Key Specifications Table
| Species | Host Cells | Protein Target |
|---|---|---|
| Human | Chem-1 | S1P3 / EDG3 |
| Description | |
|---|---|
| Catalogue Number | HTS097M |
| Trade Name |
|
| Description | ChemiSCREEN™ Membrane Preparation Recombinant Human S1P3 Lysophospholipid Receptor |
| Overview | Full-length human EDG3 cDNA encoding S1P3 |
| Background Information | Sphingosine 1-phosphate (S1P) is a bioactive lipid that binds to and activates a family of GPCRs, S1P1-5 (also known as EDG receptors). Interactions between S1P and its receptors mediate cytoskeletal rearrangement and cell migration, with functional consequences in angiogenesis, lymphocyte trafficking, and smooth muscle development (Anliker and Chun, 2004). S1P1 (Edg-1) signals exclusively through Gi, whereas S1P2 (Edg-5) and S1P3 (Edg-3) activate Gi, Gq and G12/13 (Windh et al., 1999). Although S1P1 and S1P3 promote cell migration, S1P2 inhibits cell migration in several cell types; these opposing functions appear to result from differences in the ability of each receptor to activate Gi (Sugimoto et al., 2003). Studies with knockout mice indicate that S1P2 and S1P3 have redundant functions in maintaining vascular integrity during embryonic development (Kono et al., 2004). In addition, S1P3 regulates immune responses by contributing to endothelial barrier function in splenic marginal zones (Girkontaite et al., 2004). Millipores S1P3 membrane preparations are crude membrane preparations made from our proprietary stable recombinant cell lines to ensure high-level of GPCR surface expression; thus, they are ideal HTS tools for screening of S1P3 interactions with its ligands. The membrane preparation exhibits EC50 of 1.7nM for S1P in a GTPγS binding assay. |
| Product Information | |
|---|---|
| Format | Membranes |
| Presentation | Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol with no preservatives. Packaging method: Membrane protein was adjusted to 1 mg/ml in packaging buffer, rapidly frozen, and stored at -80°C |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | Human S1P3 / EDG3 GPCR membrane preparation for GTPγS Binding & Radioligand Binding Assays. |
| Key Applications |
|
| Application Notes | GTPγS Binding and Radioligand Binding Assay. |
| Biological Information | |
|---|---|
| Entrez Gene Number |
|
| Entrez Gene Summary | This gene encodes a member of the EDG family of receptors, which are G protein-coupled receptors. This protein has been identified as a functional receptor for sphingosine 1-phosphate and likely contributes to the regulation of angiogenesis and vascular endothelial cell function. |
| Gene Symbol |
|
| GPCR Class | A |
| Host Cells | Chem-1 |
| Protein Target | S1P3 / EDG3 |
| Target Sub-Family | Lysophospholipid |
| UniProt Number |
|
| UniProt Summary | FUNCTION: SwissProt: Q99500 # Receptor for the lysosphingolipid sphingosine 1- phosphate (S1P). S1P is a bioactive lysophospholipid that elicits diverse physiological effect on most types of cells and tissues. When expressed in rat HTC4 hepatoma cells, is capable of mediating S1P-induced cell proliferation and suppression of apoptosis. SIZE: 378 amino acids; 42250 Da SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein. TISSUE SPECIFICITY: Expressed in all tissues, but most abundantly in heart, placenta, kidney, and liver. SIMILARITY: SwissProt: Q99500 ## Belongs to the G-protein coupled receptor 1 family. |
| Incubation Conditions | ASSAY CONDITIONS: Membranes are permeabilized by addition of saponin to an equal concentration by mass, then mixed with [35S]-GTPγS (final concentration of 0.1 nM) in 20 mM HEPES, pH 7.4/100 mM NaCl/10 mM MgCl2/0.5 µM GDP in a nonbinding 96-well plate. Unlabeled S1P is added to the final concentration indicated in Figure 1 (final volume 100 µL), and incubated for 30 min at 30°C. The binding reaction is transferred to a GF/B filter plate (Millipore MAHF B1H) previously prewetted with water, and washed 3 times (1 mL per well per wash) with cold 10 mM sodium phosphate, pH 7.4. The plate is dried and counted. One vial contains enough membranes for at least 200 assays (units), where one unit is the amount of membrane that will yield greater than 1000 cpm specific S1P -stimulated [35S]-GTPγS binding. The S1P3 membrane preparation is expected to be functional in a radioligand binding assay; however, the end user will need to determine the optimal radiolabeled ligand for use with this product. |
| Product Usage Statements | |
|---|---|
| Quality Assurance | EC50 in GTPγS binding assay by S1P: ~ 1.7nM |
| Usage Statement |
|
| Storage and Shipping Information | |
|---|---|
| Storage Conditions | Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw. |
| Packaging Information | |
|---|---|
| Material Size | 200 units |