Structural basis for recognition of the T cell adaptor protein SLP-76 by the SH3 domain of phospholipase Cγ1

The enzyme phospholipase Cγ1 (PLCγ1) is essential for T cell signaling and activation. Following T cell receptor ligation, PLCγ1 interacts through its SH2 and SH3 domains with the adaptors LAT and SLP-76, respectively, to form a multiprotein signaling complex that leads to activation of PLCγ1 by Syk tyrosine kinases. To identify the binding site for PLCγ1 in SLP-76, we used isothermal titration calorimetry to measure affinities for the interaction of PLCγ1-SH3 with a set of overlapping peptides spanning the central proline-rich region of SLP-76. PLCγ1-SH3 bound with high specificity to the SLP-76 motif ¹⁸⁶PPVPPQRP ¹⁹³, which represents the minimal binding site. To understand the basis for selective recognition, we determined the crystal structures of PLCγ1-SH3 in free form, and bound to a 10-mer peptide containing this site, to resolutions of 1.60 Å and 1.81 Å, respectively. The structures reveal that several key contacting residues of the SH3 shift toward the SLP-76 peptide upon complex formation, optimizing the fit and strengthening hydrophobic interactions. Selectivity results mainly from strict shape complementarity between protein and peptide, rather than sequence-specific hydrogen bonding. In addition, Pro193 of SLP-76 assists in positioning Arg192 into the compass pocket of PLCγ1-SH3, which coordinates the compass residue through an unusual aspartate. The PLCγ1-SH3/SLP-76 structure provides insights into ligand binding by SH3 domains related to PLCγ1-SH3, as well as into recognition by PLCγ1 of signaling partners other than SLP-76.