Postsynaptic membranes in the electric tissue of Narcine: IV. Isolation and characterization of the nicotinic receptor protein.
The nicotinic receptor protein of the electric tissue of Narcine was purified in several different media by partial isolation of postsynaptic membranes and affinity chromatography. Protease inhibitors were found to be necessary to prevent degradation of the protein, and both EDTA and Tris buffer were used in addition to prevent intramolecular crosslinking of 44,000 and 58,000 dalton subunits by tissue factors. The intact protein was found to have a molecular weight close to 400,000, and appears to be composed of four subunits of 44,000 daltons, two to three of 48,000, one of 58,000 and one of 65,000. All the subunits are glycoproteins and their amino acid compositions show similar hydrophobicity and acidity, suggesting similar positioning in postsynaptic membranes. Crosslinking experiments showed that acetylcholine and alpha-bungarotoxin bind to the smallest subunit, and suggest the juxtaposition of at least two of these subunits, and of all four toxin molecules bound to a receptor molecule. Morphological studies of the protein in membranes and after purification indicated cylindrial molecules with central cores.
D000109:Acetylcholine / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D004492:Edetic Acid / Q000494:pharmacology; D004557:Electric Organ / Q000032:analysis*; D005399:Fishes / Q000033:anatomy & histology*; D007526:Isoelectric Point; D008970:Molecular Weight; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D011950:Receptors, Cholinergic / Q000302:isolation & purification*; D011978:Receptors, Nicotinic / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D013570:Synaptic Membranes / Q000648:ultrastructure
D000596:Amino Acids; D011950:Receptors, Cholinergic; D011978:Receptors, Nicotinic; D010664:Phenylmethylsulfonyl Fluoride; D004492:Edetic Acid; D000109:Acetylcholine