M. Balass et al., THE ALPHA-BUNGAROTOXIN BINDING-SITE ON THE NICOTINIC ACETYLCHOLINE-RECEPTOR - ANALYSIS USING A PHAGE-EPITOPE LIBRARY, Proceedings of the National Academy of Sciences of the United Statesof America, 94(12), 1997, pp. 6054-6058
The nicotinic acetylcholine receptor (AcChoR) is a ligand-gated ion ch
annel that is activated upon binding of acetylcholine, alpha-Neurotoxi
ns, in particular alpha-bungarotoxin (alpha-BTX), bind specifically an
d with high affinity to the AcChoR and compete with binding of the nat
ural ligand, We employed a 15-mer phage-display peptide library to sel
ect epitopes reacting with alpha-BTX. Phages bearing the motif YYXSSL
as a consensus sequence were found to bind with high affinity to alpha
-BTX. The library-derived peptide (MRYYESSLKSYPD) bears amino acid seq
uence similarities to a region of the alpha-subunit of the Torpeda mus
cle AcChoR, as well as of other muscle and neuronal AcChoRs that bind
alpha-BTX. The library-derived peptide and the corresponding peptides
containing residues 187-199 of the Torpedo AcChoR alpha-subunit (WVYYT
CCPDTPYL), as well as peptides analogous to the above region in the ne
uronal AcChoR (e.g., human alpha(7); ERFYECCKEPYPD) that binds alpha-B
TX, inhibit the binding of alpha-BTX to the intact Torpedo AcChoR with
IC50 values of 10(-6) M. A synthetic peptide from a neuronal AcChoR t
hat does not bind alpha-BTX (e.g., human alpha(2); ERKYECCKEPYPD) whic
h differs by just one amino acid from the homologous peptide from the
alpha-BTX-binding protein (alpha(7))-i.e., Lys in alpha(2) and Tyr in
alpha(7)-does not inhibit the binding of alpha-BTX to Torpedo AcChoR.
These results indicate the requirement for two adjacent aromatic amino
acid residues for binding to alpha-BTX.