Recombinant Mouse ENPP-2/Autotaxin Protein, CF Summary
Ser49-Ile862, with an N-terminal 6-His tag
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
|Formulation||Supplied as a 0.2 μm filtered solution in Tris and NaCl.|
|Shipping||The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.|
|Stability & Storage:||Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
- Assay Buffer: 50 mM Tris, 20 mM CaCl2, 0.01% Brij-35 (w/v), pH 8.0
- Recombinant Mouse ENPP‑2/Autotaxin (Catalog # 6187-EN)
- Substrate: Bis(p-Nitrophenyl) Phosphate Sodium Salt (BPNPP) (Sigma, Catalog # N3002), 40 mM stock in deionized water (Note: Heating may be necessary to solubilize Substrate.)
- 0.2 M NaOH
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute rmENPP-2 to 2 ng/µL in Assay Buffer.
- Dilute Substrate to 2 mM in Assay Buffer.
- Load into plate 50 µL of 2 ng/µL rmENPP-2, and start the reaction by adding 50 µL of 2 mM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 2 mM Substrate.
- Incubate plate at room temperature for 10 minutes.
- Stop the reaction by adding 100 µL of 0.2 M NaOH to all wells.
- Read at 410 nm (absorbance) in endpoint mode.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
|Adjusted Abs* (OD) x Conversion Factor** (pmol/OD)|
|Incubation time (min) x amount of enzyme (µg)|
*Adjusted for Substrate Blank
**Derived using calibration standard p-Nitrophenol (Sigma-Aldrich, Catalog # 241326).
- rmENPP-2: 0.10 µg
- Substrate: 0.5 mM
ENPP-2, also known as Autotaxin, belongs to the ectonucleotide pyrophosphatase/phosphodiesterase (NPP) family. Some NPPs hydrolyze phosphates from nucleotides and their derivatives. ENPP-2 shares 40‑50% identity to ENPP-1 & -3, all of which contain an N‑terminal intracellular domain, a single transmembrane domain and a large extracellular domain that includes a catalytic domain, two somatomedin B-like domains, and a C-terminal nuclease like domain (1). Unlike ENPP‑1 and ENPP‑3, ENPP‑2 has weak activity against nucleotides, but exhibits a lysophospholipase D activity which allows the formation of lysophosphatidic acid (LPA) and choline from lysophosphatidylcholine (2). ENPP-2 can also hydrolyze sphingosylphosphorylcholine to produce sphingosine-1-phosphate, a lipid messenger molecule. The hydrolysis of nucleotides and lysophospholipids by ENPP2 is mediated by a single catalytic site (2). LPA and sphingosine-1-phosphate are inhibitors of ENPP-2 (3). ENPP-2 was originally found to stimulate tumor cell motility and has since been found to enhance tumor invasion and metastasis (4) and to be up‑regulated in several types of carcinomas including breast and lung (5). ENPP-2 is most highly expressed in brain and adipose tissue (6). Recombinant mouse ENPP-2 was expressed without its transmembrane and intracellular domains, resulting in the secretion of the recombinant enzyme.
- Cimpean, A. et al. (2004) Biochem. J. 381:71.
- Gijsbers, R. et al. (2003) FEBS Letters. 538:60.
- Van Meeteren, L.A. et al. (2005) J. Biol. Chem. 280:21155.
- Nam, S.W. et al. (2000) Oncogene 19:241.
- Jansen, S. et al. (2005) J. Cell Sci. 118:3081.
- Giganti, A. et al. (2008) J. Biol. Chem. 283:7776.
Product Specific NoticesCoomassie is a registered trademark of Imperial Chemical Industries Ltd.
Citations for Recombinant Mouse ENPP-2/Autotaxin Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
Citations: Showing 1 - 2
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The involvement of autotaxin in renal interstitial fibrosis through regulation of fibroblast functions and induction of vascular leakage
Authors: N Sakai, G Bain, K Furuichi, Y Iwata, M Nakamura, A Hara, S Kitajima, A Sagara, T Miyake, T Toyama, K Sato, S Nakagawa, M Shimizu, S Kaneko, T Wada
Sci Rep, 2019;9(1):7414.
Sample Types: Whole Cells
Non-invasive imaging of tumors by monitoring autotaxin activity using an enzyme-activated near-infrared fluorogenic substrate.
Authors: Madan, Damian, Ferguson, Colin G, Lee, Won Yong, Prestwich, Glenn D, Testa, Charles
PLoS ONE, 2013;8(11):e79065.
Sample Types: Complex Sample Type
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