Helical Hairpin Structure of Influenza Hemagglutinin Fusion Peptide Stabilized by Charge-Dipole Interactions between the N-terminal Amino Group and the Second Helix

摘要

The fusion domain of the influenza coat protein hemagglutinin HA2, bound to dodecyl phosphocholine (DPC) micelles, was recently shown to adopt a structure consisting of two anti-parallel α-helices, packed together in an exceptionally tight hairpin configuration. Four interhelical H^α to C=O aliphatic H-bonds were identified as factors stabilizing this fold. Here, we report evidence for an additional stabilizing force: a strong charge dipole interaction between the N-terminal Gly¹ amino group and the dipole moment of helix 2. pH titration of the amino-terminal ¹⁵N resonance, using a methylene-TROSY based 3D NMR experiment, and observation of Gly^{1 13}C′ show a strongly elevated pK value of 8.8, considerably higher than expected for an N-terminal amino group in a lipophilic environment. Chemical shifts of three C-terminal carbonyl carbons of helix 2 titrate with the protonation state of Gly¹-N, indicative of a close proximity between the N-terminal amino group and the axis of helix 2, thereby providing an optimal charge-dipole stabilization of the antiparallel hairpin fold. pK values of the side chain carboxylate groups of Glu¹¹ and Asp¹⁹ are higher by about one and 0.5 unit, respectively, than commonly seen for solvent-exposed side chains in water-soluble proteins, indicative of dielectric constants of ε = ~30 (Glu11) and ε = ~60 (Asp19), which places these groups in the headgroup region of the phospholipid micelle.