- Other Names
- Ectonucleotide Pyrophosphatase, Phosphodiesterase Family Member 6, NPP6, GPC-Cpde, Ectonucleotide Pyrophosphatase, Phosphodiesterase 6
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ENPP6 (ectonucleotide phosphodiesterase/pyrophosphatase member 6) is a choline-specific glycerophosphodiester phosphodiesterase that is part of the Nucleotide pyrophosphatases/phosphodiesterases (NPPs) family. The NPPs are ubiquitous membrane-associated or secreted ectoenzymes. The NPP family is conserved in vertebrates and the mammals have seven structurally-related ecto-enzymes. The ENPP family is divided in two groups ENPP1–3 and ENPP4–7. ENPP1–3 are multidomain proteins consisting of two N-terminal somatomedin B-like domains, a central phosphodiesterase (PDE) catalytic domain, and a C-terminal nuclease-like domain, whereas ENPP4–7 have only the PDE domain in common. In vitro, ENPP6 has phosphodiesterase activity to choline-containing compounds such as glycerophosphorylcholine (GPC) and O-phosphorylcholine N-acyl ethanolamine to produce phosphocholine. In addition, ENPP6 hydrolyzes choline-containing lysophospholipids, such as lysophophocolin (LPC), sphingosylphosphorylcholine (SPC), platelet-activating factor (PAF), and lysoPAF. Enpp6 has an active-site pocket that recognizes the choline head group of LPC substrates. ENPP6 is bound to the plasma membrane through its C-terminal glycosylphosphatidylinositol (GPI) anchor; also, the soluble form of ENPP6 has been identified.Product Details
- Human ENPP6 amino acids (Arg23 - Ser419) (Accession #BC035035) was expressed in CHO cells. The carboxy terminus contains TG-10His-GGQ tag.
- Molecular Mass
- The 412 amino acid recombinant protein has a predicted molecular mass of approximately 47.6 kD. The DTT-reduced and non-reduced protein migrate at approximately 65 kD and 100 kD respectively by SDS-PAGE.
- > 95% by SDS-PAGE gel as determined by Coomassie stained SDS-PAGE.
- 0.22 µm filtered protein solution is in 20 nM Tris, 0.3 M NaCl, pH 7.5.
- Endotoxin Level
- Less than 1.0 EU per µg cytokine as determined by the LAL method.
- 10-25 µg sizes are bottled at 200 µg/mL.
- Storage & Handling
- Unopened vial can be stored at -20°C or -70°C for six months. For maximum results, quick spin vial prior to opening. Avoid repeated freeze/thaw cycles.
- The activity of human ENPP6 is measured by its property to cleave O-(4-Nitrophenylphosphoryl) choline with a specific activity >4,000 pmol/µg/min.
- Application Notes
Bioassay Procedure and Materials
- Assay Buffer: 50 mM Tris, 0.5 M NaCl, pH 9.0
- Recombinant human ENPP6
- Substrate: O-(4-Nitrophenylphosphoryl) choline (Sigma, Catalog # N5879), 500 mM stock in deionized water
- 96-well clear plate
- Plate reader (model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute the recombinant ENPP6 to 0.4 µg/mL in assay buffer.
- Dilute the substrate at room temperature to 2 mM in assay buffer.
- Load 50 µL of 0.4 ng/µL of recombinant ENPP6 in a clear strip well plate.
- Start the reaction by adding 50 µL of 2 mM substrate. Include a substrate blank containing 50 µL assay buffer and 50 µL of substrate.
- Read at 405 nm (absorbance) in kinetic mode for 5 minutes.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
Adjusted Vmax* (OD/min) x Conversion Factor** (pmol/OD)
amount of enzyme (µg)
*Adjusted for Substrate Blank
**Derived using calibration standard p-Nitrophenol (Sigma, Catalog # 241326).
The final concentration of human recombinant ENPP6 is 0.2mg/ml (0.02 mg)
The final concentration of substrate is 1mM
BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue-ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are verified in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at firstname.lastname@example.org.
Endothelial cells of the sinusoid in the liver and developing oligodendrocytes. Expressed in kidney, brain, and liver.
- ENPP6 hydrolyzes choline-containing compound such as GPC (glycerophosphorylcholine), and supplies choline to sinusoidal endothelial cells and oligodendrocytes in development.
- Choline-containing compounds such as lysophosphatidylcholine and glycerophosphorylcholine.
- Hydrolyzes choline-containing compounds.
- Molecular Family
- Enzymes and Regulators, Phospho-Proteins
- Antigen References
- Sakagami H, et al. 2005. J. Biol. Chem. 280:23084.
- Stefan C, et al. 2005. Trends Biochem. Sci. 30:542.
- Nishimasu H, et al. 2011. Nat. Struct. Mol. Biol. 18:205.
- Greiner-Tollersrud L, et al. 2013. Neurochem. Res. 38:300.
- Morita J, et al. 2016. Sci. Rep. 6:20995.
- Gene ID
- 133121 View all products for this Gene ID
- View information about ENPP6 on UniProt.org
- Does specific activity of a recombinant protein vary between lots?
Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.
- Have your recombinants been tested for stability?
Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.
- How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?
- Use formula Specific activity (Units/mg) = 10e6/ ED50 (ng/mL)
- How does the activity of your recombinant proteins compare to competitors?
We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!
- What is the specific activity or ED50 of my recombinant protein?
The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.