Institute of Physics, Polish Academy of Sciences, Aleja Lotnikow 32/46, 02-668 Warsaw, Poland.
Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.
Int J Mol Sci. 2021 May 18;22(10):5301. doi: 10.3390/ijms22105301.
Fusion of viral and host cell membranes is a critical step in the life cycle of enveloped viruses. In the case of influenza virus, it is mediated by subunit 2 of hemagglutinin (HA) glycoprotein whose N-terminal fragments insert into the target membrane and initiate lipid exchange. These isolated fragments, known as fusion peptides (HAfp), already possess own fusogenic activity towards liposomes. Although they have long been studied with the hope to uncover the details of HA-mediated fusion, their actual mechanism of action remains elusive. Here, we use extensive molecular dynamics simulations combined with experimental studies of three HAfp variants to fully characterize their free energy landscape and interaction with lipid bilayer. In addition to customary assumed peptides localization at lipid-water interface, we characterize membrane-spanning configurations, which turn out to be metastable for active HAfps and unstable for the fusion inactive W14A mutant. We show that, while the degree of membrane perturbation by surface peptide configurations is relatively low and does not show any mutation-related differences, the effect of deeply inserted configurations is significant and correlates with insertion depth of the N-terminal amino group which is the highest for the wild type HAfp. Finally, we demonstrate the feasibility of spontaneous peptide transition to intramembrane location and the critical role of strictly conserved tryptofan residue 14 in this process.
病毒和宿主细胞膜的融合是包膜病毒生命周期中的一个关键步骤。在流感病毒的情况下，它是由血凝素（HA）糖蛋白的亚基 2介导的，其 N 端片段插入靶膜并启动脂质交换。这些分离的片段，称为融合肽（HAfp），已经具有自身对脂质体的融合活性。尽管它们已经被长期研究，希望揭示 HA 介导融合的细节，但它们的实际作用机制仍然难以捉摸。在这里，我们使用广泛的分子动力学模拟结合三种 HAfp 变体的实验研究，来充分描述它们的自由能景观和与脂质双层的相互作用。除了通常假定的肽在脂质-水界面的定位之外，我们还描述了膜贯穿构象，对于活性 HAfp 来说，这些构象是亚稳态的，而对于融合不活跃的 W14A 突变体则是不稳定的。我们表明，虽然表面肽构象对膜的扰动程度相对较低，并且没有显示出任何与突变相关的差异，但深插入构象的影响是显著的，并且与 N 端氨基的插入深度相关，对于野生型 HAfp 来说，插入深度最高。最后，我们证明了肽自发向膜内位置转变的可行性，以及严格保守的色氨酸残基 14 在这个过程中的关键作用。
