Recombinant Human SULT1B1 Protein, CF Summary
Leu2-Ile296, with an N-terminal 6-His tag
Accession # O43704
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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.
|Formulation||Supplied as a 0.2 μm filtered solution in Tris, NaCl, Glycerol and DTT.|
|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 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
- 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.
- 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.
- Prepare reaction mixture containing 0.4 mM PAPS, 0.4 mM 1-Naphthol, and 0.02 mg/mL Coupling Phosphatase 3 in Assay Buffer.
- Dilute rhSULT1B1 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 rhSULT1B1 into the plate. Include 25 µL of Assay Buffer to be used as a Control.
- Add 25 µL of reaction mixture to the wells, excluding the standard curve.
- Cover the plate with 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:
- rhSULT1B1: 1.0 µg
- PAPS: 10000 pmol (0.2 mM)
- 1-Naphthol: 0.2 mM
- Coupling Phosphatase 3: 0.5 µg
Background: Cytosolic Sulfotransferase 1B1/SULT1B1
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).
- Falany, C, N. (1997) FASEB J. 11:206.
- Gamage, N. U. et al. (2006) Toxicol. Sci. 90:5.
- Allali-Hassani, A. et al. (2007) PLoS Biol. 5:e97.
- Wang, J. et al. (1998) Mol. Pharmacol. 53:274.
- Prather, B. et al. (2012) Anal. Biochem. 423:86.
Citation for Recombinant Human SULT1B1 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
LC-MS/MS quantification of sulfotransferases is better than conventional immunogenic methods in determining human liver SULT activities: implication in precision medicine
Authors: C Xie, TM Yan, JM Chen, XY Li, J Zou, LJ Zhu, LL Lu, Y Wang, FY Zhou, ZQ Liu, M Hu
Sci Rep, 2017;7(1):3858.
Sample Types: Protein
Applications: Enzyme Assay
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Sulfotransferase Assays and Substrates
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