Düzgüneş N, Shavnin S A
Cancer Research Institute, University of California, San Francisco 94143-0128.
J Membr Biol. 1992 May;128(1):71-80. doi: 10.1007/BF00231872.
The fusion of lipid enveloped viruses with cellular membranes is thought to be mediated by the insertion into the target membrane of the N-terminal polypeptides of viral spike glycoproteins. Since membrane destabilization is a necessary step in membrane fusion, we investigated whether synthetic peptides with amino acid sequences corresponding to the N-termini of influenza virus hemagglutinin (HA2), vesicular stomatitis virus G-protein and Sendai virus F-protein, induce the destabilization and fusion of phospholipid vesicles. Membrane destabilization by the peptides was monitored by the release of aqueous contents of large unilamellar phospholipid vesicles. Aggregation was detected by a resonance energy transfer assay. Membrane fusion was followed by means of assays for the intermixing of phospholipids and of aqueous contents. The 17-amino acid HA2 peptide (HA2.17) destabilized phosphatidylcholine (PC) vesicles even at neutral pH, but the rate and extent of destabilization increased at lower pH. This peptide did not mediate appreciable release of contents from phosphatidylserine (PS) vesicles. HA2.17 induced neither aggregation nor fusion of PC or PS vesicles. In contrast, the 7-amino acid N-terminal peptide of G-protein (G.7) destabilized PS-containing membranes and not pure PC vesicles. Although G.7 caused aggregation of and lipid mixing between PS vesicles, it did not mediate any detectable intermixing of aqueous contents. The presence of cholesterol in PC membranes did not affect the destabilization caused by the N-terminal peptide of Sendai virus F-protein (F1.7), suggesting that cholesterol is not necessary for the effective interaction of this peptide with membranes, contrary to earlier proposals. Our results support the hypothesis that the hydrophobic N-terminal region of certain viral envelope proteins insert into and destabilize target membranes.
脂质包膜病毒与细胞膜的融合被认为是由病毒刺突糖蛋白N端多肽插入靶膜介导的。由于膜去稳定化是膜融合的必要步骤，我们研究了具有与流感病毒血凝素（HA2）、水泡性口炎病毒G蛋白和仙台病毒F蛋白N端氨基酸序列对应的合成肽是否能诱导磷脂囊泡的去稳定化和融合。通过大单层磷脂囊泡水相内容物的释放来监测肽对膜的去稳定化作用。通过共振能量转移分析检测聚集情况。通过磷脂和水相内容物混合的分析来跟踪膜融合。17个氨基酸的HA2肽（HA2.17）即使在中性pH值下也能使磷脂酰胆碱（PC）囊泡去稳定化，但在较低pH值下去稳定化的速率和程度会增加。该肽不会介导磷脂酰丝氨酸（PS）囊泡内容物的明显释放。HA2.17不会诱导PC或PS囊泡的聚集或融合。相比之下，G蛋白的7个氨基酸N端肽（G.7）能使含PS的膜去稳定化，而不是纯PC囊泡。尽管G.7导致了PS囊泡的聚集和脂质混合，但它不会介导任何可检测到的水相内容物的混合。PC膜中胆固醇的存在并不影响仙台病毒F蛋白N端肽（F1.7）引起的去稳定化，这表明与早期的观点相反，胆固醇对于该肽与膜的有效相互作用不是必需的。我们的结果支持这样一种假说，即某些病毒包膜蛋白的疏水N端区域插入并使靶膜去稳定化。
