title: Binding properties of SH3 peptide ligands identified from phage-displayed random peptide libraries.
abstract: Combinatorial libraries have yielded high-affinity ligands for SH3 domains of a number of different proteins. We have shown that synthetic peptides containing these SH3 ligand sequences serve as specific probes of SH3 domains. Direct binding of the N-terminal biotinylated peptide ligands was conveniently detected in ELISA, filter-blotting, and dot-blotting experiments with the use of streptavidin-conjugated enzymes. In some cases, detection of peptide-SH3 interactions required that the biotinylated peptides first were preconjugated with streptavidin to form a multivalent complex. Interestingly, these nominally tetravalent SH3 peptide ligands cross-react to varying degrees with different SH3 domains. We have used such complexes to screen lambda cDNA expression libraries and have isolated clones that encode both known and novel SH3-domain-containing proteins. Based on the success of this methodology, we propose a general strategy by which ligands of a modular domain-containing protein can be isolated from random peptide libraries and used to screen cDNA expression libraries systematically for novel modular domain-containing proteins.
authors.affiliation: Department of Biology, University of North Carolina, Chapel Hill 27599-3280, USA.
authors.forename: N G
authors.initials: NG
authors.lastname: Hoffman
authors.forename: A B
authors.initials: AB
authors.lastname: Sparks
authors.forename: J M
authors.initials: JM
authors.lastname: Carter
authors.forename: B K
authors.initials: BK
authors.lastname: Kay
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism*; D019151:Peptide Library; D011485:Protein Binding / Q000235:genetics; D018909:src Homology Domains / Q000235:genetics
chemical_list: D010446:Peptide Fragments; D019151:Peptide Library