A COMPARATIVE-ANALYSIS OF THE PHOSPHOINOSITIDE BINDING-SPECIFICITY OFPLECKSTRIN HOMOLOGY DOMAINS

Pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains are structurally related regulatory modules that are present in a variety of proteins involved in signal transduction, such as kinases, phospho lipases, GTP exchange proteins, and adapter proteins. Initially these domains were shown to mediate protein-protein interactions, but more r ecently they were also found to bind phosphoinositides. Most studies t o date have focused on binding of PH domains to phosphatidylinositol ( PtdIns)-4-P and PtdIns-4,5-P-2 and have not considered the lipid produ cts of phosphoinositide 3-kinase: PtdIns-3-P, PtdIns-3,4-P-2, and PtdI ns-3,4,5-P-3. Here we have compared the phosphoinositide specificity o f six different PH domains and the She PTB domain using all five phosp hoinositides. We show that the Bruton's tyrosine kinase PH domain bind s to PtdIns-3,4,5-P-3 with higher affinity than to PtdIns-4,5-P-2, Ptd Ins-3,4-P-2 or inositol 1,3,4,5-tetrakisphosphate (Ins-1,3,4,5 P-4). T his selectivity is decreased by the rid mutation (R28C). Selective bin ding of PtdIns-3,4,5 P-3 over PtdIns-4,5-P-2 or PtdIns-3,4-P-2 was als o observed for the amino-terminal PH domain of T lymphoma invasion and metastasis protein (Tiam-1), the PH domains of Son-of-sevenless (Sos) and, to a lesser extent, the PH domain of the beta-adrenergic recepto r kinase. The oxysterol binding protein and beta-spectrin PH domains b ound PtdIns-3,4,5-P-3 and PtdIns-4,5-P-2 with similar affinities. PtdI ns-3,4,5-P-3 and PtdIns-4,5-P-2 also bound to the PTB domain of She wi th similar affinities and lipid binding was competed with phosphotyros ine (Tyr(P)-containing peptides. These results indicate that distinct PH domains select for different phosphoinositides.