Ebola viruses are filamentous and enveloped that belongs Filoviridae family. The lipid envelope contains one type of protein (Ebola GP) arranged into oligomers. The Ebola GP is responsible for both receptor binding and membrane fusion. It is proteolytically cleaved into disulfide-linked GP1 and GP2 subunits. The N-terminal domain of the glycoprotein GP2 is the fusion peptide region, which is thought to insert directly into the target membrane. The main goal of this work is to characterize the interaction between the fusion peptide and cellular membranes. To obtain the structural information on peptide-lipid bilayer interaction, we used spectroscopic techniques with living cells and in silico approaches. The interaction between Ebola fusion peptide with VERO or BHK-21 cells induced lipid mixing of cellular membranes. However, the Ebola peptide was not able to induce lipid mixing of cholesterol-depleted VERO or BHK-21 cells indicating that cholesterol is playing a crucial role in membrane fusion. To understand the importance of cholesterol, we isolated membrane microdomains, named lipid rafts. First of all, we analyzed the peptide-lipid interaction by NMR. The chemical shift of ¹H resonances suggest that the fusion peptide interact with lipid rafts. Besides, the peptide induced aggregation of the lipid rafts, indicating fusion. To further understand the interaction between Ebola fusion peptide and membranes we used in silico approaches. The data show that Ebola fusion peptide interacts with membranes because of the hydrophobicity of the carbonic skeleton of lipids, although the structure is stabilized by the polar heads of the lipids. After that, we showed the interaction between aromatic amino acid residues that probably contributed to structure stability. This work is the first indicative of direct interaction between Ebola fusion peptide and lipid rafts, a crucial information since the fusion peptide is a potential target for the development of antiviral drugs.