Sequence specificity of SHP-1 and SHP-2 Src homology 2 domains. Critical roles of residues beyond the pY+3 position.
A combinatorial phosphotyrosyl (pY) peptide library was screened to determine the amino acid preferences at the pY+4 to pY+6 positions for the four SH2 domains of protein-tyrosine phosphatases SHP-1 and SHP-2. Individual binding sequences selected from the library were resynthesized and their binding affinities and specificities to various SH2 domains were further evaluated by SPR studies, stimulation of SHP-1 and SHP-2 phosphatase activity, and in vitro pulldown assays. These studies reveal that binding of a pY peptide to the N-SH2 domain of SHP-2 is greatly enhanced by a large hydrophobic residue (Trp, Tyr, Met, or Phe) at the pY+4 and/or pY+5 positions, whereas binding to SHP-1 N-SH2 domain is enhanced by either hydrophobic or positively charged residues (Arg, Lys, or His) at these positions. Similar residues at the pY+4 to pY+6 positions are also preferred by SHP-1 and SHP-2 C-SH2 domains, although their influence on the overall binding affinities is much smaller compared with the N-SH2 domains. A structural model was generated to qualitatively interpret the contribution of the pY+4 and pY+5 residues to the overall binding affinity. Examination of pY motifs from known SHP-1 and SHP-2-binding proteins shows that many of the pY motifs contain a hydrophobic or positively charged residue(s) at the pY+4 and pY+5 positions.
Imhof
Diana
D
Department of Chemistry, Ohio State University, Columbus, Ohio 43210, USA.
Wavreille
Anne-Sophie
AS
May
Andreas
A
Zacharias
Martin
M
Tridandapani
Susheela
S
Pei
Dehua
D
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000378:metabolism*; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D017386:Sequence Homology, Amino Acid; D018909:src Homology Domains
D047908:Intracellular Signaling Peptides and Proteins; D010446:Peptide Fragments; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases