Localization of histidine residues relevant for the binding of α-bungarotoxin to the acetylcholine receptor α-subunit in V8-proteolytic fragments

• Published in: Neurochemistry international 1996-05, Vol.28 (5), p.557-567
• Main Authors: Lacorazza, H.Daniel, López, Ricardo A., Venera, Graciela D., Biscoglio de Jiménez Bonino, Mirtha
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Biological and medical sciences; Bungarotoxins - chemistry; Cell receptors; Cell structures and functions; Electric Fish; Freshwater; Fundamental and applied biological sciences. Psychology; Histidine - analysis; Molecular and cellular biology; Molecular Sequence Data; Monoamines receptors (catecholamine, serotonine, histamine, acetylcholine); Peptide Fragments - chemistry; Receptors, Cholinergic - chemistry; Sequence Homology, Amino Acid; Torpedo californica
• ISSN: 0197-0186; 1872-9754
• DOI: 10.1016/0197-0186(95)00113-1

Histidine residues have been shown to be critical for α-BgTx binding to the acetylcholine receptor (Lacorazza et al., 1992; Bouzat et al., 1993; Lacorazza et al., 1995). Receptor subunits from Discopyge tschudii were modified with diethylpyrocarbonate (DEP). DEP treatment produces a concentration-dependent decrease of [ 125I]α-BgTx binding to the α-subunit. The neurotoxin binding capacity was fully restored by adding the nucleophile hydroxylamine. By proteolytic mapping of the α-subunit with V8-protease, we determined that the binding capacity to the fragment αV8–19 decreased 80% by DEP treatment. In addition, the [ 125I]α-BgTx binding to the same fragment decreased by 70% when the subunits were reduced and affinity-alkylated. We report the N-terminal sequence of both subunits and V8-fragments (αV8–10, αV8–13, and αV8–18), which constitute a first contribution to the knowledge of the primary structure of the Discopyge tschudii receptor. We propose that the fragment αV8–19 contains one or more of the histidine residues involved in the α-BgTx binding and probably includes the Cys α192–193 disulfide bond. Only two histidine residues are present in the extracellular sequence of Torpedo california for such fragments: His α186 and α204.