SPECIFICITY OF THE PTB DOMAIN OF SHC FOR BETA-TURN-FORMING PENTAPEPTIDE MOTIFS AMINOTERMINAL TO PHOSPHOTYROSINE

She phosphorylation in cells following growth factor, insulin, cytokin e, and lymphocyte receptor activation leads to its association with Gr b2 and activation of Ras, In addition to being a cytoplasmic substrate of tyrosine kinases, She contains an SH2 domain and a non-SH2 phospho tyrosine binding (PTB) domain, Here we show that the She PTB domain, b ut not the SH2 domain, binds with high affinity (ID50 congruent to 1 m u M) to phosphopeptides corresponding to the sequence surrounding Tyr( 250) Of the polyoma virus middle T (mT) antigen (LLSNPTpYS-VMRSK), Tru ncation studies show that five residues amino terminal to tyrosine are required for high affinity binding, whereas all residues carboxyl-ter minal to tyrosine can be deleted without loss of affinity, Substitutio n studies show that tyrosine phosphorylation is required and residues at -5, -3, -2, and -1 positions relative to pTyr are important for thi s interaction, H-1 NMR studies demonstrate that the phosphorylated mT antigen-derived sequence forms a stable beta turn in solution, and cor relations between structure and function indicate that the beta turn i s important for PTB domain recognition. These results show that PTB do mains are functionally distinct from SH2 domains, Whereas SH2 domain b inding specificity derives from peptide sequences carboxyl-terminal to phosphotyrosine, the She PTB domain gains specificity by interacting with beta turn-forming sequences amino-terminal to phosphotyrosine.