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 transm embrane channel that binds nerve-released acetylcholine to elicit rapi d transport of small cations. Composed of two alpha subunits and one b eta, one gamma, and one delta subunit, the receptor is a cooperative p rotein containing two sites that bind agonists, curariform antagonists , and snake alpha-toxins. Until recently the two binding sites were th ought to reside entirely within each of the two alpha subunits, but af finity labeling and expression studies have demonstrated contributions by the gamma and delta subunits. Affinity labeling and mutagenesis st udies have identified residues of the alpha subunit that contribute to the binding site, but the corresponding gamma- and delta-subunit resi dues remain unknown. By making gamma-delta chimeras and following the nearly 100-fold difference in curare affinity for the two binding site s, the present work identified residues of the gamma and delta subunit s likely to be near the binding site. Two sets of binding determinants were identified in homologous positions of the gamma and delta subuni ts. The determinants lie on either side of a disulfide loop found with in the major extracellular domain of the subunits. This loop is common to all acetylcholine, gamma-aminobutyrate, and glycine receptor subun its.