Recombinant Human SULT2A1 Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
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Recombinant Human SULT2A1 Protein, CF Summary

Details of Functionality
Measured by its ability to transfer sulfate from PAPS to t-Dehydroandrosterone (DHEA). The specific activity is >15 pmol/min/μg, as measured under the described conditions.
Source
E. coli-derived human Cytosolic Sulfotransferase 2A1/SULT2A1 protein
Ser2-Glu285, with an N-terminal Met and 6-His tag
Accession #
N-terminal Sequence
Met
Protein/Peptide Type
Recombinant Enzymes
Gene
SULT2A1
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
35 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
SDS-PAGE
36 kDa, reducing conditions
Publications
Read Publications using
5828-ST in the following applications:

Packaging, Storage & Formulations

Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.
Buffer
Supplied as a 0.2 μm filtered solution in Tris, NaCl and DTT.
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Assay Procedure
  • Assay Buffer: 50 mM Tris, 15 mM MgCl2, pH 7.5 (supplied in 10X form in kit)
  • Recombinant Human Cytosolic Sulfotransferase 2A1/SULT2A1 (rhSULT2A1) (Catalog # 5828-ST)
  • 3'-Phosphoadenosine-5'-phosphosulfate (PAPS) (Catalog # ES019)
  • T-Dehydroandrosterone (DHEA)  (Sigma, Catalog # D4000), 1 mM in 50% DMSO, 50% deionized water
  • Universal Sulfotransferase Activity Kit (Catalog # EA003)
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute Coupling Phosphatase 3 to 0.1 mg/mL in Assay Buffer.
  2. Prepare reaction mixture by combining 140 µL of 1 mM PAPS, 70 µL of 1 mM DHEA, 70 µL of 0.1 mg/mL Coupling Phosphatase 3, and 70 µL of Assay Buffer.
  3. Dilute rhSULT2A1 to 40 µg/mL in Assay Buffer.
  4. Dilute 1 mM Phosphate Standard 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.
  5. Continue 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 5.0 nmol per well.
  6. Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
  7. Load 25 µL of the 40 µg/mL rhSULT2A1 into the plate. Include a Control containing 25 µL of Assay Buffer.
  8. Add 25 µL of reaction mixture to the wells, excluding the standard curve.
  9. Cover the plate with parafilm or a plate sealer and incubate at 37 °C for 20 minutes.
  10. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  11. Add 100 µL of deionized water to all wells. Mix briefly.
  12. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate for 20 minutes at room temperature.
  13. Read plate at 620 nm (absorbance) in endpoint mode.
  14. 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:
  • rhSULT2A1: 1.0 µg
  • PAPS: 10000 pmoles (0.2 mM)
  • DHEA: 0.1 mM
  • Coupling Phosphatase 3: 0.5 µg

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human SULT2A1 Protein, CF

  • alcohol/hydroxysteroid sulfotransferase
  • Cytosolic Sulfotransferase 2A1
  • Dehydroepiandrosterone sulfotransferase
  • DHEAS
  • DHEA-ST
  • DHEA-STbile salt sulfotransferase
  • EC 2.8.2.14
  • HST
  • hSTa
  • Hydroxysteroid Sulfotransferase
  • ST2
  • ST2A1
  • ST2A3
  • STDbile-salt sulfotranasferase 2A1
  • Sulfotransferase 2A1
  • sulfotransferase family, cytosolic, 2A, dehydroepiandrosterone(DHEA)-preferring, member 1
  • SULT2A1

Background

Cytosolic sulfotransferases are a family of Phase II drug-metabolizing enzymes that catalyze the sulfation of many endogenous and xenobiotic substrates (1‑3). They have important roles in the metabolism of many endogenous compounds including steroids, bile acids, thyroid hormones and monoamine neurotransmitters. They are distributed throughout the body and serve to inactivate and increase water-solubility of xenobiotics and therapeutic drugs. They are distinct from Golgi resident sulfotransferases by lacking N-terminal signal-anchor domains and residing only in the cytoplasm. SULT2A1 is primarily expressed in liver and adrenal tissues where it sulfates steroids and bile acids (4‑6). It is also called DHEA/bile acid sulfotransferase and plays roles in maintaining cholesterol and bile acid homeostasis by increasing cholesterol catabolism and, at the same time, preventing toxicity from bile acid accumulation. The enzymatic activity of the recombinant human SULT2A1 is measured using a phosphatase-coupled assay (7).
  1. Falany, C. N. (1997) FASEB J. 11:206.
  2. Gamage, N. U. et al. (2006) Toxicol. Sci. 90:5.
  3. Allali-Hassani, A. et al. (2007) PLoS Biol. 5:e97.
  4. Comer, K. A. et al. (1993) Biochem. J. 289:233.
  5. Otterness, D.M. et al. (1992) Mol. Pharmacol. 41:865.
  6. Luu-The, V. et al. (1995) DNA Cell Biol. 14: 511.
  7. Prather, B. et al. (2012) Anal. Biochem. 423:86.

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Bioinformatics

Gene Symbol SULT2A1
Uniprot