Grb2 SH3 binding to peptides from Sos: evaluation of a general model for SH3-ligand interactions.
BACKGROUND
Grb2 acts as an adaptor protein in the transduction of signals from receptor tyrosine kinases to Ras. It binds to phosphotyrosine on the cytoplasmic tail of cell-surface receptors via its central SH2 domain, and to its immediate downstream target, Sos, via two SH3 domains. The basis of the Grb2-Sos interaction is not fully understood. We previously proposed a model for SH3 domain binding specificity, based on two solution structures of the Src SH3 domain complexed with high-affinity ligands, in which the ligands are bound in a polyproline type II conformation in two distinct orientations, class I and class II. Here, we have used this model to predict the identity and orientation of Grb2 SH3 ligands in the human Sos protein.


RESULTS
Six contiguous fragments from the carboxy-terminal portion of hSos (amino acids 1000-1333), each containing a single potential SH3 binding site, were expressed in E. coli as GST fusion proteins. Four of these proteins were predicted to associate with SH3 domains. The amino-terminal Grb2 SH3 domain was shown to bind strongly to only these four fragments.


CONCLUSIONS
We have used a general model for SH3-ligand interactions to predict the nature of Grb2 SH3 interactions with the hSos protein. Comparison of the four hSos sequences that bind Grb2 revealed a preference for the PXXPXR motif, consistent with the predicted class II-type binding interaction. The interaction between Grb2 and hSos peptides is predominantly via the amino-terminal SH3 domain, although the carboxy-terminal SH3 domain may increase the overall stability of the Grb2-hSos complex.
Simon
J A
JA
Howard Hughes Medical Institute, Cambridge, MA, USA.
Schreiber
S L
SL
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001665:Binding Sites; D015153:Blotting, Western; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D051380:GRB2 Adaptor Protein; D006801:Humans; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry; D020664:Son of Sevenless Proteins; D018909:src Homology Domains / Q000235:genetics*
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D008024:Ligands; D008565:Membrane Proteins; D010446:Peptide Fragments; D010455:Peptides; D010761:Phosphorus Radioisotopes; D011506:Proteins; D011993:Recombinant Fusion Proteins; D020664:Son of Sevenless Proteins