The α5(H105R) mutation impairs α5 selective binding properties by altered positioning of the α5 subunit in GABAA receptors containing two distinct types of α subunits

GABA(A) receptors are pentameric ligand-gated ion channels that are major mediators of fast inhibitory neurotransmission. Clinically relevant GABAA receptor subtypes are assembled from alpha 5(1-3, 5), beta 1-3 and the gamma 2 subunit. They exhibit a stoichiometry of two alpha, two beta and one gamma subunit, with two GABA binding sites located at the alpha/beta and one benzodiazepine binding site located at the alpha/gamma subunit interface. Introduction of the H105R point mutation into the alpha 5 subunit, to render alpha 5 subunit-containing receptors insensitive to the clinically important benzodiazepine site agonist diazepam, unexpectedly resulted in a reduced level of alpha 5 subunit protein in alpha 5(H105R) mice. In this study, we show that the alpha 5(H105R) mutation did not affect cell surface expression and targeting of the receptors or their assembly into macromolecular receptor complexes but resulted in a severe reduction of alpha 5-selective ligand binding. Immunoprecipitation studies suggest that the diminished alpha 5-selective binding is presumably due to a repositioning of the alpha 5(H105R) subunit in GABA(A) receptor complexes containing two different alpha subunits. These findings imply an important role of histidine 105 in determining the position of the alpha 5 subunit within the receptor complex by determining the affinity for assembly with the gamma 2 subunit.