title: The structural loop II of cobrotoxin is the main binding region for nAChR and epitope in the region is conformation-dependent.
abstract: Modification of positively charged residues, Lys and Arg, in cobrotoxin revealed that Lys-27, Lys-47, Arg-28, Arg-30, Arg-33, and Arg-36 of cobrotoxin were essential for the lethality and binding activity to nicotinic acetylcholine receptor (nAChR). The antigenicity of cobrotoxin was drastically diminished when Lys-47, Arg-28, Arg-30, Arg-33, and Arg-36 were modified, while that of the Lys-27-modified derivative was not significantly changed. The CD spectra of cobrotoxin displayed similar patterns after modification of Lys-27, Lys-47, and Arg-28. These findings suggest that Lys-27, Lys-47, and Arg-28 residues may be related to the direct binding to nAChR, and that there is no involvement of Lys-27 in the antigenic determinants of cobrotoxin. Extending the modification to Arg-30, Arg-33, and Arg-36 caused progressive conformational changes of cobrotoxin and resulted in decreased binding activity to antibody and nAChR. This indicates that Arg-30, Arg-33, and Arg-36 may be of structural importance for maintaining the active conformation of cobrotoxin. These results, together with the facts that Tyr-25, Tyr-35, and Trp-29 of cobrotoxin are important in nAChR binding activity, but Trp-29 and Tyr-35 residues are not essential for the antigenicity, suggest that the structural loop II of cobrotoxin is the main binding region for nAChR and the epitope in that region is conformation-dependent.
authors.affiliation: Department of Biochemistry, Kaohsiung Medical College, Taiwan.
authors.forename: K W
authors.initials: KW
authors.lastname: Kuo
authors.forename: L S
authors.initials: LS
authors.lastname: Chang
authors.forename: C C
authors.initials: CC
authors.lastname: Chang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D002942:Circular Dichroism; D003039:Cobra Neurotoxin Proteins / Q000737:chemistry* / Q000378:metabolism*; D004557:Electric Organ / Q000378:metabolism*; D000939:Epitopes / Q000737:chemistry / Q000378:metabolism; D008239:Lysine; D008961:Models, Structural; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011863:Radioimmunoassay; D011978:Receptors, Nicotinic / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014101:Torpedo
chemical_list: D003039:Cobra Neurotoxin Proteins; D000939:Epitopes; D010446:Peptide Fragments; D011978:Receptors, Nicotinic; D001120:Arginine; D008239:Lysine