Markov state models of proton- and gate-dependent activation in a pentameric ligand-gated ion channel

Ligand-gated ion channels conduct currents in response to chemical stimuli, mediating electrochemical signaling in neurons and other excitable cells. For many channels the mechanistic details of gating remain unclear, partly due to limited structural data and simulation timescales. Here, we used enhanced sampling to simulate the pH-gated channel GLIC, and construct Markov state models (MSMs) of gating transitions. Consistent with new functional recordings reported here in oocytes, our analysis revealed differential effects of protonation and mutation on free-energy wells. Clustering of closed-versus open-like states enabled estimation of open probabilities and transition rates in each condition, while higher-order clustering affirmed conformational trends in gating. Furthermore, our models uncovered state- and protonation-dependent symmetrization among subunits. This demonstrates the applicability of MSMs to map energetic and conformational transitions between ion-channel functional states, and how they correctly reproduce shifts upon activation or mutation, with implications for modeling neuronal function and developing state-selective drugs.