HIV-1包膜糖蛋白介导的融合是由HIV-1 gp41 N-和c -螺旋区域的协调合并驱动的，这导致6-螺旋束的形成。这两个区域被认为是抑制HIV-1进入的肽和抗体的主要靶点。然而，控制这种抑制的参数尚未阐明。在HIV-1介导的细胞-细胞融合过程中，我们通过使用分别结合N-和c -螺旋区域的试剂，通过定量视频显微镜监测HIV-1 gp41构象状态的时间序列来解决这个问题。表达env的细胞与靶细胞在37℃的不同时间孵育，然后洗涤。被触发的gp41对NC-1单克隆抗体的反应性在引物状态下迅速增加到最大水平，但一旦形成稳定的融合连接，反应性就会下降。相比之下，与gp41的c -螺旋区结合的5-helix的反应性随着时间的推移而不断增加。肽N36（Mut(e,g)）降低了NC-1单克隆抗体结合并增强了5-螺旋结合，这与该分子促进gp41三聚体解离的观点一致。这种失活途径可能对进入抑制剂和候选疫苗的设计很重要。
HIV-1 envelope glycoprotein-mediated fusion is driven by the concerted coalescence of the HIV-1 gp41 N- and C-helical regions, which results in the formation of 6-helix bundles. These two regions are considered prime targets for peptides and antibodies that inhibit HIV-1 entry. However, the parameters that govern this inhibition have yet to be elucidated. We address this issue by monitoring the temporal sequence of conformational states of HIV-1 gp41 during the course of HIV-1-mediated cell-cell fusion by quantitative video microscopy using reagents that bind to N- and C-helical regions, respectively. Env-expressing cells were primed by incubation with target cells at different times at 37 degrees C followed by washing. The reactivity of triggered gp41 to the NC-1 monoclonal antibody, which we demonstrate here to bind to N-helical gp41 trimers, increased rapidly to a maximal level in the primed state but decreased once stable fusion junctions had formed. In contrast, reactivity with 5-helix, which binds to the C-helical region of gp41, increased continuously as a function of time following the priming. The peptide N36(Mut(e,g)) reduced NC-1 monoclonal antibody binding and enhanced 5-helix binding, consistent with the notion that this molecule promotes dissociation of gp41 trimers. This inactivation pathway may be important for the design of entry inhibitors and vaccine candidates.