Recombinant Human SULT1B1 Protein, CF

• Reactivity: Hu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Human SULT1B1 Protein, CF Summary

• Details of Functionality: Measured by its ability to transfer sulfate from PAPS to 1-Napthol. The specific activity is >30 pmol/min/μg, as measured under the described conditions.
• Source: E. coli-derived human Cytosolic Sulfotransferase 1B1/SULT1B1 protein
  Leu2-Ile296, with an N-terminal 6-His tag
  Accession # O43704
• Accession #: O43704
• N-terminal Sequence: His
• Protein/Peptide Type: Recombinant Enzymes
• Gene: SULT1B1
• 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

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 36 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

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, Glycerol 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 1B1/SULT1B1 (rhSULT1B1) (Catalog # 5959-ST)
  • 3'-Phosphoadenosine-5'-phosphosulfate (PAPS) (Catalog # ES019)
  • 1-Naphthol (Sigma, Catalog # N1000), 10 mM in deionized water
  • Universal Sulfotransferase Activity Kit (Catalog # EA003)
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute 1 mM Phosphate Standard provided by the Universal Sulfotransferase 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.
  2. Prepare additional curve points 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.
  3. Prepare reaction mixture containing 0.4 mM PAPS, 0.4 mM 1-Naphthol, and 0.02 mg/mL Coupling Phosphatase 3 in Assay Buffer.
  4. Dilute rhSULT1B1 to 40 µg/mL in Assay Buffer.
  5. Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
  6. Load 25 µL of the 40 µg/mL rhSULT1B1 into the plate. Include 25 µL of Assay Buffer to be used as a Control.
  7. Add 25 µL of reaction mixture to the wells, excluding the standard curve.
  8. Cover the plate with a plate sealer and incubate at 37 °C for 20 minutes.
  9. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  10. Add 100 µL of deionized water to all wells. Mix briefly.
  11. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate for 20 minutes at room temperature.
  12. Read plate at 620 nm (absorbance) in endpoint mode.
  13. 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:
  • rhSULT1B1: 1.0 µg
  • PAPS: 10000 pmol (0.2 mM)
  • 1-Naphthol: 0.2 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 SULT1B1 Protein, CF

• Cytosolic Sulfotransferase 1B1
• EC 2.8.2
• EC 2.8.2.-
• EC 2.8.2.1
• EC 2.8.2.4
• MGC13356
• ST1B1
• ST1B2
• ST1B2sulfotransferase family cytosolic 1B member 1
• Sulfotransferase 1B1
• Sulfotransferase 1B2
• sulfotransferase family, cytosolic, 1B, member 1
• SULT1B1
• SULT1B2
• Thyroid hormone sulfotransferase

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 functions 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. Cytosolic sulfotransferases are distinct from Golgi resident sulfotransferases by lacking N-terminal signal-anchorage domains and residing only in the cytoplasm. SULT1B1 is primarily expressed in the liver, peripheral blood leukocytes, colon, spleen and small intestine and can sulfate thyroid hormones and small phenols (4). Human SULT1B1 shows 72.3% amino acid sequence identity to mouse SULT1B1.The enzymatic activity of the recombinant human GAL3ST2 is measured using a phosphatase-coupled assay (4).

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. Wang, J. et al. (1998) Mol. Pharmacol. 53:274.
5. Prather, B. et al. (2012) Anal. Biochem. 423:86.

Publications for Cytosolic Sulfotransferase 1B1/SULT1B1 (5959-ST)(1)

Publication using 5959-ST

• C Xie, TM Yan, JM Chen, XY Li, J Zou, LJ Zhu, LL Lu, Y Wang, FY Zhou, ZQ Liu, M Hu LC-MS/MS quantification of sulfotransferases is better than conventional immunogenic methods in determining human liver SULT activities: implication in precision medicine Sci Rep, 2017-06-20;7(1):3858. 2017-06-20 [PMID: 28634336] (Enzyme Assay, Human)

Bioinformatics

• Gene Symbol: SULT1B1
• Uniprot:
  • O43704()