Amino acid residues within the sequence region alpha 55-74 of Torpedo nicotinic acetylcholine receptor interacting with antibodies to the main immunogenic region and with snake alpha-neurotoxins.
The sequence region 55-74 of the alpha-subunit of the acetylcholine receptor (AChR) from Torpedo californica electroplax comprises the amino-terminal end of a sequence segment--residues alpha 67-76--forming the main immunogenic region (MIR), which is most frequently recognized by anti-AChR autoantibodies in myasthenia gravis. The synthetic sequence alpha 55-74 of Torpedo AChR binds alpha-bungarotoxin (alpha BTX), suggesting that amino acid residues within this sequence region may contribute to formation of an alpha BTX binding site. Using single-residue substituted synthetic analogues of the sequence alpha 55-74 of Torpedo AChR, in which each residue was sequentially substituted by either glycine or alanine, we sought identification of the amino acids involved in interaction with alpha-neurotoxins and with three different anti-MIR monoclonal antibodies (mAbs 6, 22, and 198). Substitution of Arg55, Arg57, Trp60, Arg64, Leu65, Arg66, Trp67, or Asn68 strongly inhibited alpha-toxin binding, whereas substitutions of Ile61, Val63, Pro69, Ala70, Asp71, or Tyr72 had marginal effects. Substitutions within the region alpha 68-72 significantly diminished binding of anti-MIR mAbs, although residue preferences differed among mAbs. Further, substituting Trp60 substantially reduced binding of mAb 198, and moderately affected binding of mAb 6, and substitution of Asp62 slightly but consistently affected binding of mAbs 6 and 22.
Department of Pharmacology, University of Minnesota, Minneapolis 55455.
J L
JL
Wahlsten
J M
JM
Lindstrom
B M
BM
Conti-Tronconi
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001665:Binding Sites; D002038:Bungarotoxins / Q000378:metabolism; D003039:Cobra Neurotoxin Proteins / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000276:immunology / Q000378:metabolism*; D011978:Receptors, Nicotinic / Q000276:immunology / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014101:Torpedo
D000911:Antibodies, Monoclonal; D002038:Bungarotoxins; D003039:Cobra Neurotoxin Proteins; D010446:Peptide Fragments; D011978:Receptors, Nicotinic; C025154:alpha-cobratoxin