Sequences surrounding the Src-homology 3 domain of phospholipase Cγ-1 increase the domain's association with Cbl

SH3 domains are protein modules that interact with proline-rich polypeptide fragments. Cbl is an adapter-like protein known to interact with several SH3 domains, including the PLC gamma 1 SH3 domain and the Grb2 amino terminal SH3 domain. Here we explore whether sequences surrounding the PLC gamma 1 SH3 domain core sequence (aa.796-851) can affect the binding to Cbl, a target used as a prototypical ligand. Consistent with previous reports, our results demonstrated a weak binding of Cbl to GST fusion proteins that strictly encompass the structural core of the PLC gamma 1 SH3 domain but a high-avidity binding to the Grb2 amino-terminal SH3 domain. Inclusion of amino acids immediately flanking the PLC gamma 1 SH3 core domain, however, substantially increased binding of Cbl to a level comparable to that of the Grb2 amino-terminal SH3 domain. The interaction of this extended PLC gamma 1 SH3 domain fusion protein with Cbl was shown to depend entirely upon the interaction of the domain with a proline-rich motif in Cbl, ruling out the possibility that amino acids adjacent to the core SH3 domain of PLC gamma 1 provide independent Cbl binding. These data suggest that sequences surrounding the SH3 domain of PLC gamma 1 may contribute to or stabilize the association of the domain with the target protein, thus increasing its binding efficiency. (C) 1998 Academic Press.