The human immunodeficiency virus utilizes its gp41 fusion protein to mediate virus-cell membrane fusion. The conserved disulfide loop region in the gp41 hairpin conformation reverses the protein chain, such that the N-terminal heptad repeat and the C-terminal heptad repeat regions interact to form the six-helix bundle. Hence, it is conceivable that the sequential folded N- and C-terminal parts of the loop region also interact. We show that the N- and C-terminal parts of the loop preferably form disulfide-bonded heterodimers with slow oxidation kinetics. Furthermore, when the two parts were linked to a single polypeptide to form the full-length loop, only an intramolecular disulfide-bonded loop was formed. Fluorescence studies revealed that an interaction takes place between the N- and C-terminal parts of the loop in solution, which was sustained in membranes. Functionally, only a combination of the N- and C-loop parts induced lipid mixing of model liposomes, the level of which increased 8-fold when they were connected to a single polypeptide chain. In both cases, the activity was independent of the oxidation state of the cysteines. Overall, the data (i) provide evidence of a specific interaction between the N- and C-terminal parts of the loop, which can further stabilize gp41 hairpin conformation, and (ii) suggest that the interaction between the N- and C-terminal parts of the loop is sufficient to induce lipid merging without forming a disulfide bond.
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