TOXINS SELECTIVE FOR SUBUNIT INTERFACES AS PROBES OF NICOTINIC ACETYLCHOLINE-RECEPTOR STRUCTURE

The pentameric structure of the nicotinic acetylcholine receptor with two of the five subunit interfaces serving as ligand binding sites off ers an opportunity to distinguish features on the surfaces of the subu nits and their ligand specificity characteristics. This problem has be en approached through the study of assembly of subunits and binding ch aracteristics of selective peptide toxins. The receptor, with its circ ular order of homologous subunits (alpha gamma alpha delta beta), asse mbles in only one arrangement, and through mutagenesis, the residues g overning assembly can be ascertained. Selectivity of certain toxins is sufficient to readily distinguish between sites at the alpha gamma an d alpha delta interfaces. By interchanging residues on the gamma and d elta subunits, and ascertaining how they interact with the alpha-subun it, determinants forming the binding sites can be delineated. The alph a-conotoxins, which contain two disulfide loops and 12-14 amino acids, show a 10 000-fold preference for the alpha delta over the alpha gamm a subunit interface with alpha epsilon falling between the two. The wa glerins, as 22-24 amino acid peptides with a single core disulfide loo p, show a 2000-fold preference for alpha epsilon over the alpha gamma and alpha delta interfaces. Finally, the 6700 Da short alpha-neurotoxi n from N. mossambica mossambica shows a 10 000-fold preference for the alpha gamma and alpha delta interfaces over alpha epsilon. Selective mutagenesis enables one to also distinguish alpha-neurotoxin binding a t the alpha gamma and alpha delta subunits. This information, when cou pled with homology modeling of domains and site-directed residue modif ication, reveals important elements of receptor structure and conforma tion. ((C) Elsevier, Paris).