Recombinant Human beta-Glucuronidase/GUSB Protein, CF Summary
Met1-Thr651, with a C-terminal 6-His tag
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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 and NaCl.|
|Shipping||The product is shipped with polar packs. 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: 100 mM Sodium Acetate, pH 3.5
- Recombinant Human beta ‑Glucuronidase/GUSB (rhGUSB) (Catalog # 6144-GH)
- Substrate: 4-Methylumbelliferyl beta -D-glucuronide (Sigma, Catalog # M9130), 50 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhGUSB to 4 ng/μL in Assay Buffer.
- Dilute Substrate to 2 mM in Assay Buffer.
- Load into a plate 50 μL of 4 ng/μL rhGUSB and start the reaction by adding 50 μL of 2 mM Substrate. For Substrate Blanks, load 50 μL of Assay Buffer and 50 μL of 2 mM Substrate.
- Read plate at excitation and emission wavelengths of 365 nm and 445 nm, respectively, in kinetic mode for 5 minutes.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
|Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU)|
|amount of enzyme (µg)|
*Adjusted for Substrate Blank
**Derived using calibration standard 4-Methylumbelliferone (4-MU) (Sigma, Catalog # M1381).
- rhGUSB: 0.200 μg
- Substrate: 1 mM
Human beta -Glucuronidase (EC 18.104.22.168) encoded by the GUSB gene is a lysosomal hydrolase involved in the stepwise degradation of glucuronic acid-containing glycosaminoglycans that include heparan sulfate, chondroitin sulfate and hyaluronan (1). The enzyme is only active on the glucuronic acid of the non-reducing end. The native protein has been reported as a tetrameric glycoprotein composed of identical subunits (1, 2). Mutations in the GUSB gene are linked to mucopolysaccharidosis type VII (3). Accumulation of partially degraded glycosaminoglycans, with glucuronic acid residues at the non-reducing termini, are usually found in the lysosomes of patients with the disease (3). It has also been reported that this enzyme may contribute to the depletion of chondroitin from cartilage and thereby facilitate the damage of joints in rheumatoid arthritis (4).
- Shipley, J.M. et al. (1993) Am. J. Hum. Genet. 52:517.
- Oshima, A, et al. (1987) Proc. Natl. Acad. Sci. USA 84:685.
- Bell, C.E. Jr. et al. (1977) J. Clin. Invest. 59:97.
- Ortutay, Z. et al. (2003) Arthritis Rheum. 48:2163.
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