Solid-state 15N-NMR Investigations on the pH dependent Conformation of Trp-41 of the M2 Proton Channel of Influenza A Virus

The M2 proton-conductive channel of influenza A virus is a tetrameric bundle which forms an integral membrane protein that is activated after endocytosis in the endosome at low pH. For proton gating the side chains His-37 and Trp-41 are identified to play an important role. Here, we investigated these side chain conformations of a synthetic 25-residue peptide containing the M2 trans-membrane domain (M2TMD) by solid state nuclear magnetic resonance (NMR) experiments and quantum mechanics/molecular mechanics (QM/MM) simulations. The trans-membrane peptide was reconstituted as an alpha-helical tetrameric bundle in liquid crystalline lipid membranes in the form of multilamellar vesicles and/or oriented model membranes. The M2TMD was labelled with N-15(epsilon l) at Trp-41. We applied different static N-15-NMR experiments and investigated the motionally narrowed N-15 chemical shift tensors of Trp-41 for closed and opened states. The following side chain torsion angles for Trp-41 were obtained: (chi(1)=-110 degrees +/- 30 degrees, chi 2=-+130 degrees +/- 30 degrees) and (chi(1)=55 degrees +/- 30 degrees, chi(1)=-130 degrees +/- 30 degrees) in the closed and open state.