Phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P3] and its immediate breakdown product PtdIns(3,4)P2 function as second messengers in growth factor- and insulin-induced signalling pathways. One of the ways that these 3-phosphoinositides are known to regulate downstream signalling events is by attracting proteins that possess specific PtdIns-binding pleckstrin homology (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 PtdIns(3,4,5)P3 and PtdIns(3,4)P2 with similar affinity. Recently, a new PH-domain-containing protein, termed tandem PH-domain-containing protein (TAPP) 1, was described which is the first protein reported to bind PtdIns(3,4)P2 specifically. Here we describe the crystal structure of the PtdIns(3,4)P2-binding PH domain of TAPP1 at 1.4 Å (1 Å = 0.1 nm) resolution in complex with an ordered citrate molecule. The structure is similar to the known structure of the PH domain of DAPP1 around the D-3 and D-4 inositol-phosphate-binding sites. However, a glycine residue adjacent to the D-5 inositol-phosphate-binding site in DAPP1 is substituted for a larger alanine residue in TAPP1, which also induces a conformational change in the neighbouring residues. We show that mutation of this glycine to alanine in DAPP1 converts DAPP1 into a TAPP1-like PH domain that only interacts with PtdIns(3,4)P2, whereas the alanine to glycine mutation in TAPP1 permits the TAPP1 PH domain to interact with PtdIns(3,4,5)P3.
- protein structure
The coordinates and structure factors will appear in the PDB under the accession numbers 1eaz and r1eazsf respectively.
Abbreviations used: ARF, ADP-ribosylation factor; BTK, Bruton's tyrosine kinase; DAPP1, dual adaptor for phosphotyrosine and 3-phosphoinositides; GST, glutathione S-transferase; PH, pleckstrin homology; PI 3-kinase, phosphoinositide 3-kinase; SHIP, SH2-containing inositol phosphatase; GRP1; general receptor for phosphoinositides-1; TAPP, tandem PH-domain-containing protein; TAPP1-PHCT, PtdIns(3,4)P2-binding PH domain of TAPP1; VL, variable loop.
- The Biochemical Society, London ©2001