Molecular dissection of subunit interfaces in the acetylcholine receptor: identification of residues that determine curare selectivity.
The acetylcholine receptor from vertebrate skeletal muscle is a transmembrane channel that binds nerve-released acetylcholine to elicit rapid transport of small cations. Composed of two alpha subunits and one beta, one gamma, and one delta subunit, the receptor is a cooperative protein containing two sites that bind agonists, curariform antagonists, and snake alpha-toxins. Until recently the two binding sites were thought to reside entirely within each of the two alpha subunits, but affinity labeling and expression studies have demonstrated contributions by the gamma and delta subunits. Affinity labeling and mutagenesis studies have identified residues of the alpha subunit that contribute to the binding site, but the corresponding gamma- and delta-subunit residues remain unknown. By making gamma-delta chimeras and following the nearly 100-fold difference in curare affinity for the two binding sites, the present work identified residues of the gamma and delta subunits likely to be near the binding site. Two sets of binding determinants were identified in homologous positions of the gamma and delta subunits. The determinants lie on either side of a disulfide loop found within the major extracellular domain of the subunits. This loop is common to all acetylcholine, gamma-aminobutyrate, and glycine receptor subunits.
Sine
S M
SM
Department of Physiology and Biophysics, Mayo Clinic, Rochester, MN 55905.
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003472:Curare / Q000494:pharmacology*; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D017354:Point Mutation; D011485:Protein Binding; D011487:Protein Conformation; D011978:Receptors, Nicotinic / Q000737:chemistry* / Q000187:drug effects* / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014403:Tubocurarine / Q000031:analogs & derivatives / Q000378:metabolism
D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D011978:Receptors, Nicotinic; D011993:Recombinant Fusion Proteins; D003472:Curare; C015836:dimethyltubocurarine; D014403:Tubocurarine