Cryo-EM structure of the human α1β3γ2 GABAA receptor in a lipid bilayer

Abstract

Type A γ-aminobutyric acid receptors (GABAARs) are pentameric ligand-gated ion channels (pLGICs) and the main drivers of fast inhibitory neurotransmission in the vertebrate nervous system. Their dysfunction is implicated in a range of neurological disorders, including depression, epilepsy and schizophrenia. Amongst the numerous assemblies theoretically possible, α1β2/3γ2 GABAARs are most prevalent in the brain. The β3 subunit plays an important role in maintaining inhibitory tone and expression of this subunit alone is sufficient to rescue inhibitory synaptic transmission in a CRISPR/Cas9 derived β1-3 triple knockout. To date, efforts to generate accurate structural models for heteromeric GABAARs have been hampered by the use of engineered receptors and the presence of detergents. Significantly, some recent cryo-EM reconstructions report collapsed conformations which disagree with the prototypical pLGIC, the Torpedo nicotinic acetylcholine receptor, the large body of structural work on homologous homopentameric receptor variants, and the logic of an ion channel architecture. To address this problem, here we present a high-resolution cryo-EM structure of the full-length human α1β3γ2L, a major synaptic GABAAR isoform, functionally reconstituted in lipid nanodiscs. The receptor is bound to a positive allosteric modulator megabody and in a desensitized conformation. Unexpectedly, each GABAAR pentamer harbours two phosphatidylinositol 4,5-bisphosphate (PIP2) molecules, whose head groups occupy positively-charged pockets in the intracellular juxtamembrane regions of α1-subunits. Beyond this level, the intracellular M3-M4 loops are largely disordered, possibly because interacting post-synaptic proteins were not included. This structure illustrates the molecular principles of heteromeric GABAA receptor organization and provides the reference framework for future mechanistic investigations of GABA-ergic signaling and pharmacology.