Recombinant Human Tissue alpha-L-Fucosidase/FUCA1, CF
Recombinant Human Tissue alpha-L-Fucosidase/FUCA1, CF Summary
Learn more about Fluorescent Glycan Labeling and DetectionProduct Specifications
Gln32-Lys466 with a C-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.
7039-GH
| 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 Procedure
- Assay Buffer: 50 mM Sodium Acetate, 5 mM MgCl2, pH 4.5
- Recombinant Human Tissue alpha ‑L‑Fucosidase/FUCA1 (rhFUCA1) (Catalog # 7039-GH)
- Substrate: 4-Methylumbelliferyl-alpha -L-fucopyranoside (Research Products International Corp, Catalog # M65200), 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 rhFUCA1 to 2 ng/μL in Assay Buffer.
- Dilute Substrate to 1.6 mM in Assay Buffer.
- Load into a plate 50 μL of 2 ng/μL rhFUCA1, and start the reaction by adding 50 μL of 1.6 mM Substrate. For Substrate Blanks, load 50 μL of Assay Buffer and 50 μL of 1.6 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).
- rhFUCA1: 0.100 μg
- Substrate: 0.8 mM
Reconstitution Calculator
Background: Tissue alpha-L-Fucosidase/FUCA1
Fucosylated glycoconjugates play numerous roles in biological processes, such as development, apoptosis and the immune responses, and are involved in the pathology of inflammation, cancer and cystic fibrosis (1, 2, 3, 4). Tissue alpha ‑L‑Fucosidase, also known as FUCA1, is a lysosomal enzyme responsible for hydrolyzing alpha ‑L‑fucose moieties from glycolipids and oligosaccharides (5). The mature enzyme in the liver has a molecular weight of 200 kDa and exists in the native state as a tetramer (6). Defects in FUCA1 are the cause of fucosidosis (7, 8, 9), an autosomal recessive lysosomal storage disease characterized by the accumulation of fucose-containing glycolipids and glycoproteins in various tissues. Serum alpha ‑L‑fucosidase has been identified as a useful marker for oral cancer (10).
- Hiraishi, K. et al. (1993) Glycobiology 3:381.
- Solter, D. and Knowles, B.B. (1978) Proc. Natl. Acad. Sci. USA 75:5565.
- Becker, D.J. and Lowe, J.B. (2003) Glycobiology 13:41R.
- Liu, T.W. et al. (2009) Proc. Natl. Acad. Sci. USA 106:14581.
- Fukushima, H. et al. (1985) Proc. Natl. Acad. Sci. USA 82:1262.
- Alhadeff, J.A. et al. (1975) J. Biol. Chem. 250:7106.
- O’Brien, J.S. et al. (1987) Enzyme 38:45.
- Durand, P. et al. (1969) J. Pediatr. 75:665.
- Johnson, K. and Dawson, G. (1985) Biochem. Biophys. Res. Commun. 133:90.
- Shah, M. et al. (2008) Cancer 113:336.
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