Abstract
This report investigates the homotetrameric membrane protein structure of the S31N M2 protein from Influenza A virus in the presence of a high molar ratio of lipid. The structured regions of this protein include a single transmembrane helix and an amphipathic helix. Two structures of the S31N M2 conductance domain from Influenza A virus have been deposited in the Protein Data Bank (PDB). These structures present different symmetries about the channel main axis. We present new magic angle spinning and oriented sample solid-state NMR spectroscopic data for S31N M2 in liquid crystalline lipid bilayers using protein tetramer:lipid molar ratios ranging from 1:120 to 1:240. The data is consistent with an essentially 4-fold-symmetric structure very similar to the M2 WT structure that also has a single conformation for the four monomers, except at the His37 and Trp41 functional sites when characterized in samples with a high molar ratio of lipid. While detergent solubilization is well recognized today as a nonideal environment for small membrane proteins, here we discuss the influence of a high lipid to protein ratio for samples of the S31N M2 protein to stabilize an essentially 4-fold-symmetric conformation of the M2 membrane protein. While it is generally accepted that the chemical and physical properties of the native environment of membrane proteins needs to be reproduced judiciously to achieve the native protein structure, here we show that not only the character of the emulated membrane environment is important but also the abundance of the environment is important for achieving the native structure. This is a critical finding as a membrane protein spectroscopist's goal is always to generate a sample with the highest possible protein sensitivity while obtaining spectra of the native-like structure.