title: Histidine 186 of the nicotinic acetylcholine receptor alpha subunit requires the presence of the 192-193 disulfide bridge to interact with alpha-bungarotoxin.
abstract: We have previously shown that two histidine residues of the nicotinic acetylcholine receptor are relevant for alpha-bungarotoxin binding. This paper studies: (1) the interaction between alpha-bungarotoxin and the peptide alpha173-202--synthesized according to the sequence of the Torpedo californica receptor alpha subunit--and between the toxin and the same peptide containing His186 modified with ethoxyformic anhydride or substituted by Ala; (2) the influence of the presence of Cys192-Cys193 disulfide bridge on such interactions. Solid-phase and in-solution competition assays were performed: ethoxyformylation of His186 or its substitution by Ala led to a significant drop in the toxin binding capacity only for peptides containing the bridge. Circular dichroism and fourth derivate spectra of all peptides were also analyzed. Results strongly indicate the involvement of His186 in the toxin binding to those peptides with the bridge--also present in the native receptor molecules--but not to their reduced forms; on the other hand, they give further support to the already established premise that, though the bridge does not participate directly in receptor-toxin binding, its presence is relevant to define the appropriate conformation of the interaction area.
authors.affiliation: Instituto de Química y Fisicoquímica Biológicas (UBA--CONICET) ,Facultad de Farmacia y Bioquímica, Buenos Aires, Argentina.
authors.forename: F D
authors.initials: FD
authors.lastname: Testai
authors.forename: G D
authors.initials: GD
authors.lastname: Venera
authors.forename: C
authors.initials: C
authors.lastname: Pena
authors.forename: M J
authors.initials: MJ
authors.lastname: de Jimenez Bonino
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002038:Bungarotoxins / Q000493:pharmacokinetics*; D002942:Circular Dichroism; D004220:Disulfides; D004557:Electric Organ; D006639:Histidine; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D011978:Receptors, Nicotinic / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D014101:Torpedo; D064569:alpha7 Nicotinic Acetylcholine Receptor
chemical_list: D002038:Bungarotoxins; D004220:Disulfides; D010446:Peptide Fragments; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; D006639:Histidine