title: Solubilization of the benzodiazepine/gamma-aminobutyric acid receptor complex: comparison of the detergents octylglucopyranoside and 3-[(3-cholamidopropyl)-dimethylammonio]1-propanesulfonate (CHAPS).
abstract: Octylglucopyranoside (OCTG) was three times more efficient than 3-[(3-cholamidopropyl)-dimethylammonio]1-propanesulfonate (CHAPS) in solubilizing the benzodiazepine (BDZ)/gamma-aminobutyric acid (GABA) receptor complex from rat cerebellar synaptic membranes. OCTG-solubilized receptor preparations had ligand binding characteristics that were significantly different from those of the CHAPS-solubilized receptors. The inclusion of phospholipids in the solubilization media improved the binding characteristics of both soluble receptor preparations and appeared absolutely necessary for the maintenance of chloride facilitation of flunitrazepam (FNZ) binding to OCTG-solubilized receptors. FNZ and ethyl-beta-carboline-3-carboxylate bound to OCTG-solubilized preparations with equilibrium dissociation constants of 2.2 nM and 1.6 nM, respectively, and chloride (150 mM) and GABA (100 microM) + chloride facilitated the binding of FNZ by 15% and 55%, respectively; these ligand binding characteristics are similar to those of membrane-located BDZ receptors. Cartazolate, a pyrazolopyridine that facilitated the binding of FNZ to membrane-located and CHAPS-solubilized receptors, did not facilitate FNZ binding to OCTG-solubilized receptors. These results are discussed in terms of an interaction between the membrane lipid phosphatidylserine (PS) and cartazolate; PS appears to have the capacity to inhibit the effects of cartazolate on FNZ binding. Storage of the soluble receptor preparations for 24 h at 4 degrees C resulted in the loss of several characteristic BDZ receptor binding properties. Incorporation of the OCTG-solubilized receptor complex into liposomes prevented these losses but this procedure did not protect the CHAPS-solubilized receptors. We conclude that OCTG may have some advantages over CHAPS as the detergent of choice for the solubilization and reconstitution in liposomes of a functional BDZ/GABA receptor-chloride ionophore complex.
mesh_terms: D000818:Animals; D002243:Carbolines / Q000378:metabolism; D002531:Cerebellum / Q000032:analysis*; D002712:Chlorides / Q000494:pharmacology; D002793:Cholic Acids; D004355:Drug Stability; D005445:Flunitrazepam / Q000378:metabolism; D005960:Glucosides; D006027:Glycosides; D008297:Male; D010743:Phospholipids / Q000494:pharmacology; D011720:Pyrazoles / Q000494:pharmacology; D051381:Rats; D011963:Receptors, GABA-A / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D013570:Synaptic Membranes / Q000032:analysis*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
chemical_list: D002243:Carbolines; D002712:Chlorides; D002793:Cholic Acids; D005960:Glucosides; D006027:Glycosides; D010743:Phospholipids; D011720:Pyrazoles; D011963:Receptors, GABA-A; C018619:octyl-beta-D-glucoside; D005680:gamma-Aminobutyric Acid; D005445:Flunitrazepam; C024915:beta-carboline-3-carboxylic acid ethyl ester; C006315:cartazolate; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate