From Ligand Binding to Pore Opening in the Mosquito Odorant Receptor AaegOR10

Mosquito olfaction is central to host seeking and other behaviors relevant to vector ecology and disease transmission. Mosquito odorant receptors (ORs) are ligand-gated ion channels, yet how ligand binding is coupled to pore opening remains poorly understood. Here, we combine more than 400 $\mu$s of aggregate molecular dynamics simulations and adaptive sampling with Markov state models (MSMs) and electrophysiological measurements to characterize the gating dynamics of \textit{Aedes aegypti} odorant receptor 10 (AaegOR10). Interaction analysis identifies M288 as a persistent contact between $o$-cresol and S6, connecting the orthosteric pocket to a conformation-dependent hydrophobic network involving I293, F294, and F356 at the S6-S7b interface. The MSMs place the experimentally determined open and closed structures within a multistate conformational landscape and identify a principal intermediate along the dominant gating pathway. A localized S6 kink in this state is associated with weakened S6-S7b coupling and increased flexibility of the pore-lining S7b helix. Ligand binding shifts the conformational equilibrium toward open states and shortens the transition times from the closed state to both the intermediate and open states. We also identify an off-path closed state characterized by a pronounced inward displacement of S6, with phospholipid headgroup insertion observed in some sampled conformations. Mutational analysis and voltage-clamp recordings support the functional relevance of M288, the S6-S7b interface, and the kink-prone S6 region. Together, these results describe a multistate pathway linking ligand recognition to AaegOR10 pore gating.