Structural basis for activation and potentiation in a human α5β3 GABAA receptor

Abstract

General anesthetics like etomidate mediate amnesic as well as sedative effects through different populations of gamma amino butyric acid receptors (GABAARs) in the central nervous system. The amnesic effects have largely been attributed to 5-subunit-containing receptors in the hippocampus, indicating a clear role in learning and memory for these receptors. The 5 subunit is thought to primarily coassemble with {beta}3 and, in some cases, {gamma}2 subunits, generating a variety of receptor subtypes with differential functional and pharmacological properties. However, the stoichiometry, structure, and gating mechanisms of these different subpopulations are not well understood. Here we report structures of human 5{beta}3 GABAARs with various modulators, assembled in two stoichiometries. Our cryo-EM structures, combined with electrophysiology in Xenopus oocytes, support a primary assembly of 2:3 :{beta} subunits, though a minority population of 1:4 :{beta} indicates multiple assemblies are possible. Differential glycosylation of the 5 and {beta}3 subunits enabled reconstruction of the heteromeric complex even in the absence of added fiducials. In the resting state of the receptor, Zn2+ binds to histidine residues at the M2-17' position in the {beta}3 subunit, blocking ion passage. In the activated state, GABA binding to the orthosteric site is associated with global rearrangements propagating to unbinding of the 17' Zn2+ atoms and opening of the 9' hydrophobic gate. Unlike the 1{beta}3 receptor where GABA is effectively a partial agonist, saturating GABA binding to 5{beta}3 appears to drive activation of nearly all receptors, resulting in a single desensitized state observed under the cryo-EM conditions. In agreement with this high GABA efficacy, the GABA-bound structure is virtually unaffected by further addition of the anesthetic etomidate at the {beta}- transmembrane domain interface. These structures offer new detailed models for gating and modulation of a GABAAR subtype critical to learning and memory, including prospective templates for structure-based drug discovery.