The nicotinic acetylcholine receptor is found at neuromuscular junctions, where its function is to depolarize muscle cells at the postsynaptic membrane, triggering muscle contraction. Depolarization occurs by channel opening, induced by the binding of acetylcholine, to allow the passage of small cations. Neurotoxins inhibit the binding of acetylcholine, by binding a large portion of the acetylcholine receptor -subunit, blocking at least part of the acetylcholine binding site. The best structure for the receptor is at the low resolution level of 22 A, using a combination of electron microscopy and x-ray diffraction data. NMR spectroscopy for this large membrane bound complex is clearly beyond current technological capabilities. However, peptides corresponding to the portion of the receptor near the acetylcholine binding site have been found to bind - bungarotoxin with about the same affinity as the entire - subunit binds this neurotoxin. It is proposed here to determine the structure of these peptides bound to -bungarotoxin, using two and three-dimensional NMR spectroscopy, including heteronuclear techniques with selective isotopic labeling. This will allow for the first time the details of the interactions between a portion of the acetylcholine receptor containing the major determinant of the neurotoxin binding site, and at least part of the acetylcholine binding site to be determined. By studying complexes formed with different peptides the relative importance of specific binding interactions between - bungarotoxin and the acetylcholine receptor, will be clarified. This will contribute to the general knowledge of non-covalent interactions between proteins.
