The stability of the intact envelope glycoproteins is a major determinant of sensitivity of HIV/SIV to peptidic fusion inhibitors.

C-peptides derived from the HIV envelope glycoprotein transmembrane subunit gp41 C-terminal heptad repeat (C-HR) region are potent HIV fusion inhibitors. These peptides interact with the gp41 N-terminal heptad repeat (N-HR) region and block the gp41 six-helix bundle formation that is required for fusion. However, the parameters that govern this inhibition have yet to be elucidated. We address this issue by comparing the ability of C34, derived from HIV-1, HIV-2 and SIV gp41, to inhibit HIV-1, HIV-2 and SIV envelope-mediated fusion and the ability of these peptides to form stable six-helix bundles with N36 peptides derived from gp41 of these three viruses. The ability to form six-helix bundles was examined by circular dichroism spectroscopy, and HIV/SIV Env-mediated membrane fusion was monitored by a dye transfer assay. HIV-1 N36 formed stable helix bundles with HIV-1, HIV-2 and SIV C34, which all inhibited HIV-1 Env-mediated fusion at IC(50)<10nM. The three C34 peptides were poor inhibitors of HIV-2 and SIV fusion (IC(50)>100nM), although HIV-2 and SIV N36 formed stable helix bundles with SIV C34. Priming experiments with sCD4 indicate that, in contrast to HIV-1, HIV-2 and SIV Env do not expose their N-HR region to SIV C34 following CD4 binding, but rapidly proceed to co-receptor engagement and six-helix bundle formation resulting in fusion. Our results suggest that several factors, including six-helix bundle stability and the ability of CD4 to destabilize the envelope glycoprotein, serve as determinants of sensitivity to entry inhibitors.