An Allosteric Cholesterol Site in Glycine Receptors Characterized Through Molecular Simulations

Glycine receptors are pentameric ligand-gated ion channels that conduct chloride ions across postsynaptic membranes to facilitate fast inhibitory neurotransmission. In addition to gating by the glycine agonist, interactions with lipids and other compounds in the surrounding membrane environment modulate their function, but molecular details of these interactions remain unclear - in particular for cholesterol. To identify such interactions, here we report on coarse-grained simulations in a model neuronal membrane for three zebrafish glycine-receptor structures, representing apparent resting, open, and desensitized states. We then converted the systems to all-atom models to examine detailed lipid interactions, and observe cholesterol bound to the receptor at an outer-leaflet intersubunit site in a state-dependent manner, indicating that it can bias receptor function. Finally, using a modified perturbation-response scanning approach, we applied short atomistic simulations to identify amino-acid translations correlated with gating conformational changes. Frequent cholesterol contacts in atomistic simulations clustered with residues identified by perturbation analysis and overlapped with mutations influencing channel function and pathology. Cholesterol binding at this site was also observed in a recently reported pig heteromeric glycine receptor. These results indicate state-dependent lipid interactions relevant to allosteric transitions of heteromeric glycine receptors, including specific amino-acid contacts applicable to biophysical modeling and pharmaceutical design.