Histidine 186 of the nicotinic acetylcholine receptor α subunit requires the presence of the 192–193 disulfide bridge to interact with α-bungarotoxin

• Published in: Neurochemistry international 2000, Vol.36 (1), p.27-33
• Main Authors: Testai, Fernando D, Venera, Graciela D, Peña, Clara, Biscoglio de Jiménez Bonino, Mirtha J
• Format: Article
• Language: English
• Subjects: a-Bungarotoxin; alpha7 Nicotinic Acetylcholine Receptor; Amino Acid Sequence; Animals; Binding Sites; Binding, Competitive; Biological and medical sciences; Bungarotoxins - pharmacokinetics; Cell receptors; Cell structures and functions; Circular Dichroism; Disulfides; Electric Organ; Fundamental and applied biological sciences. Psychology; Histidine; Kinetics; Ligand-gated ionic channels; Molecular and cellular biology; Molecular Sequence Data; Monoamines receptors (catecholamine, serotonine, histamine, acetylcholine); Nicotinic acetylcholine receptor; Nicotinic receptor histidine residues; Nicotinic receptor structure-function relationship; Peptide Fragments - chemistry; Protein Conformation; Receptors, Nicotinic - chemistry; Receptors, Nicotinic - isolation & purification; Receptors, Nicotinic - metabolism; Torpedo; α-bungarotoxin-nicotinic receptor interaction
• ISSN: 0197-0186; 1872-9754
• DOI: 10.1016/S0197-0186(99)00099-6

We have previously shown that two histidine residues of the nicotinic acetylcholine receptor are relevant for α-bungarotoxin binding. This paper studies: (1) the interaction between α-bungarotoxin and the peptide α173–202 — synthesized according to the sequence of the Torpedo californica receptor α 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.