Recombinant Human ST8SIA1 Protein, CF
R&D Systems | Catalog # 6716-GT
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Key Product Details
- R&D Systems CHO-derived Recombinant Human ST8SIA1 Protein (6716-GT)
- Quality control testing to verify active proteins with lot specific assays by in-house scientists
- All R&D Systems proteins are covered with a 100% guarantee
Source
CHO
Accession Number
Applications
Enzyme Activity
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Learn more about Fluorescent Glycan Labeling and Detection
Product Specifications
Source
Chinese Hamster Ovary cell line, CHO-derived human ST8 alpha-2,8-Sialyltransferase 8A/ST8SIA1 protein
Tyr49-Ser356, with a C-terminal 6‑His tag
Tyr49-Ser356, with a C-terminal 6‑His tag
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
Tyr49
Predicted Molecular Mass
36 kDa
SDS-PAGE
40-60 kDa, reducing conditions
Activity
Measured by its ability to transfer Neu5Ac from CMP-Neu5Ac to fetuin of fetal calf serum.
The specific activity is >15 pmol/min/μg, as measured under the described conditions.
The specific activity is >15 pmol/min/μg, as measured under the described conditions.
Formulation, Preparation, and Storage
6716-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.
|
Background: ST8 alpha-2,8-Sialyltransferase 8A/ST8SIA1
References
- Kolter, T. et al. (2002) J. Biol. Chem. 277:25859.
- Cheresh, D.A. et al. (1986) J. Cell. Biol. 102:688.
- Nakayama, J. et al. (1986) J. Biol. Chem. 271:3684.
- Nara, K. et al. (1994) Proc. Natl. Acad. Sci. USA. 91:7952.
- Haraguchi, M. et al. (1994) Proc. Natl. Acad. Sci. USA. 91:10455.
- Crespo, P.M. et al. (2010) J. Biol. Chem. 285:29179.
- Wu, Z.L. et al. (2010) Glycobiology doi: 10.1093/glycob/cwq187.
Long Name
ST8 alpha-N-acetyl-neuraminide alpha-2,8-sialyltransferase 1
Alternate Names
GD3S, SIAT8A, ST8SiaI
Gene Symbol
ST8SIA1
UniProt
Additional ST8 alpha-2,8-Sialyltransferase 8A/ST8SIA1 Products
Product Documents for Recombinant Human ST8SIA1 Protein, CF
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Human ST8SIA1 Protein, CF
For research use only
Related Research Areas
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Protocols
View specific protocols for Recombinant Human ST8SIA1 Protein, CF (6716-GT):
Materials
- Sialyltransferase Activity Kit (Catalog # EA002)
- Assay Buffer: 50 mM MES, 5 mM MgCl2, pH 6.5
- Recombinant Human alpha -2,8‑Sialyltransferase 8A/ST8SIA1 (rhST8SIA1) (Catalog # 6716-GT)
- Donor Substrate: CMP-Neu5Ac (Sigma, Catalog # C8271), 10 mM stock in deionized water
- Acceptor Substrate: Fetuin (Sigma, Catalog # F3385), 50 mg/mL stock in deionized water
- 96-well Clear Plate (Catalog # DY990)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute 1 mM Phosphate Standard provided by the Sialyltransferase Activity Kit by adding 40 μL of the 1 mM Phosphate Standard to 360 μL of 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 Assay Buffer. The standard curve has a range of 0.078 to 5 nmoles per well.
- Dilute CMP-Neu5Ac to 2.5 mM in Assay Buffer.
- Dilute Coupling Phosphatase 2 to 25 µg/mL in Assay Buffer.
- Prepare reaction mixture by combining 50 µL of 2.5 mM CMP-Neu5Ac, 50 µL of 25 µg/mL Coupling Phosphatase 2, 62.5 µL of 50 mg/mL Fetuin, and 150 µL of Assay Buffer.
- Dilute rhST8SIA1 to 40 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 Assay Buffer.
- Load 25 µL of the 40 µg/mL rhST8SIA1 into the plate. Include a Control containing 25 µL of Assay Buffer.
- Add 25 µL of reaction mixture to the wells, excluding the standard curve.
- Cover the plate with parafilm or a plate sealer 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) = |
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:
- rhST8SIA1: 1.0 µg
- Coupling Phosphatase 2: 0.1 µg
- Fetuin: 250 µg
- CMP-Neu5Ac: 0.2 mM