The sites of neurotoxicity in alpha-cobratoxin.
We have chemically modified groups of amino acids in the sequence of alpha-cobratoxin and have studied the derivatives as to their affinity of binding to the acetylcholine receptor protein from Torpedo marmorata. (i) The toxin derivatives which were fully modified at lysine (penta-epsilon-N,N-dimethyl lysine; penta-epsilon-N-acetyl lysine), arginine (penta-N7,N8-(1,2-dihydroxycyclohex-1,2-ylene arginine), and tyrosine (mononitrotyrosine) all had significant remaining toxicity and affinity of binding. (ii) The "extra" disulfide of alpha-cobratoxin was selectively reduced and alkylated. Depending on the charge, size, and hydrophobicity of the attached groups, derivatives were obtained that bound to the acetylcholine receptor with higher (di-S-carboxyamidomethyl), about equal (di-S-pyridylethyl), or lower (di-iodoacetaminoethylnaphthylamine-5-sulfonic acid) affinity than the unmodified toxin. (iii) A fully reduced and carbamidomethylated derivative of alpha-cobratoxin obtained by repeating the procedure for selective reduction six times still bound with appreciable affinity (KD approximately 3 X 10(-6) M) to the acetylcholine receptor. We conclude that neither a single positively charged residue nor tyrosine nor the integrity of the disulfides is absolutely essential for toxicity. Furthermore, the single tyrosine and the area around the extra disulfide do not participate in the binding to the receptor. Together with previous findings on this interaction, this suggests a multipoint attachment of toxin and receptor involving several locally separate structural elements of the toxin.
D000215:Acylation; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000032:analysis; D003039:Cobra Neurotoxin Proteins / Q000378:metabolism* / Q000633:toxicity; D003512:Cyclohexanones / Q000494:pharmacology; D004546:Elapid Venoms / Q000378:metabolism*; D007700:Kinetics; D008745:Methylation; D051379:Mice; D010196:Pancreatic Elastase; D010446:Peptide Fragments / Q000032:analysis; D011950:Receptors, Cholinergic / Q000378:metabolism*; D013774:Tetranitromethane / Q000494:pharmacology; D014101:Torpedo; D014443:Tyrosine / Q000032:analysis
D003039:Cobra Neurotoxin Proteins; D003512:Cyclohexanones; D004546:Elapid Venoms; D010446:Peptide Fragments; D011950:Receptors, Cholinergic; D014443:Tyrosine; C025154:alpha-cobratoxin; D001120:Arginine; D010196:Pancreatic Elastase; D013774:Tetranitromethane