A Predicted Binding Site for Cholesterol on the GABAA Receptor

Jérôme Hénin,Reza Salari,Sruthi Murlidaran,Grace Brannigan

doi:10.1016/j.bpj.2014.03.024

Abstract

Modulation of the GABA type A receptor (GABAAR) function by cholesterol and other steroids is documented at the functional level, yet its structural basis is largely unknown. Current data on structurally related modulators suggest that cholesterol binds to subunit interfaces between transmembrane domains of the GABAAR. We construct homology models of a human GABAAR based on the structure of the glutamate-gated chloride channel GluCl of Caenorhabditis elegans. The models show the possibility of previously unreported disulfide bridges linking the M1 and M3 transmembrane helices in the α and γ subunits. We discuss the biological relevance of such disulfide bridges. Using our models, we investigate cholesterol binding to intersubunit cavities of the GABAAR transmembrane domain. We find that very similar binding modes are predicted independently by three approaches: analogy with ivermectin in the GluCl crystal structure, automated docking by AutoDock, and spontaneous rebinding events in unbiased molecular dynamics simulations. Taken together, the models and atomistic simulations suggest a somewhat flexible binding mode, with several possible orientations. Finally, we explore the possibility that cholesterol promotes pore opening through a wedge mechanism.

Highlights

Rapid inhibition in the vertebrate central nervous system is mediated largely by the neurotransmitter g-aminobutyric acid (GABA) acting on the GABA type A (GABAAR), a ClÀ channel in the Cys-loop superfamily of ionotropic receptors [1,2,3]
Three homology models of the a1b1g2 GABAA receptor were built based on the recent crystal structure of the glutamate-gated chloride channel (GluCl) from C. elegans [32] (PDB code 3RHW)
We investigate the implications of potential cholesterol binding to intersubunit ivermectin/neurosteroid sites on the GABA type A receptor (GABAAR)

Summary

Introduction

Rapid inhibition in the vertebrate central nervous system is mediated largely by the neurotransmitter g-aminobutyric acid (GABA) acting on the GABA type A (GABAAR), a ClÀ channel in the Cys-loop superfamily of ionotropic receptors [1,2,3]. Dysfunctional GABAARs have been implicated in multiple human diseases, including epilepsy [4], schizophrenia [5,6,7], Alzheimer’s disease [8], and alcohol dependence [9,10]. Drugs treating these disorders, as well as multiple classes of anesthetics [11,12,13] and sedatives [14,15] target the GABAAR. The mechanisms through which both endogenous and exogenous lipophilic modulators exert their effects on the GABAAR have not been conclusively determined. No high-resolution crystal structures of the GABAAR have been published; a general understanding of Cysloop receptor structure has emerged over the past decade thanks to the publication of cryoelectron microscopy (cryoEM) images at medium resolution [20,21] obtained for a cationic homolog (the nicotinic acetylcholine receptor (nAChR)) and, more recently, high-resolution crystal structures of prokaryotic cationic members of the family, Dickeya dadantii and Gloeobacter violaceus ligand-gated ion channels (ELIC [22,23] and GLIC [24,25,26,27,28], respectively)

Methods

Results

Conclusion