Recombinant Human GSTM1 Protein, CF Summary
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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, DTT 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.
- Assay Buffer: 50 mM Sodium Phosphate, pH 7.0
- Recombinant Human Glutathione S‑Transferase mu 1/GS (rhGSTM1) (Catalog # 6894-GT)
- L-Glutathione, reduced (GSH) (Amresco, Catalog # 0399), 250 mM stock in deionized water
- Substrate: 1-Bromo-2,4-dinitrobenzene (BDNB) (Sigma, Catalog # 262226), 75 mM stock in ethanol
- UV plate (Costar, Catalog # 3635)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute GSH to 1 mM in Assay Buffer.
- Dilute rhGSTM1 to 0.2 ng/μL in Assay Buffer containing 1 mM GSH.
- Dilute Substrate to 2 mM in Assay Buffer containing 1 mM GSH.
- Load into a plate 50 μL of 0.2 ng/μL rhGSTM1 and start the reaction by adding 50 μL of 2 mM Substrate. For Substrate Blanks, load 50 μL of Assay Buffer containing 1 mM GSH and 50 μL of 2 mM Substrate.
- Read plate at a wavelength of 340 nm (bottom read) in kinetic mode for 5 minutes.
- 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 0.32 cm
Note: the output of many spectrophotometers is in mOD. Per Well:
- rhGSTM1: 0.010 μg
- Substrate: 1 mM
- GSH: 1 mM
Background: Glutathione S-Transferase mu 1/GSTM1
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. GSTM1 is a cytosolic protein that belongs to the mu class of the GST superfamily. The gene encoding GSTM1 is mapped onto human chromosome 1p13.3 and is known to be highly polymorphic (1). Mostly notably, the widely occurring GSTM1-null genotype has been linked to a variety of cancers including lung (2), gastric (3), bladder (4) and prostate (5). In addition to its role in releasing oxidative stress, GSTM1 has also been suggested to act as a hormone binding protein and play a role in maintaining hormone homeostasis in the body (6, 7).
- Pearson, W. R. et al. (1993) Am. J. Hum. Genet. 53:220.
- Mohr, L.C. et al. (2003) Anticancer. Res. 23:2111.
- Wang, H. et al. (2010) Dig. Dis. Sci. 55:1824.
- Engel, L. S. et al. (2002) Am. J. Epidermiol. 156:95.
- Mo, Z. et al. (2009) Prostate. 69:662.
- Mukherjee, S. B. et al. (1999) Biochem. J. 340:309.
- Ishigaki, S. et al. (1989) Arch. Biochem. Biophys. 273:265.
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