Membrane curvature sensing and symmetry breaking of the M2 proton channel from Influenza A

The M2 proton channel aids in the exit of mature influenza viral particles from the host plasma membrane through its ability to stabilize regions of high negative gaussian curvature (NGC) that occur at the neck of budding virions. The channels are homo-tetramers that contain a cytoplasm-facing amphipathic helix (AH) that is necessary and sufficient for NGC generation; however, constructs containing the transmembrane spanning helix, which facilitates tetramerization, exhibit enhanced curvature generation. Here we used all-atom molecular dynamics (MD) simulations to explore the conformational dynamics of M2 channels in lipid bilayers revealing that the AH is dynamic, quickly breaking the 4-fold symmetry observed in most structures. Next, we carried out MD simulations with the protein restrained in 4-fold and 2-fold symmetric conformations to determine the impact on the membrane shape. While each pattern was distinct, all configurations induced pronounced curvature in the outer leaflet with rather subtle lipid tilt, while conversely, the inner leaflets adjacent to the AHs showed minimal curvature and significant lipid tilt. The MD-generated profiles at the protein-membrane interface were then extracted and used as boundary conditions in a continuum elastic membrane model to calculate the membrane bending energy of each conformation embedded in different membrane surfaces characteristic of a budding virus. The calculations show that all three M2 conformations are stabilized in concave spherical caps and destabilized in convex spherical caps, the latter reminiscent of a budding virus. Only C2-broken symmetry conformations are stabilized in NGC surfaces, by 1-3 kBT depending on the AH domain arrangement. The most favored conformation is stabilized in saddles with curvatures corresponding to 33 nm radii. In total, our work provides atomistic insight into the curvature sensing capabilities of M2 channels and how enrichment in the nascent viral particle depends on protein shape and membrane geometry.