Recombinant Mouse ST3GAL5/GM3 Synthase Protein, CF
R&D Systems | Catalog # 8454-GT
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Key Product Details
- R&D Systems CHO-derived Recombinant Mouse ST3GAL5/GM3 Synthase Protein (8454-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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Product Specifications
Source
Chinese Hamster Ovary cell line, CHO-derived mouse ST3GAL5/GM3 Synthase protein
Thr87-His414, with an N-terminal 6-His tag
Thr87-His414, with an N-terminal 6-His tag
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Endotoxin Level
<0.1 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
His
Predicted Molecular Mass
38 kDa
SDS-PAGE
45-60 kDa, reducing conditions
Activity
Measured by its ability to transfer Neu5Ac from CMP-Neu5Ac to alpha -lactose.
The specific activity is >5 pmol/min/μg, as measured under the described conditions.
The specific activity is >5 pmol/min/μg, as measured under the described conditions.
Formulation, Preparation, and Storage
8454-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: ST3GAL5/GM3 Synthase
References
- Varki, A. (1999) Glycobiology 2:25.
- Ishii, A. et al (1998) J. Biol. Chem. 273:31652.
- Uemura, S. et al (2009) Mol. Biol. Cell 20:3088.
- Simpson, M. A. et al. (2004) Nat. Genet. 36:1225.
- Tagami, S. et al. (2002) J. Biol. Chem. 277:3085.
- Wu, Z.L. et al. (2011) Glycobiology 21:727.
Long Name
ST3 beta-Galactoside alpha-2,3-Sialyltransferase 5
Alternate Names
3S-T, Ganglioside GM3 Synthase, SATI, Siat9, ST3GALV
Gene Symbol
ST3GAL5
UniProt
Additional ST3GAL5/GM3 Synthase Products
Product Documents for Recombinant Mouse ST3GAL5/GM3 Synthase 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 Mouse ST3GAL5/GM3 Synthase Protein, CF
For research use only
Related Research Areas
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Protocols
View specific protocols for Recombinant Mouse ST3GAL5/GM3 Synthase Protein, CF (8454-GT):
Materials
- Assay Buffer: 25 mM MES, pH 7.0
- Recombinant Mouse ST3GAL5/GM3 Synthase (rmST3GAL5) (Catalog # 8454-GT)
- Donor Substrate: CMP-Sialic Acid (CMP-Neu5Ac) (Sigma, Catalog # C8271), 10 mM stock in deionized water
- Acceptor Substrate: alpha -Lactose (Sigma, Catalog # L2643), 300 mM stock in deionized water
- Sialyltransferase Activity Kit (Catalog # EA002)
- 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.
- Prepare standard curve by performing six one-half serial dilutions of the 100 µM Phosphate stock in Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
- Prepare reaction mixture containing 0.4 mM CMP-Sialic Acid, 80 mM alpha -lactose, and 2 µg/mL Coupling Phosphatase 2.
- Dilute rmST3GAL5 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 rhST6GAL1 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 room temperature for 3 hours.
- Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
- Add 100 µL of deionized water to all wells.
- Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate for 20 minutes at room 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:
- rmST3GAL5: 1.0 µg
- Coupling Phosphatase 2: 0.05 µg
- CMP-Sialic Acid: 0.2 mM
- alpha -Lactose: 40 mM
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