Dataset Information

Molecular basis of phosphatidylinositol 4-phosphate and ARF1 GTPase recognition by the FAPP1 pleckstrin homology (PH) domain.

ABSTRACT: Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane trafficking, the molecular basis of its dual function remains unclear. Here, we report a 1.9 Å resolution crystal structure of the FAPP1 PH domain and detail the molecular mechanisms of the PtdIns(4)P and ARF1 recognition. The FAPP1 PH domain folds into a seven-stranded ?-barrel capped by an ?-helix at one edge, whereas the opposite edge is flanked by three loops and the ?4 and ?7 strands that form a lipid-binding pocket within the ?-barrel. The ARF1-binding site is located on the outer side of the ?-barrel as determined by NMR resonance perturbation analysis, mutagenesis, and measurements of binding affinities. The two binding sites have little overlap, allowing FAPP1 PH to associate with both ligands simultaneously and independently. Binding to PtdIns(4)P is enhanced in an acidic environment and is required for membrane penetration and tubulation activity of FAPP1, whereas the GTP-bound conformation of the GTPase is necessary for the interaction with ARF1. Together, these findings provide structural and biochemical insight into the multivalent membrane anchoring by the PH domain that may augment affinity and selectivity of FAPP1 toward the TGN membranes enriched in both PtdIns(4)P and GTP-bound ARF1.

SUBMITTER: He J

PROVIDER: S-EPMC3099681 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Molecular basis of phosphatidylinositol 4-phosphate and ARF1 GTPase recognition by the FAPP1 pleckstrin homology (PH) domain.

He Ju J Scott Jordan L JL Heroux Annie A Roy Siddhartha S Lenoir Marc M Overduin Michael M Stahelin Robert V RV Kutateladze Tatiana G TG

The Journal of biological chemistry 20110322 21

Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane trafficking, the molecular basis of its dual function remains unclear. Here, we report a 1.9 Å resolution crystal structure of the FAP ...[more]

PMID: 21454700

Similar Datasets

Project description:Background:Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] is an important regulator of several cellular processes and a precursor for other second messengers which are involved in cell signaling pathways. Signaling proteins preferably interact with PI(4,5)P2 through its pleckstrin homology (PH) domain. Efforts are underway to design small molecule-based antagonist, which can specifically inhibit the PI(4,5)P2/PH-domain interaction to establish an alternate strategy for the development of drug(s) for phosphoinositide signaling pathways. Methods:Surface plasmon resonance, molecular docking, circular dichroism, competitive Förster resonance energy transfer, isothermal titration calorimetric analyses and liposome pull down assay were used. Results:In this study, we employed 1,2,3-triazol-4-yl methanol containing small molecule (CIPs) as antagonists for PI(4,5)P2/PH-domain interaction and determined their inhibitory effect by using competitive-surface plasmon resonance analysis (IC50 ranges from 53 to 159 nM for PI(4,5)P2/PLC?1-PH domain binding assay). We also used phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3], phosphatidylinositol 3,4-bisphosphate [PI(3,4)P2], PI(4,5)P2 specific PH-domains to determine binding selectivity of the compounds. Various physicochemical analyses showed that the compounds have weak affect on fluidity of the model membrane but, strongly interact with the phospholipase C ?1 (PLC?1)-PH domains. The 1,2,3-triazol-4-yl methanol moiety and nitro group of the compounds are essential for their exothermic interaction with the PH-domains. Potent compound can efficiently displace PLC?1-PH domain from plasma membrane to cytosol in A549 cells. Conclusions:Overall, our studies demonstrate that these compounds interact with the PIP-binding PH-domains and inhibit their membrane recruitment. General significance:These results suggest specific but differential binding of these compounds to the PLC?1-PH domain and emphasize the role of their structural differences in binding parameters. These triazole-based compounds could be directly used/further developed as potential inhibitor for PH domain-dependent enzyme activity.

Dataset Information

Molecular basis of phosphatidylinositol 4-phosphate and ARF1 GTPase recognition by the FAPP1 pleckstrin homology (PH) domain.

Publications

Molecular basis of phosphatidylinositol 4-phosphate and ARF1 GTPase recognition by the FAPP1 pleckstrin homology (PH) domain.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets