The membrane-interacting abilities of three sequences representing the putative fusogenic subdomain of the Ebola virus transmembrane protein have been investigated. In the presence of calcium, the sequence EBOGE (GAAIGLAWIPYFGPAAE) efficiently fused unilamellar vesicles composed of phosphatidylcholine, phosphatidylethanolamine, cholesterol, and phosphatidylinositol (molar ratio, 2:1:1:0.5), a mixture that roughly resembles the lipid composition of the hepatocyte plasma membrane. Analysis of the lipid dependence of the process demonstrated that the fusion activity of EBOGE was promoted by phosphatidylinositol but not by other acidic phospholipids. In comparison, EBOEA (EGAAIGLAWIPYFGPAA) and EBOEE (EGAAIGLAWIPYFGPAAE) sequences, which are similar to EBOGE except that they bear the negatively charged glutamate residue at the N terminus and at both the N and C termini, respectively, induced fusion to a lesser extent. As revealed by binding experiments, the glutamate residue at the N terminus severely impaired peptide-vesicle interaction. In addition, the fusion-competent EBOGE sequence did not associate significantly with vesicles lacking phosphatidylinositol. Tryptophan fluorescence quenching by vesicles containing brominated phospholipids indicated that the EBOGE peptide penetrated to the acyl chain level only when the membranes contained phosphatidylinositol. We conclude that binding and further penetration of the Ebola virus putative fusion peptide into membranes might be governed by the nature of the N-terminal residue and by the presence of phosphatidylinositol in the target membrane. Moreover, since insertion of such a peptide leads to membrane destabilization and fusion, the present data would be compatible with the involvement of this sequence in Ebola virus fusion
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