Immunochemical tests of acetylcholine receptor subunit models

Abstract

Acetylcholine receptors of fish electric organs and mammalian skeletal muscle comprise four structurally homologous glycoprotein subunits in the mole ratio α₂βγδ (refs 1–4). All four subunits have leader sequences and are exposed on both sides of the membrane^5,6. From amino acid sequencing, three groups^5,7,8 have predicted that each subunit has four hydrophobic α-helical transmembranous domains. Because the N-terminus of each subunit is thought to remain on the extracellular surface after cleavage of the leader sequence, this model predicts that the N- and C-termini are both on the extracellular side. An alternative model proposed by two other groups^9,10 predicts that there is, in addition, a fifth amphipathic transmembranous domain which would place the C-terminus on the cytoplasmic side. Here, using anti-subunit sera and monoclonal antibodies and their reaction with synthetic subunit peptides, we demonstrate that the C-terminus is in fact on the cytoplasmic surface. We also show that, contrary to other predictions¹¹, the most hydrophilic sequence on the extracellular domain of α-subunits is not the main immunogenic region.