We have investigated membrane interactions and perturbations induced by NH₂-DKWASLWNWFNITNWLWYIK-COOH (HIV_c), representing the membrane interface-partitioning region that precedes the transmembrane anchor of the human immunodeficiency virus type-1 gp41 fusion protein. The HIV_c peptide bound with high affinity to electrically neutral vesicles composed of dioleoylphosphatidylcholine, dioleoylphosphatidylethanolamine and cholesterol (molar ratio, 1:1:1), and induced vesicle leakage and lipid mixing. Infrared spectra suggest that these effects were promoted by membrane-associated peptides adopting an α-helical conformation. A sequence representing a defective gp41 phenotype unable to mediate both cell–cell fusion and virus entry, was equally unable to induce vesicle fusion, and adopted a non-helical conformation in the membrane. We conclude that membrane perturbation and adoption of the α-helical conformation by this gp41 region might be functionally meaningful.