Crystal structure of the phosphatidylinositol 3,4-bisphosphate-binding pleckstrin homology (PH) domain of tandem PH-domain-containing protein 1 (TAPP1): molecular basis of lipid specificity

Phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P-3] and its immedia te breakdown product PtdIns(3,4)P-2 function as second messengers in growth factor- and insulin-induced signalling pathways. One of the ways that thes e 3-phosphoinositides are known to regulate downstream signalling events is by attracting proteins that possess specific PtdIns-binding pleckstrin hom ology (PH) domains to the plasma membrane. Many of these proteins, such as protein kinase B, phosphoinositide-dependent kinase 1 and the dual adaptor for phosphotyrosine and 3-phosphoinositides (DAPP1) interact with both PtdI ns(3,4,5)P-3 and PtdIns(3,4)P-3 with similar affinity. Recently, a new PH-d omain-containing protein, termed tandem PH-domain-containing protein (TAPP) 1, was described which is the first protein reported to bind PtdIns(3,4)P- 2 specifically. Here we describe the crystal structure of the PtdIns(3,4)P- 2-binding PH domain of TAPP1 at 1.4 Angstrom (1 Angstrom = 0.1 nm) resoluti on in complex with an ordered citrate molecule. The structure is similar to the known structure of the PH domain of DA-PP1 around the D-3 and D-4 inos itol-phosphate-binding sites. However, a glycine residue adjacent to the D- 5 inositol-phosphate-binding site in DAPP1 is substituted for a larger alan ine residue in TAPP1. which also induces a conformational change in the nei ghbouring residues. We show that mutation of this glycine to alanine in DAP P1 converts DAPP1 into a TAPP1-like PH domain that only interacts with PtdI ns(3,4)P-2, whereas the alanine to glycine mutation in TAPP1permits the TAP P1 PH domain to interact with PtdIns(3,4,5)P-3.