Two neighboring residues of loop A of the alpha1 subunit point towards the benzodiazepine binding site of GABAA receptors.

Institute of Biochemistry and Molecular Medicine, University of Bern, Bühlstrasse 28, CH-3012 Bern, Switzerland.
FEBS Letters (Impact Factor: 3.58). 11/2007; 581(24):4718-22. DOI:10.1016/j.febslet.2007.08.068
Source: PubMed

ABSTRACT Benzodiazepines are widely used drugs exerting sedative, anxiolytic, muscle relaxant, and anticonvulsant effects by acting through specific high affinity binding sites on some GABA(A) receptors. It is important to understand how these ligands are positioned in this binding site. We are especially interested here in the conformation of loop A of the alpha(1)beta(2)gamma(2) GABA(A) receptor containing a key residue for the interaction of benzodiazepines: alpha(1)H101. We describe a direct interaction of alpha(1)N102 with a diazepam- and an imidazobenzodiazepine-derivative. Our observations help to better understand the conformation of this region of the benzodiazepine pocket in GABA(A) receptor.

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    ABSTRACT: Benzodiazepines (BZDs) exert their therapeutic actions by binding to the GABA(A) receptor (GABA(A)R) and allosterically modulating GABA-induced chloride currents (I(GABA)). A variety of ligands with divergent structures bind to the BZD site, and the structural mechanisms that couple their binding to potentiation of I(GABA) are not well understood. In this study, we measured the effects of individually mutating 22 residues throughout the BZD binding pocket on the abilities of eszopiclone, zolpidem, and flurazepam to potentiate I(GABA). Wild-type and mutant α(1)β(2)γ(2) GABA(A)Rs were expressed in Xenopus laevis oocytes and analyzed using a two-electrode voltage clamp. GABA EC(50), BZD EC(50), and BZD maximal potentiation were measured. These data, combined with previous radioligand binding data describing the mutations' effects on BZD apparent binding affinities (J Neurosci 28:3490-3499, 2008; J Med Chem 51:7243-7252, 2008), were used to distinguish residues within the BZD pocket that contribute to BZD efficacy and BZD binding. We identified six residues whose mutation altered BZD maximal potentiation of I(GABA) (BZD efficacy) without altering BZD binding apparent affinity, three residues whose mutation altered binding but had no effect on BZD efficacy, and four residues whose mutation affected both binding and efficacy. Moreover, depending on the BZD ligand, the effects of some mutations were different, indicating that the structural mechanisms underlying the ability of BZD ligands with divergent structures to potentiate I(GABA) are distinct.
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