Recombinant Human SULT1E1 Protein, CF
Recombinant Human SULT1E1 Protein, CF Summary
Product Specifications
Asn2-Ile294, with an N-terminal 6-His tag
Analysis
Product Datasheets
Carrier Free
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.
5545-ST
| Formulation | Supplied as a 0.2 μm filtered solution in Tris, NaCl 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 Procedure
- Universal Sulfotransferase Activity Kit (Catalog # EA003)
- 10X Assay Buffer (supplied in kit): 500 mM Tris, 150 mM MgCl2, pH 7.5
- Recombinant human SULT-1E1 (rhSULT-1E1) (Catalog # 5545-ST)
- 3'-Phosphoadenosine-5'-phosphosulfate (PAPS) (Catalog # ES019)
- 1-Naphthol (Sigma, Catalog # N1000), 10 mM in deionized water
- 96-well Clear Plate (Catalog # DY990)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Prepare 1X Assay Buffer by diluting 10X stock 10-fold with deionized water.
- 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 1X 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 1X Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
- Prepare a reaction mixture containing 0.4 mM PAPS, 0.4 mM 1-Naphthol, and 0.02 mg/mL Coupling Phosphatase 3 in 1X Assay Buffer.
- Dilute rhSULT-1E1 to 20 µg/mL in 1X Assay Buffer.
- Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of 1X Assay Buffer.
- Load 25 µL of the 20 µg/mL rhSULT-1E1 into the plate. Include a Control containing 25µL of 1X Assay Buffer.
- Add 25 µL of reaction mixture to the wells, excluding the standard curve.
- Seal plate 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 sealed plate 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 fitting and adjusted for Control.
Per Reaction:- rhSULT-1E1: 0.5 µg
- PAPS: 10 nmoles (0.2 mM)
- 1-Naphthol: 0.2 mM
- Coupling Phosphatase 3: 0.5 µg
Reconstitution Calculator
Background: Cytosolic Sulfotransferase 1E1/SULT1E1
Cytosolic Sulfotransferases catalyze the sulfation of many hormones, neurotransmitters, drugs, and xenobiotic compounds. They are distinct from Golgi resident sulfotransferases by the absence of transmembrane domains and are located in the cytoplasm. SULT1E1 is widely known as an estrogen sulfotransferase and may control estrogen levels by converting free estradiol to its inactive sulfate conjugate (1). Known substrates of this enzyme also include dehydroepiandrosterone, pregnenolone, ethinylestradiol, equalenin, diethylstilbesterol, 4-nitrophenol, 1-napthol and resveratrol (2, 3). SULT1E1 activity is found in reproductive organs (4), intestine and liver (3, 5). The enzymatic activity of the recombinant human SULT1E1 is measured using a phosphatasecoupled assay (6).
- Gong, H. et al. (2008) Cancer Res. 68:7386.
- Falany, C.N. et al. (1995) J. Steroid Biochem. Mol. Biol. 52:529.
- Miksits, M. et al. 2005, Xenobiotica 35:1101.
- Takase, Y. et al. (2007) The Prostate 67:405.
- Schrag, M.L. et al. (2004) Drug Metabolism and Disposition 32:1299.
- Prather, B. et al. (2012) Anal. Biochem. 423:86.
Citations for Recombinant Human SULT1E1 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.
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Citations: Showing 1 - 2
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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, 2017-06-20;7(1):3858.
Species: Human
Sample Types: Protein
Applications: Enzyme Assay -
A versatile polyacrylamide gel electrophoresis based sulfotransferase assay.
Authors: Wu ZL, Ethen CM, Larson S, Prather B, Jiang W
BMC Biotechnol., 2010-02-10;10(0):11.
Applications: Enzyme Assay
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