Orientation of Peptide Fragments from Sos Proteins Bound to the N-Terminal SH3 Domain of Grb2 Determined by NMR Spectroscopy

Abstract

NMR spectroscopy has been used to characterize the protein-protein interactions between the mouse Grb2 (mGrb2) N-terminal SH3 domain complexed with a 15-residue peptide (SPLLPKLPP-KTYKRE) corresponding to residues 1264-1278 of the mouse Sos-2 (mSos-2) protein. Intermolecular interactions between the peptide and 13C-15N-labeled SH3 domain were identified in half-reverse-filtered 2D and 3D NOESY experiments. Assignments for the protons involved in interactions between the peptide and the SH3 domain were confirmed in a series of NOESY experiments using a set of peptides in which different leucine positions were fully deuterated. The peptide ligand-binding site of the mGrb2 N-terminal SH3 domain is defined by the side chains of specific aromatic residues (Tyr7, Phe9, Trp36, Tyr52) that form two hydrophobic subsites contacting the side chains of the peptide Leu4 and Leu7 residues. An adjacent negatively charged subsite on the SH3 surface is likely to interact with the side chain of a basic residue at peptide position 10 that we show to be involved in binding. The peptide-binding site of the SH3 is characterized by large perturbations of amide chemical shifts when the peptide is added to the SH3 domain. The mGrb2 N-terminal SH3 domain structure in the complex is well-defined (backbone RMSD of 0.56 +/- 0.21 calculated over the backbone N, C alpha, and C atoms of residues 1-54). The structure of the peptide in the complex is less well-defined but displays a distinct orientation.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of Interfacial Water Molecules in Proline-rich Ligand Recognition by the Src Homology 3 Domain of Abl

“…Despite the highly hydrophobic character of the ligand, this interaction is characterized by an extremely negative binding enthalpy (Ϫ92 kJ⅐mol Ϫ1 ) (12), which is notably bigger than the values reported for other SH3 complexes that typically range between Ϫ20 and Ϫ50 kJ⅐mol Ϫ1 (13)(14)(15)(16)(17). In previous work, we identified a set of fully buried water molecules at the binding interface that mediate the interactions between the peptide ligand and the SH3 domain and postulated that these water-mediated hydrogen bonds, which generate an extended and considerably more polar binding interface, are key for understanding the thermodynamic behavior of the system (12).…”

“…Nonetheless, all thermodynamic studies of SH3 ligand binding reported to date have surprisingly revealed an invariantly negative binding enthalpy that is partially compensated by unfavorable entropic contributions (12)(13)(14)(15)(16)(17). This thermodynamic behavior, which cannot be rationalized exclusively in terms of direct interactions between hydrophobic surfaces, reveals an underlying complexity in the recognition of proline-rich ligands by SH3 domains.…”

Disabled-2 inactivation is an early step in ovarian tumorigenicity

“…The domain structure suggests that Dab2 is an adaptor protein involved in protein-protein interaction . The C-terminal proline-rich domain of Dab2-contains Grb2-binding motifs similar to that of the Ras guanine nucleotide exchanger Sos Lim et al, 1994;Wittekind et al, 1994). It has been shown previously that Dab2 binds Grb2 in vitro and in vivo, competing with Sos .…”

The LAT Story: A Tale of Cooperativity, Coordination, and Choreography

“…Sos1 binds Grb2 constitutively (Wittekind et al 1994;Houtman et al 2006). Following TCR ligation, Grb2 recruits Sos1 to the site of Ras localization at the membrane.…”