13C-13C correlation spectroscopy of membrane-associated influenza virus fusion peptide strongly supports a helix-turn-helix motif and two turn conformations.

The influenza virus fusion peptide (IFP) is the N-terminal domain of the viral hemagglutinin protein, binds to the endosomal membrane, and plays a critical role in fusion between the viral and endosomal membranes which is a primary step in infection. The IFP is also an important system for testing simulation methods for membrane-associated peptides. In detergent, the IFP forms helix-turn-helix and helix-turn-strand structures at pH 5.0 and 7.4, respectively, while simulations in membranes by different groups have yielded conflicting results with some reports of a continuous helix without a turn. In this study, (13)C-(13)C NMR correlation spectra were obtained for the membrane-associated IFP and the (13)C chemical shifts supported a helix-turn-helix motif at both pH 5.0 and 7.4 with an alternate turn conformation at pH 5.0 that was absent at pH 7.4. The alternate conformation was correlated with protonation of the side chain of Glu-11 in the turn and with greater fusion at pH 5.0. The structures are overall consistent with the hypothesis of "inverted V" membrane location of the IFP with insertion of the N-terminal region into the membrane and contact of the turn with the lipid/water interface. The positions of hydrophobic residues in the pH 5.0 structure may favor membrane insertion with resultant increased membrane perturbation and fusion rate. In addition to their functional relevance, these IFP structures are important reference data for simulations of the membrane-associated IFP which can in principle detect the full conformational distribution of the IFP.