Mammalian muscle acetylcholine receptor purification and characterization.

Nicotinic acetylcholine receptor (AcChR) was purified from fetal calf muscle by an affinity chromatographic method utilizing alpha-neurotoxin from Naja naja siamensis as an immobilized ligand. Preparations of AcChR with an average specific activity of 5 nmol of alpha-toxin bound/mg of protein were obtained, i.e., 75% of the theoretical specific activity assuming identity with Torpedo AcChR. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified AcChR consistently showed the presence of five polypeptides, having apparent Mr's of 42 000, 44 000, 49 000, 55 000, and 58 000, respectively. The peptide of Mr 44K was demonstrated to be actin. The amino acid composition of fetal calf AcChR was shown to be similar to that of Torpedo AcChR. In addition, calf AcChR contained large amounts of amino sugars. The sedimentation coefficient of the purified calf AcChR was found to be 9.25 +/- 0.25, i.e., similar to the monomeric form of electric organ AcChR. Determination of the isoelectric point of alpha-bungarotoxin/calf AcChR complexes revealed the presence of two charged forms, having pI values of 5.16 +/- 0.13 and 6.05 +/- 0.18, respectively.