Recombinant Human Fucosyltransferase 11/FUT11 Protein, CF
Recombinant Human Fucosyltransferase 11/FUT11 Protein, CF Summary
Learn more about Fluorescent Glycan Labeling and DetectionProduct Specifications
Gly25-Leu492, with an N-terminal 6-His tag
Analysis
Product Datasheets
Carrier Free
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.
5964-GT
| 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 Procedure
- Glycosyltransferase Activity Kit (Catalog # EA001)
- 10X Assay Buffer (supplied in kit): 250 mM Tris, 100 mM CaCl2, pH 7.5
- MnCl2 (supplied in kit): 100 mM
- Recombinant Human Fucosyltransferase 11/FUT11 (rhFUT11) (Catalog # 5964-GT)
- GDP-Fucose (Sigma, Catalog # G4401), 1.6 mM stock in deionized water
- 96-well Clear Plate (Catalog # DY990)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Prepare 1X Assay Buffer containing 10 mM MnCl2 by combining 10X stocks and diluting 10-fold with deionized water.
- Dilute 1 mM Phosphate Standard provided by the Glycosyltransferase Kit by adding 40 µL of the 1 mM Phosphate Standard to 360 µL of 1X Assay Buffer for a 100 µM stock. This is the first point of the standard curve.
- Complete the standard curve by performing six one-half serial dilutions of the 100 µM Phosphate stock using 1X Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
- Prepare reaction mixture containing 0.2 mM GDP-Fucose and 8 µg/mL Coupling Phosphatase 1 in Assay Buffer.
- Dilute rhFUT11 to 80 ng/µL in Assay Buffer.
- Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
- Load 25 µL of 80 ng/µL rhFUT11 into empty wells of the same plate as the curve. Include a Control containing 25 μL of Assay Buffer.
- Add 25 µL of the reaction mixture to all wells, excluding the standard curve.
- Seal plate and incubate at room temperature for 1 hour.
- Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
- Add 100 µL of deionized water to all wells. Mix briefly.
- Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate sealed plate for 20 minutes at room temperature.
- Read plate at 620 nm (absorbance) in endpoint mode.
- Calculate specific activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted phosphate released* (nmol) x (1000 pmol/nmol) |
| Incubation time (min) x amount of enzyme (µg) |
*Derived from the phosphate standard curve using linear fitting and adjusted for Control.
Per Reaction:- rhFUT11: 2 µg
- Coupling Phosphatase 1: 0.2 µg
- GDP-Fucose: 100 µM
Reconstitution Calculator
Background: Fucosyltransferase 11/FUT11
Fucose is frequently found at terminal sites on various glycans and is essential for the generation of many sugar epitopes. Well-known fucose containing glycans include Lewis and ABO blood group antigens. Lewis epitopes are key elements involved in leukocyte homing and extravasation process, thus are essential for lymphocyte maturation and natural defense functions (1). O-fucosylation on Notch receptor is found to be essential for its signaling function (2). So far, 11 carbohydrate specific fucosyltransferases are found in humans (3). FUT1 and FUT2 are alpha 1-2 fucosyltransferases and are responsible for ABO blood group antigen synthesis (4). FUT3, FUT4, FUT5, FUT6, FUT7 and FUT9 are alpha 1-3/4 fucosyltransferases and are responsible for Lewis antigen generation (5). FUT10 and FUT11 are newly cloned alpha 1-3 fucosyltransferases that are distinct from the alpha 1-3/4 fucosyltransferase subfamily and are able to introduce fucose to the innermost core GlcNAc of the N-glycan on conalbumin glycopeptides and biantennary N-glycan acceptors but not onto short lactosaminyl acceptor substrates (6). Predicted as a type II transmembrane protein and a Golgi resident enzyme, the exact function of this enzyme needs to be further characterized. The activity of this enzyme has been measured with a phosphatasecoupled method (7).
- Weston, B. W. et al. (1992) J. Biol.Chem. 267:4152.
- Stahl, M. et al. (2008) J. Biol.Chem. 283:13638.
- Becker, D.J. et al. (2003) Glycobiology 13:41R.
- Kelly, R. J. et al. (1995) J. Biol.Chem. 270:4640.
- Weston, B. W. et al. (1992) J. Biol.Chem. 267:24575.
Citation for Recombinant Human Fucosyltransferase 11/FUT11 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.
1 Citation: Showing 1 - 1
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Core-6 fucose and the oligomerization of the 1918 pandemic influenza viral neuraminidase.
Authors: Wu Z, Zhou H, Ethen C, N Reinhold V
Biochem Biophys Res Commun, 2016-03-21;473(2):524-9.
Applications: Bioassay
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