Protonation equilibria and pore-opening structure of the dual-histidine influenza B virus M2 transmembrane proton channel from solid-state NMR

• Main Authors: Williams, Jonathan K., Shcherbakov, Alexander A., Wang, Jun, Hong, Mei
• Other Authors: Univ Arizona, Dept Pharmacol & Toxicol
• Language: en
• Published: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC 2017
• Online Access: http://hdl.handle.net/10150/626055; http://arizona.openrepository.com/arizona/handle/10150/626055

The influenza A and B viruses are the primary cause of seasonal flu epidemics. Common to both viruses is the M2 protein, a homotetrameric transmembrane proton channel that acidifies the virion after endocytosis. Although influenza A M2 (AM2) and B M2 (BM2) are functional analogs, they have little sequence homology, except for a conserved HXXXW motif, which is responsible for proton selectivity and channel gating. Importantly, BM2contains a second titratable histidine, His-27, in the tetrameric transmembrane domain that forms a reverse WXXXH motif with the gating tryptophan. To understand how His-27 affects the proton conduction property of BM2, we have used solid-state NMR to characterize the pH-dependent structure and dynamics of His-27. In cholesterol-containing lipid membranes mimicking the virus envelope, N-15 NMR spectra show that the His-27 tetrad protonates with higher pKa values than His-19, indicating that the solvent-accessible His-27 facilitates proton conduction of the channel by increasing the proton dissociation rates of His-19. AM2is inhibited by the amantadine class of antiviral drugs, whereas BM2 has no known inhibitors. Wemeasured the N-terminal interhelical separation of the BM2 channel using fluorinated Phe-5. The interhelical F-19-F-19 distances show a bimodal distribution of a short distance of 7 angstrom and a long distance of 15-20 angstrom, indicating that the phenylene rings do not block small-molecule entry into the channel pore. These results give insights into the lack of amantadine inhibition of BM2 and reveal structural diversities in this family of viral proton channels.