Sensitive immunoassay shows selective association of peripheral and integral membrane proteins of the inhibitory glycine receptor complex.
The inhibitory glycine receptor of mammalian spinal cord is a ligand-gated chloride channel that, on affinity purification, contains two subunits of 48-kilodalton (kD) and 58-kD molecular mass in addition to an associated 93-kD protein. Ligand-binding 48-kD subunit and 93-kD protein were quantified in the CNS of the adult rat using a newly developed dot receptor assay (detection limit less than or equal to 1 fmol/assay) which employs monoclonal antibodies specific for glycine receptor polypeptides. The 93-kD protein was found to codistribute at a fixed stoichiometry with the 48-kD subunit throughout the CNS of the rat. Moreover, the 93-kD protein cofractionated with the ligand-binding subunit on solubilization and affinity chromatography or immunoprecipitation. However, both proteins were separated on sucrose gradient centrifugation of detergent extracts of spinal cord membranes in accord with earlier observations on purified receptor. These data prove that the 93-kD polypeptide is selectively associated with the membrane core of the strychnine-sensitive glycine receptor. The regional distribution of glycine receptor polypeptides was also determined in the CNS of the spastic rat mutant. In contrast to hereditary spasticity in mouse and cattle, no reduction of glycine receptors was found in the spastic rat.
Becker
C M
CM
Zentrum für Molekulare Biologie, Universität Heidelberg, F.R.G.
Hoch
W
W
Betz
H
H
D000818:Animals; D002490:Central Nervous System / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D005591:Chemical Fractionation; D002846:Chromatography, Affinity; D000939:Epitopes; D007118:Immunoassay / Q000379:methods* / Q000592:standards; D008565:Membrane Proteins / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats; D011919:Rats, Inbred Strains; D011922:Rats, Mutant Strains; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter / Q000276:immunology / Q000378:metabolism*; D012016:Reference Values; D012680:Sensitivity and Specificity; D013035:Spasm / Q000378:metabolism; D013331:Strychnine / Q000378:metabolism; D014316:Tritium
D000939:Epitopes; D008565:Membrane Proteins; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter; D014316:Tritium; D013331:Strychnine