Solubilization of kainic acid binding sites from rat brain.
Kainic acid binding sites were solubilized from rat brain using a combination of Triton X-100 and digitonin. The highest percentage of solubilized binding sites (45%) was obtained by treating brain membranes with 1% Triton-X-100 and 0.2% digitonin in 0.5 M potassium phosphate containing 20% glycerol. The solubilized binding sites were stable and amenable to analysis by gel filtration and lectin affinity chromatography. Computer assisted analyses demonstrated that the solubilized sites displayed high- and low-affinity binding constants similar to the membrane-bound sites. Competition experiments further supported the pharmacological similarities of the solubilized and membrane-bound sites. Gel filtration chromatography of the solubilized binding site indicated that the detergent-bound complex had a Stokes radius of 82.7 A. The [3H]kainic acid binding site appears to be glycosylated based on its capability to bind to lectins. The lectin, wheatgerm agglutinin, proved to be a potentially useful tool for characterization because the solubilized binding sites were bound and eluted in relatively high yield.
D000818:Animals; D001667:Binding, Competitive; D001923:Brain Chemistry; D002462:Cell Membrane / Q000032:analysis; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002845:Chromatography; D004072:Digitonin; D007608:Kainic Acid / Q000378:metabolism; D008297:Male; D017830:Octoxynol; D011092:Polyethylene Glycols; D051381:Rats; D011919:Rats, Inbred Strains; D018092:Kainic Acid Receptors; D017981:Receptors, Neurotransmitter / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D014909:Wheat Germ Agglutinins / Q000378:metabolism
D004072:Digitonin; D007608:Kainic Acid; D017830:Octoxynol; D011092:Polyethylene Glycols; D018092:Kainic Acid Receptors; D017981:Receptors, Neurotransmitter; D014909:Wheat Germ Agglutinins