AFFINITY, SPECIFICITY, AND KINETICS OF THE INTERACTION OF THE SHC PHOSPHOTYROSINE BINDING DOMAIN WITH ASPARAGINE-X-X-PHOSPHOTYROSINE MOTIFSOF GROWTH-FACTOR RECEPTORS

The phosphotyrosine binding (PTB) domain specifically binds to tyrosin e-phosphorylated proteins, but differs in structure and mechanism of a ction from the SH2 domain family, We quantitated the affinity, specifi city, and kinetics of the interaction of the SHC PTB domain with a seq uence motif, asparagine-X-X-phosphotyrosine (NXX(pY)), found in severa l receptor tyrosine kinases and oncogenic proteins. PTB domain-mediate d interaction with the NXX(pY) motif of c-ErbB2 was characterized by s imilar overall affinity but slower kinetics than that reported for SH2 domains, This suggested that unlike SH2 domains, PTB domains may not rapidly exchange among associated proteins. Furthermore, when directly and quantitatively compared, PTB domain binding specificity did not s ignificantly overlap with a panel of seven SH2 domains, Thus, signalin g pathways involving PTB and SH2 domain-mediated interactions can be r egulated separately. Finally, our data define the minimal SHC PTB doma in binding motif as NXX(pY), not NPX(pY) as suggested by other authors , and suggest a high affinity motif, hydrophobic residue-(D/E)-N-X-X-p Y-(W/F), found in the Trk and ErbB receptor tyrosine kinase families. We conclude that PTB domains mediate specific protein-protein interact ions independent from those mediated by SH2 domains.