Analysis of exposed regions on the main extracellular domain of mouse acetylcholine receptor alpha subunit in live muscle cells by binding profiles of antipeptide antibodies.

To study the structural organization of the main extracellular domain of the nicotinic acetylcholine receptor (AChR) alpha subunit in live muscle cells, we examined the native membrane-bound receptors in cultured mouse skeletal muscle cells for their ability to bind a panel of antibodies against uniform-sized overlapping synthetic peptides which collectively represent this entire domain. The binding profile indicated that the regions alpha 23-49, alpha 78-126, alpha 146-174, and alpha 182-210 are accessible to binding with antibody. Residues alpha 23-49, alpha 78-126, and alpha 194-210 contain binding regions for alpha-neurotoxin and some myasthenia gravis autoantibodies. A comparison of this binding profile with the profile obtained for membrane-bound Torpedo californica AChR in isolated membrane fractions showed some similarities as well as significant differences between the subunit organization in the isolated membrane fraction and that in the membrane of live muscle cells. Regions alpha 89-104 and alpha 158-174, which are exposed in the isolated membrane fraction, are also exposed in the live cell. On the other hand, regions alpha 23-49, and alpha 182-210, which are exposed in the live cell, are not accessible in the isolated membrane and, furthermore, the region alpha 1-16, which has marginal accessibility in the cell, becomes highly accessible in the membrane isolates. The exposed regions defined by this study may be the primary targets for the initial autoimmune attack on the receptors in experimental autoimmune myasthenia gravis.