title: Binding kinetics of gamma-aminobutyric acidA receptor noncompetitive antagonists: trioxabicyclooctane, dithiane, and cyclodiene insecticide-induced slow transition to blocked chloride channel conformation.
abstract: Binding kinetics and affinities are determined for 25 antagonists interacting with the noncompetitive blocker site of the gamma-aminobutyric acidA receptor complex present in bovine brain membranes. Four radiolabeled noncompetitive antagonists are 4-tert-butylbicyclophosphoro[35S]thionate ([35S]TBPS), 4-tert-butylbicycloortho[3',4'-3H2]benzoate, 4'-cyano-4-sec-[3,4-3H2]butylbicycloorthobenzoate, and the new 4'-ethynyl-4-n-[2,3-3H2]propylbicycloorthobenzoate. The other 21 antagonists are unlabeled inhibitors of three chemical classes (other trioxabicyclooctane, dithiane, and cyclodiene insecticides). The radioligands bind to a single noninteracting site in the membranes, based on linear Scatchard plots and monophasic association and dissociation kinetics. The kinetics of unlabeled ligands are estimated by their effect on the [35S]TBPS association curve, using the theoretical model of Motulsky and Mahan [Mol. Pharmacol. 25:1-9 (1984)]. The receptor affinities of trioxabicyclooctanes and dithianes correlate with their association rates, whereas those of cyclodienes correlate with their dissociation rates. The low association rate constants for all ligands (< or = 3 x 10(7) M-1 min-1 at 25 degrees) are consistent with a slow transition to a blocked receptor conformation upon binding of these channel blockers. The association rate-controlled affinity for the trioxabicyclooctanes and dithianes is suggestive of an induced-fit model in which binding of the ligand initiates a conformational change in the receptor complex to the blocked state.
mesh_terms: D000818:Animals; D001643:Bridged Bicyclo Compounds / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism; D018118:Chloride Channels; D058787:GABA-A Receptor Antagonists; D007306:Insecticides / Q000737:chemistry / Q000378:metabolism*; D007473:Ion Channels / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011869:Radioligand Assay; D011963:Receptors, GABA-A / Q000737:chemistry
chemical_list: D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D018118:Chloride Channels; D058787:GABA-A Receptor Antagonists; D007306:Insecticides; D007473:Ion Channels; D008565:Membrane Proteins; D011963:Receptors, GABA-A; C037476:tert-butylbicyclophosphorothionate