Structure and dynamics of a multidomain ligand-gated ion channel revealed under acidic conditions

Pentameric ligand-gated ion channels are critical mediators of electrochemical signal transduction across evolution, including key targets of biophysical studies and therapeutic development. However, the intrinsically allosteric, polymodal, modular nature of gating in this protein family presents persistent challenges to biophysical characterization. The bacterial channel DeCLIC constitutes a provocative model system for structure, function, and dynamics in this family, including a modulatory N-terminal domain (NTD). Previous closed structures of DeCLIC support a rationale for its inhibition by calcium via a site of conserved relevance in other family members; however, mechanisms of gating in DeCLIC and properties of its open state have remained unclear. Here we integrated structure-function methods including cryogenic electron microscopy (cryo-EM), molecular dynamics simulations and small-angle neutron scattering under acidic conditions to characterize a previously unreported conformation of DeCLIC with a fully hydrated pore. In contrast to previous structures, the low-pH open conformation captured by cryo-EM was stable and permeable in simulations, and consistent with solution-phase behavior as measured by small-angle scattering. We further captured an alternative closed state of the channel, evidently promoted by depletion of modulatory calcium at low pH, exhibiting dynamic rearrangements in the N-terninal domain. The expanded-pore structure evidently corresponds to a functional open state of DeCLIC, while calcium-site and NTD dynamics drive channel closure, providing a detailed template for biophysical characterization of modulatory mechanisms in ligand-gated ion channels and related systems.