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Mimicking the nicotinic receptor binding site by a single chain Fv selected by competitive panning from a synthetic phage library
Author(s) -
Bracci Luisa,
Pini Alessandro,
Lozzi Luisa,
Lelli Barbara,
Battestin Paola,
Spreafico Adriano,
Bernini Andrea,
Niccolai Neri,
Neri Paolo
Publication year - 2001
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2001.00398.x
Subject(s) - phage display , receptor , ligand (biochemistry) , bungarotoxin , nicotinic agonist , biopanning , nicotinic acetylcholine receptor , peptide library , acetylcholine receptor , binding site , epitope , chemistry , biology , biochemistry , microbiology and biotechnology , antibody , peptide , peptide sequence , immunology , gene
We have developed a novel competitive method to select from a phage display library a single chain Fv which is able to mimic the α‐bungarotoxin binding site of the muscle nicotinic receptor. The single chain Fv was selected from a large synthetic library using α‐bungarotoxin‐coated magnetic beads. Toxin‐bound phages were then eluted by competition with affinity purified nicotinic receptor. Recognition of the toxin by the anti‐α‐bungarotoxin single chain Fv was very similar to that of the receptor, such as indicated by the epitope mapping of α‐bungarotoxin through overlapping synthetic peptides. Moreover, several positively charged residues located in the toxin second loop and in the C‐terminal region were found to be critical, to a similar extent, for toxin recognition by the single chain Fv and the receptor. However, although the anti‐α‐bungarotoxin single chain Fv seems to mimic the toxin binding site of the nicotinic receptor, it does not bind other nicotinic agonists or antagonists. Our results suggest that competitive selection of anti‐ligand antibody phages can allow the production of receptor‐mimicking molecules directly and exclusively targeted at one specific ligand. Since physiologically and pharmacologically different ligands can produce opposite effects on receptor functions, such selective ligand decoys can have important therapeutic applications.