title: Characterization of gamma-aminobutyric acid-benzodiazepine receptor complexes by protection against inactivation by group-specific reagents.
abstract: The chemical topography of the gamma-aminobutyric acid (GABA) and benzodiazepine (BZ) receptors was investigated in a thoroughly washed cortical membrane preparation of the rat. Chemical modification by several amino- and tyrosyl-selective reagents and the protection from it by direct and allosteric ligands of the GABA-BZ receptor complex were used to identify the residues at the binding sites. Inhibition of specific GABA binding by p-diazobenzenesulfonic acid (DSA), tetranitromethane (TNM), and N-acetylimidazole and the selective and complete protection from it by GABA and muscimol suggest the presence of a tyrosine residue at the GABAA site. TNM, like DSA, selectively decreased the number of the low-affinity GABA receptors, and this could be completely protected only by GABA concentrations that can saturate the low-affinity sites. TNM pretreatment also abolished the muscimol enhancement of [3H]diazepam binding, which suggests that the low-affinity GABA receptor sites are responsible for this enhancement. Inhibition of GABA binding by pyridoxal-5-phosphate (PLP) and the selective protection by GABA and muscimol support the presence of a lysine residue at the GABAA receptor site. Complete and selective protection from diethylpyrocarbonate (DEP) inhibition of [3H]diazepam binding by flurazepam suggests the presence of a histidine residue at the BZ site. Flurazepam selectively protected from inhibition of [3H]diazepam binding by N-bromosuccinimide and N-acetylimidazole, but not that by DSA and TNM, which does not allow a unanimous conclusion regarding the presence of tyrosine or tryptophan residues at the BZ site.
mesh_terms: D000818:Animals; D001569:Benzodiazepines / Q000378:metabolism*; D002540:Cerebral Cortex / Q000378:metabolism*; D003975:Diazepam / Q000378:metabolism; D003979:Diazonium Compounds / Q000494:pharmacology; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007202:Indicators and Reagents / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000378:metabolism*; D011963:Receptors, GABA-A; D013425:Sulfanilic Acids / Q000494:pharmacology; D013774:Tetranitromethane / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D003979:Diazonium Compounds; D007093:Imidazoles; D007202:Indicators and Reagents; D011956:Receptors, Cell Surface; D011963:Receptors, GABA-A; D013425:Sulfanilic Acids; D001569:Benzodiazepines; D005680:gamma-Aminobutyric Acid; D013774:Tetranitromethane; D004047:Diethyl Pyrocarbonate; D003975:Diazepam; C000469:diazobenzenesulfonic acid; C012160:N-acetylimidazole