Recombinant Human ST3GAL1 Protein, CF
Recombinant Human ST3GAL1 Protein, CF Summary
Learn more about Fluorescent Glycan Labeling and DetectionProduct Specifications
Arg57-Arg340, with an N-terminal HA (YPYDVPDYA) tag
Analysis
Product Datasheets
6905-GT
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.
6905-GT
| Formulation | Supplied as a 0.2 μm filtered solution in Tris and NaCl. |
| Shipping | The product is shipped with dry ice or equivalent. 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
- Sialyltransferase Activity Kit (Catalog # EA002)
- 10X Assay Buffer: 250 mM Tris, pH 7.5
- MnCl2 (supplied in kit): 100 mM
- Recombinant Human beta -Gal alpha -2,3-Sialyltransferase/ST3GAL1 (rhST3GAL1) (Catalog # 6905-GT)
- CMP-Neu5Ac (Sigma, Catalog # C8271), 10 mM stock in deionized water
- beta 1-3 Galactosyl-N-acetyl galactosamine (V-labs, Catalog # GN213), 50 mM stock in deionized water
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Prepare 1X Assay Buffer containing 10 mM MnCl2 by combining equal volumes of 10X Assay Buffer and 100 mM MnCl2 and diluting 5-fold with deionized water.
- Dilute Phosphate Standard 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 nmoles per well.
- Prepare reaction mixture containing 0.4 mM CMP-Neu5Ac, 1 mM beta 1-3 Galactosyl-N-acetyl galactosamine, 4 μg/mL Coupling Phosphatase 2 in 1X Assay Buffer.
- Dilute rhST3GAL1 to 1 µg/mL in Assay Buffer.
- Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of 1X Assay Buffer.
- Load 25 µL of the 1 µg/mL rhST3GAL1 into the plate. Include a Control containing 25 µL of 1X Assay Buffer.
- Add 25 µL of reaction mixture to the wells, excluding the standard curve.
- Seal plate and incubate at 37 °C for 20 minutes.
- 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 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 or 4-parameter fitting and adjusted for Control.
Per Reaction:- rhST3GAL1: 0.025 μg
- Coupling Phosphatase 2: 0.1 μg
- CMP-Neu5Ac: 200 µM
- beta 1-3 Galactosyl-N-acetyl galactosamine: 0.5 mM
Background: ST3 beta-Gal alpha-2,3-Sialyltransferase 1/ST3GAL1
Sialyltransferases add sialic acid to glycoproteins or glycosphingolipids and play important roles in many biological processes including immune recognition, pathogen infection, and cell adhesion (1). ST3GAL1 is a type II membrane protein localized in the trans‑Golgi network that transfers sialic acid to galactose residues of core 1 O‑glycans (3Gal beta 1‑3GalNAc) to produce sialyl‑T antigen (NeuAc alpha 2‑3Gal beta 1‑3GalNAc) (2, 3). ST3GAL1 is ubiquitously expressed in most tissues, with highest expression in the placenta, liver and skeletal muscle (4). Over‑expression of ST3GAL1 may be involved in the tumorigenesis of both breast cancer (5, 6) and bladder cancer (7). The enzyme activity of the recombinant human ST3GAL1 was determined using a phosphatase‑coupled glycosyltransferase assay (8).
- Varki, A. (1999) Glycobiology 2:25.
- Jeanneau, C. et al. (2004) J. Biol. Chem. 279:13461.
- Dalziel, M. et al. (2001) J. Biol. Chem. 276:11007.
- Kitagawa, H. and Paulson, J.C. (1994) J. Biol. Chem. 269:17872.
- Picco, G. et al. (2010) Glycobiology 20:1241.
- Burchell, J. et al. (1999) Glycobiology 9:1307.
- Videira, P.A. et al. (2009) BMC Cancer 9:357.
- Wu, Z.L. et al. (2011) Glycobiology 21:727.
Citation for Recombinant Human ST3GAL1 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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Fluorescent glycan fingerprinting of SARS2 spike proteins
Authors: ZL Wu, JM Ertelt
Scientific Reports, 2021-10-14;11(1):20428.
Species: Human
Sample Types: Recombinant Protein
Applications: Bioassay
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