Identification of epitopes within a highly immunogenic region of acetylcholine receptor by a phage epitope library.
We have employed a hexapeptide phage-epitope library to identify epitopes for a mAb (mAb 5.14), which is directed to a determinant within a highly immunogenic, cytoplasmic region of the alpha-subunit of acetylcholine receptor (AChR). We have selected two different peptide-presenting phages (SWDDIR-phage and LWILTR-phage) which interact specifically with mAb 5.14. This interaction is specifically inhibited by AChR and by synthetic peptides corresponding to the hexapeptides presented by the selected phages. Although mAb 5.14 binds to AChR in its native as well as its denatured form, the selected hexapeptides do not exist as such in the AChR molecule. However, three amino acid sequence homologies with these hexapeptides were shown to be present in the cytoplasmic region of Torpedo AChR. By extending the selected hexapeptides, at one or both ends, with amino acid residues flanking the hexapeptides in the phage, we obtained mimotopes with an up to two order of magnitude higher affinity to the Ab. These extended peptides were able to efficiently block the binding of mAb 5.14 to both peptide-presenting phages, and to AChR.
Barchan
D
D
Department of Chemical Immunology, Weizmann Institute of Science, Rehovot, Israel.
Balass
M
M
Souroujon
M C
MC
Katchalski-Katzir
E
E
Fuchs
S
S
D000595:Amino Acid Sequence; D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000276:immunology; D017951:Antigen Presentation; D001435:Bacteriophages / Q000235:genetics; D001667:Binding, Competitive / Q000276:immunology; D015153:Blotting, Western; D015723:Gene Library; D016056:Immunodominant Epitopes / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000276:immunology; D011950:Receptors, Cholinergic / Q000276:immunology*; D014101:Torpedo
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D016056:Immunodominant Epitopes; D009842:Oligopeptides; D011950:Receptors, Cholinergic