Melikyan Gregory B, Egelhofer Marc, von Laer Dorothee
Department of Molecular Biophysics and Physiology, Rush University Medical Center, Chicago, Illinois 60612, USA.
J Virol. 2006 Apr;80(7):3249-58. doi: 10.1128/JVI.80.7.3249-3258.2006.
Soluble peptides derived from the C-terminal heptad repeat domain of human immunodeficiency virus type 1 (HIV-1) gp41 are potent inhibitors of HIV-1 entry and gp41-induced fusion. Target membrane-anchored variants of these peptides have been shown to retain inhibitory activity. Both soluble and membrane-anchored C peptides (MACs) are thought to block fusion by binding to the N-terminal coiled coil domain of gp41 and preventing formation of the final six-helix bundle structure. However, interactions of target MACs with gp41 must be restricted to a subset of trimers that have their hydrophobic fusion peptides inserted into the target membrane. This unique feature of MACs was used to identify the intermediate step of fusion at which gp41 engaged the target membrane. Fusion between HIV envelope-expressing effector cells and target cells was measured by fluorescence microscopy. Expression of MACs in target cells led to less than twofold reduction in the extent of fusion. However, when reaction was first arrested by adding lysolipids that disfavored membrane merger, and the lipids were subsequently removed by washing, control cells supported fusion, whereas those that expressed MACs did not. The drastically improved potency of MACs implies that, at lipid-arrested stage, gp41 bridges the viral and target cell membranes and therefore more optimally binds the membrane-anchored peptides. Experimental demonstration of this intermediate shows that, similar to fusion induced by many other viral glycoproteins, engaging the target membrane by HIV-1 gp41 permits coupling between six-helix bundle formation and membrane merger.
来源于人类免疫缺陷病毒1型（HIV-1）gp41 C末端七肽重复结构域的可溶性肽是HIV-1进入和gp41诱导融合的有效抑制剂。这些肽的靶向膜锚定变体已被证明保留抑制活性。可溶性和膜锚定C肽（MACs）都被认为通过与gp41的N末端卷曲螺旋结构域结合并阻止最终六螺旋束结构的形成来阻断融合。然而，靶向MACs与gp41的相互作用必须局限于其疏水融合肽插入靶膜的三聚体亚群。MACs的这一独特特性被用于确定gp41与靶膜结合的融合中间步骤。通过荧光显微镜测量表达HIV包膜的效应细胞与靶细胞之间的融合。在靶细胞中表达MACs导致融合程度降低不到两倍。然而，当通过添加不利于膜融合的溶血磷脂使反应首先停止，随后通过洗涤去除脂质时，对照细胞支持融合，而表达MACs的细胞则不支持。MACs显著提高的效力意味着，在脂质阻断阶段，gp41桥接病毒和靶细胞膜，因此能更优化地结合膜锚定肽。这一中间步骤的实验证明表明，与许多其他病毒糖蛋白诱导的融合类似，HIV-1 gp41与靶膜结合允许六螺旋束形成与膜融合之间的偶联。
