Structural basis for specificity switching of the Src SH2 domain.

The Src SH2 domain binds pYEEI-containing phosphopeptides in an extended conformation with a hydrophobic pocket, which includes ThrEF1, binding Ile(pY +3). Mutating ThrEF1 to tryptophan switches specificity to an Asn(pY +2) requirement, yielding a biological mimic of the Grb2 SH2 domain. Here we show that the Src ThrEF1Trp SH2 domain mutant binds pYVNV phosphopeptides in a beta turn conformation, which, despite differing conformations of the interacting tryptophan, closely resembles the native Grb2/pYVNV cognate peptide binding mode. The ThrEF1Trp substitution therefore switches specificity by physically occluding the pTyr +3 binding pocket and by providing additional interaction surface area for Asn(pY +2). This demonstrates structurally how novel SH2 domain specificities may rapidly evolve through single amino acid substitutions and suggests how new signaling pathways may develop.