Recombinant Human GSTP1 Protein, CF

Catalog # Availability Size / Price Qty
6455-GT-010

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Recombinant Human GSTP1 Protein, CF Summary

Product Specifications

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by the conjugation of reduced glutathione to 1-bromo-2,4-dinitrobenzene. The specific activity is >25,000 pmol/min/μg, as measured under the described conditions.
Source
E. coli-derived human Glutathione S-Transferase pi 1/GSTP1 protein
Met1-Glu210
Accession #
N-terminal Sequence
Analysis
Met
Predicted Molecular Mass
23 kDa
SDS-PAGE
24 kDa, reducing conditions

Product Datasheets

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6455-GT

Carrier Free

What does CF mean?

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.

What formulation is right for me?

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.

6455-GT

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
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.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Assay Buffer: 100 mM NaH2PO4, pH 7.0
  • Recombinant Human Glutathione S‑Transferase pi 1/GSTP1 (rhGSTP1) (Catalog # 6455-GT)
  • Substrate: 1-bromo-2,4-dinitrobenzene (BDNB) (Sigma, Catalog # 262226), 75 mM stock in ethanol
  • L-Glutathione, reduced (GSH) (Amresco, Catalog # 399), 250 mM stock in deionized water
  • UV Plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhGSTP1 to 0.2 ng/μL in Assay Buffer.
  2. Dilute GSH to 4 mM in Assay Buffer.
  3. Combine equal volumes of 0.2 ng/μL rhGSTP1 and 4 mM GSH for 0.1 ng/μL rhGSTP1 with 2 mM GSH.
  4. Dilute Substrate to 2 mM in Assay Buffer.
  5. Load into a UV plate 50 μL of the rhGSTP1/GSH mixture. Include a substrate blank containing 25 μL of Assay Buffer with 25 μL of the 4 mM GSH prepared in step 2.
  6. Start the reaction by adding 50 μL of 2 mM Substrate to well.
  7. Read in kinetic mode for 5 minutes at an absorbance of 340 nm.
  8. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank 
     **Using the extinction coefficient 9600 M-1cm-1 
     ***Using the path correction 0.320 cm
     Note: the output of many spectrophotometers is in mOD. Per Well:
  • rhGSTP1: 0.005 μg
  • GSH: 1 mM
  • Substrate: 1 mM
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Background: Glutathione S-Transferase pi 1/GSTP1

Glutathione S-Transferases (GSTs) are members of the phase II detoxification enzyme family that conjugate glutathione to various electrophilic compounds, including metabolites generated by oxidative processes in the body, environmental toxins or carcinogens, and anti-cancer drugs. GSTP1 is a cytosolic protein that belongs to pi class of the GST superfamily. It is crystallized as a homodimer (1), but also exists in solution as an equilibrium mixture of monomer and dimer, depending on the protein concentration (2). Four genetic variants of GSTP1 with different enzymatic activities have been identified, which indicates the particular allelic form expressed in tissues could contribute to variation in catalytic efficiency and biological functions (3, 4). Human GSTP1 is present at elevated levels in many tumor cells, and has unique properties as a cancer marker (5). Genetic polymorphisms and expression patterns of GSTP1 have been associated with a variety of effects on human cancer, anti-cancer drug resistance, and asthma (6). In addition to its role as a drug-metabolizing enzyme, GSTP1 has ligand binding properties and regulates kinase signaling pathways through protein-protein interactions (7).

References
  1. Reinemer, P. et al. (1992) J. Mol. Biol. 227:214.
  2. Huang, Y.C. et al. (2008) J. Biol. Chem. 283:32880.
  3. Ali-Osman, F. et al. (1997) J. Biol. Chem. 272:10004.
  4. Hu, X. et al. (1998) Cancer Res. 58:5340.
  5. Sato, K. et al. (1992) Tohoku J. Exp. Med. 168:97.
  6. Townsend, D.M. and K.D. Tew (2003) Oncogene 22:7369.
  7. Adler, V. et al. (1999) EMBO J. 18:1321.
Entrez Gene IDs
2950 (Human)
Alternate Names
deafness, X-linked 7; DFN7; EC 2.5.1.18; FAEES3; fatty acid ethyl ester synthase III; glutathione S-transferase P; Glutathione STransferase pi 1; Glutathione S-Transferase pi 1; GST class-pi; GST3DFN7; GSTP; GSTP1; GSTP1-1; PI

Product Specific Notices

Coomassie is a registered trademark of Imperial Chemical Industries Ltd.

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