Type I and type II gamma-aminobutyric acid/benzodiazepine receptors: purification and analysis of novel receptor complex from neonatal cortex.
The gamma-aminobutyric acid (GABA) type A receptor was purified several thousandfold by affinity chromatography from rat cerebellum, adult cortex, and neonatal cortex. Competition for the benzodiazepine binding site by CL 218872 indicated that cerebellar receptors were predominantly type I, adult cortical receptors were a mixture of subtypes, and neonatal cortex was enriched in type II receptor. The receptor purified from neonatal cortex contained predominantly a 54-kilodalton (kDa), beta-subunit-like protein, whereas receptors from cerebellum and adult cortex contained nearly equal amounts of a 50-kDa, alpha-subunit-like protein and a 54-kDa polypeptide. Peptide maps of trypsin-digested 54-kDa subunits from cerebellum, adult cortex, and neonatal cortex exhibited very similar profiles, a result indicating considerable homology between these proteins in the receptor subtypes. A 59-kDa subunit protein was detected in the receptor complex purified from neonatal cortex. Like the 50-kDa, alpha-subunit of the type I receptor, this protein was photolabeled with [3H]flunitrazepam. The photolabeled peptide fragments, produced by trypsin digestion of these alpha 50- and alpha 59-subunits, exhibited the same retention times on reverse-phase HPLC. A less highly purified GABAA receptor preparation from adult rat spinal cord possessed characteristics that were very similar to those of the receptors purified from neonatal cortex.
Sato
T N
TN
Department of Biology, Georgetown University, Washington, D.C. 20057.
Neale
J H
JH
D000818:Animals; D000831:Animals, Newborn; D002540:Cerebral Cortex / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005445:Flunitrazepam / Q000378:metabolism; D009118:Muscimol / Q000378:metabolism; D009994:Osmolar Concentration; D010449:Peptide Mapping; D051381:Rats; D011963:Receptors, GABA-A / Q000032:analysis*; D013116:Spinal Cord / Q000378:metabolism; D014316:Tritium
D011963:Receptors, GABA-A; D014316:Tritium; D009118:Muscimol; D005445:Flunitrazepam