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The basic function of the Src homology 3 (SH3) domain is considered to be binding to proline-rich sequences containing a PxxP motif. Recently, many SH3 domains, including those from Grb2 and Pex13p, were reported to bind sequences lacking a PxxP motif. We report here that the 22 residue peptide lacking a PxxP motif, derived from p47phox, binds to the C-terminal SH3 domain from p67phox. We applied the NMR cross-saturation method to locate the interaction sites for the non-PxxP peptides on their cognate SH3 domains from p67phox, Grb2 and Pex13p. The binding site of the Grb2 SH3 partially overlapped the conventional PxxP-binding site, whereas those of p67Pphox and Pex13p SH3s are located in different surface regions. The non-PxxP peptide from p47phox binds to the p67Pphox SH3 more tightly when it extends to the N-terminus to include a typical PxxP motif, which enabled the structure determination of the complex, to reveal that the non-PxxP peptide segment interacted with the p67phox SH3 in a compact helix-turn-helix structure (PDB entry 1K4U). [ABSTRACT FROM AUTHOR]

An analysis of the backbone dynamics of the C-terminal Src homology 3 (SH3) domain of p67phoxSH3(C), in complex with a 32-residue high-affinity (Kd = 24 nM) peptide, Pf, from the C-terminal region of p47phox is presented. This paper represents the first detailed analysis of the backbone dynamics and the ligand-induced changes therein of a high-affinity, high-specificity interaction involving an SH3 domain. The dynamic features are compared with those in the high-affinity, highly specific interaction between the SH3 domain of C-terminal Src kinase (Csk-SH3) and a proline-rich peptide from proline-enriched phosphatase (PEP). Both systems share common dynamic features especially in the canonical PxxP motif recognition surface where slow micro- to millisecond time scale dynamics persist on complex formation especially in several residues that are implicated in ligand recognition and in stabilizing the SH3 fold. These residues are highly conserved in SH3 domains. Ile505, which lies outside the PxxP recognition motif on p67phoxSH3(C) and is key in conferring high specificity to the p67phoxSH3(C)/Pf interaction, becomes more disordered upon complex formation. This behavior is similar to that seen in the residues that constitute the specificity surface in Csk-SH3. [ABSTRACT FROM AUTHOR]