Purification and characterization of nicotinic acetylcholine receptors from muscle.
The nicotinic acetylcholine receptor was purified from normal and denervated rat skeletal muscle. The purification protocol included alpha-cobratoxin biospecific adsorption, ion exchange chromatography, and gel filtration steps. The highest specific activity achieved was 7.5 pmol of 125I-alpha-bungarotoxin binding sites per microgram protein. Sodium dodecyl sulfate gel electrophoresis of purified AChR revealed subunits with molecular weights of 42,000 and 66,000 daltons and a minor component with a molecular weight of 52,000 daltons. Normal muscle AChR is comprised of one toxin binding component. Upon denervation a second component appears, but both components are increased as a consequence of denervation. A dissociation constant of 1.5 x 10(-8)M was determined for d-tubocurarine from receptor from both normal and denervated muscle. A dissociation constant of 1 x 10(-7)M for acetylcholine, perhaps analogous to the high affinity acetylcholine binding observed in electric fish receptor, was determined.
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002038:Bungarotoxins / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D003039:Cobra Neurotoxin Proteins / Q000378:metabolism; D003472:Curare / Q000494:pharmacology; D006614:Hindlimb; D008970:Molecular Weight; D009121:Muscle Denervation; D009132:Muscles / Q000032:analysis*; D051381:Rats; D011950:Receptors, Cholinergic / Q000302:isolation & purification*; D011978:Receptors, Nicotinic / Q000302:isolation & purification* / Q000378:metabolism
D002038:Bungarotoxins; D003039:Cobra Neurotoxin Proteins; D011950:Receptors, Cholinergic; D011978:Receptors, Nicotinic; C025154:alpha-cobratoxin; D003472:Curare; D000109:Acetylcholine