Viroporins comprise protein channels that are embedded in viral membranes facilitating the transport of various ions and molecules across the membrane. Such proteins are involved in different steps of the viral life cycle and are therefore promising drug targets. The global climate change fosters the spread of vector-borne viral diseases, such as West Nile and Dengue fever. Therefore, it is important to understand structure and function of these viroporins in order to develop global health strategies in the future.
M2 is a tetrameric proton channel from the influenza A virus and is so far the only viroporin which has been the target of two licensed drugs that block effectively the wild-type of M2. Due to its simple structure, only one α-helix spans the membrane, it comprises an ideal model system for viroporins in general and proton channels in particular.
Despite its simplistic architecture, the M2 channel plays a crucial role in the infection process of the influenza virus. Upon entry into an infected cell via endocytosis, the virus is exposed to a highly acidic environment, activating the M2 channel and allowing protons to enter the viral capsid.
In the present study, three UniSysCat PIs - A. Lange, H. Sun and T. Utesch from the FMP, joined with the Kozuch group from the FU Berlin to characterize the conducting domain of the tetrameric M2 channel, a construct comprising the amino acids 18–60, by proton-detected solid-state NMR under native-like conditions in lipid bilayers at different pH values relevant to the virus life cycle. Thereby, different conformations were detected: the nonconducting, closed state at pH 7.8, the opening at pH 6.0 and at pH 4.5 the conducting, fully open state. In the closed state at pH 7.8, the researchers detected two sets of resonances from the functionally important histidine side chain (H37), which were assigned with means of quantum mechanics/molecular mechanics (QM/MM) simulations to hydrogen-bonded and free H37 side chains that occur in varying ratios in the tetrameric arrangement. Further, also some backbone signals appear twice, which suggests conformational heterogeneity. The structural arrangement appears rather rigid, explaining the nonconducting nature of the channel. By lowering the pH to 6.0 the dynamics of the side chains increases, as derived from their disappearance in cross polarization based solid-state NMR spectra. The dynamic arrangement results from the additional protonation of the four H37 side chains and fosters an efficient transport of protons through the channel. Finally, at pH 4.5, the conformational heterogeneity observed at higher pH values disappears completely, and a unique set of highly resolved resonances becomes visible. This suggests a well-defined, acid-activated state of the M2 channel. Notably, in this state, the signals of the His37 side chains disappeared due to dynamics, as well as the signals of the amphipathic helix (residues 45–52). In summary, this study provides strong evidence to a model of proton conduction through M2 that relies on dynamic H37 side chains, paving the way for an atomic structure of the acid-activated state of M2.
Structural Transition from Closed to Open for the Influenza A M2 Proton Channel as Observed by Proton-Detected Solid-State NMR
Swantje Mohr, Caspar Schattenberg, Tillmann Utesch, Henry Sawczyc, Veniamin Chevelkov, Sascha Lange, Jacek Kozuch, Han Sun, and Adam Lange
Journal of the American Chemical Society 2025 147 (31), 27537-27551
DOI: 10.1021/jacs.5c05111