Binding of the Grb2 SH2 domain to phosphotyrosine motifs does not change the affinity of its SH3 domains for Sos proline-rich motifs.
Phosphotyrosine peptide binding to Grb2 induces tryptophan fluorescence changes in the Src homology 2 (SH2) domain. Affinities are in the nanomolar range, the Shc peptide having the highest affinity, followed by peptides mimicking Grb2 binding sites on EGF and HGF receptors, the putative sites on insulin and IGF-1 receptors having much lower affinities. Proline-rich peptide binding to the SH3 domains induces fluorescence changes mainly in the C-terminal SH3. Affinities are in the micromolar range, the highest affinity peptides mimicking the first proline-rich motif of the Sos C-terminus. Additional residues before this PVPPPVPP motif provide a minor contribution to the binding, but the two residues after this motif are important and may contribute to specificity. The affinity of each SH3 for each proline-rich motif is too low to account for the high stability of the Grb2-Sos complex, suggesting that Grb2 recognizes other structural features in the Sos C-terminus. Binding of a phosphotyrosine peptide to the SH2 has no effect on the SH3s. Thus the binding of Grb2 to a receptor or to an associated protein phosphorylated on tyrosines is unlikely to activate the exchange factor activity of Sos through a conformational change transmitted from the SH2 to the SH3 domains.
Cussac
D
D
Institut de Pharmacologie Moléculaire et Cellulaire du CNRS, Valbonne, France.
Frech
M
M
Chardin
P
P
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001483:Base Sequence; D066246:ErbB Receptors / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000378:metabolism*; D019000:Phosphotyrosine; D011392:Proline / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D020664:Son of Sevenless Proteins; D013050:Spectrometry, Fluorescence; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D008565:Membrane Proteins; D010455:Peptides; D011506:Proteins; D011994:Recombinant Proteins; D020664:Son of Sevenless Proteins; D019000:Phosphotyrosine; D014443:Tyrosine; D011392:Proline; D066246:ErbB Receptors