Defining SH2 domain and PTP specificity by screening combinatorial peptide libraries.
Src homology 2 (SH2) domains mediate protein-protein interactions by recognizing short phosphotyrosyl (pY) peptide motifs in their partner proteins. Protein tyrosine phosphatases (PTPs) catalyze the dephosphorylation of pY proteins, counteracting the protein tyrosine kinases. Both types of proteins exhibit primary sequence specificity, which plays at least a partial role in dictating their physiological interacting partners or substrates. A combinatorial peptide library method has been developed to systematically assess the sequence specificity of SH2 domains and PTPs. A "one-bead-one-compound" pY peptide library is synthesized on 90-microm TentaGel beads and screened against an SH2 domain or PTP of interest for binding or catalysis. The beads that carry the tightest binding sequences against the SH2 domain or the most efficient substrates of the PTP are selected by an enzyme-linked assay and individually sequenced by a partial Edman degradation/mass spectrometry technique. The combinatorial method has been applied to determine the sequence specificity of 8 SH2 domains from Src and Csk kinases, adaptor protein Grb2, and phosphatases SHP-1, SHP-2, and SHIP1 and a prototypical PTP, PTP1B.
Wavreille
Anne-Sophie
AS
Department of Chemistry and Ohio State Biochemistry Program, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210, USA.
Garaud
Mathieu
M
Zhang
Yanyan
Y
Pei
Dehua
D
D020650:Combinatorial Chemistry Techniques / Q000379:methods*; D051380:GRB2 Adaptor Protein / Q000737:chemistry; D019151:Peptide Library; D019000:Phosphotyrosine / Q000378:metabolism; D025941:Protein Interaction Mapping / Q000379:methods; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000737:chemistry; D020539:Sequence Analysis, Protein / Q000379:methods; D013379:Substrate Specificity; D018909:src Homology Domains
D051380:GRB2 Adaptor Protein; D019151:Peptide Library; D019000:Phosphotyrosine; D016392:Proto-Oncogene Proteins pp60(c-src); D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases