title: Identification of amino acid residues responsible for the alpha5 subunit binding selectivity of L-655,708, a benzodiazepine binding site ligand at the GABA(A) receptor.
abstract: L-655,708 is a ligand for the benzodiazepine site of the gamma-aminobutyric acid type A (GABA(A)) receptor that exhibits a 100-fold higher affinity for alpha5-containing receptors compared with alpha1-containing receptors. Molecular biology approaches have been used to determine which residues in the alpha5 subunit are responsible for this selectivity. Two amino acids have been identified, alpha5Thr208 and alpha5Ile215, each of which individually confer approximately 10-fold binding selectivity for the ligand and which together account for the 100-fold higher affinity of this ligand at alpha5-containing receptors. L-655,708 is a partial inverse agonist at the GABA(A) receptor which exhibited no functional selectivity between alpha1- and alpha5-containing receptors and showed no change in efficacy at receptors containing alpha1 subunits where amino acids at both of the sites had been altered to their alpha5 counterparts (alpha1Ser205-Thr,Val212-Ile). In addition to determining the binding selectivity of L-655,708, these amino acid residues also influence the binding affinities of a number of other benzodiazepine (BZ) site ligands. They are thus important elements of the BZ site of the GABA(A) receptor, and further delineate a region just N-terminal to the first transmembrane domain of the receptor alpha subunit that contributes to this binding site.
authors: Neuroscience Research Centre, Merck Sharp and Dohme Research Laboratories, Harlow, Essex, UK.
authors: M A
authors: MA
authors: Casula
authors: F A
authors: FA
authors: Bromidge
authors: G V
authors: GV
authors: Pillai
authors: P B
authors: PB
authors: Wingrove
authors: K
authors: K
authors: Martin
authors: K
authors: K
authors: Maubach
authors: G R
authors: GR
authors: Seabrook
authors: P J
authors: PJ
authors: Whiting
authors: K L
authors: KL
authors: Hadingham
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D014151:Anti-Anxiety Agents / Q000378:metabolism; D001386:Azides / Q000378:metabolism; D001569:Benzodiazepines / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D002243:Carbolines / Q000378:metabolism; D018076:DNA, Complementary; D005260:Female; D005442:Flumazenil / Q000378:metabolism; D005445:Flunitrazepam / Q000378:metabolism; D058785:GABA-A Receptor Agonists; D006801:Humans; D007093:Imidazoles / Q000378:metabolism* / Q000494:pharmacology; D007532:Isoleucine / Q000737:chemistry*; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D011724:Pyridazines / Q000378:metabolism; D011725:Pyridines / Q000378:metabolism; D011963:Receptors, GABA-A / Q000737:chemistry* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D013912:Threonine / Q000737:chemistry*; D014162:Transfection; D014982:Xenopus laevis; D000077334:Zolpidem
chemical_list: D014151:Anti-Anxiety Agents; D001386:Azides; D002243:Carbolines; D018076:DNA, Complementary; D058785:GABA-A Receptor Agonists; D007093:Imidazoles; C105490:L 655,708; D008024:Ligands; D021122:Protein Subunits; D011724:Pyridazines; D011725:Pyridines; D011963:Receptors, GABA-A; D011993:Recombinant Fusion Proteins; D007532:Isoleucine; D001569:Benzodiazepines; D013912:Threonine; D005442:Flumazenil; D005445:Flunitrazepam; C021604:CL 218872; D000077334:Zolpidem; C042957:Ro 15-4513; C036150:beta-carboline-3-carboxylic acid methyl ester