Recombinant Human Glucosamine (N-acetyl)-6-Sulfatase, CF Summary
Val37-Leu552 & Thr44-Leu552, both with a C-terminal 10-His tag
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 and Glycerol.|
|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: 50 mM Sodium Acetate, 250 mM NaCl, pH 5.0
- Recombinant Human Glucosamine (N-acetyl)‑6-Sulfatase/GNS (rhGNS) (Catalog # 2484-SUC)
- Substrate: 4-Nitrocatechol Sulfate (PNCS) (Sigma, Catalog # N-7251), 100 mM stock in deionized water
- 0.2 M NaOH
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute rhGNS to 2 µg/mL in Assay Buffer.
- Dilute PNCS substrate to 4 mM in Assay Buffer.
- Combine 75 µL of 2 µg/mL rhGNS and 75 µL 4 mM PNCS. Include a Substrate Blank containing Assay Buffer in place of rhGNS.
- Incubate at 37 °C for 2 hours.
- Stop reactions and develop color by adding 150 µL of 0.2 M NaOH to each reaction.
- Load into the wells of a clear 96-well plate 200 µL from each reaction.
- Read at 510 nm (absorbance) in endpoint mode.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
|Adjusted Abs* (OD) x Conversion Factor** (pmole/OD)|
|Incubation time (min) x amount of enzyme (µg)|
*Adjusted for Substrate Blank
**Derived using calibration standard p-Nitrocatechol (Sigma, Catalog # N15553).Per Well:
- rhGNS: 0.1 µg
- Substrate: 1 mM
Background: Glucosamine (N-acetyl)-6-Sulfatase/GNS
A member of the sulfatase family, GNS is required for the lysosomal degradation of the glycosaminoglycans (GAG) heparan sulfate and keratan sulfate (1, 2). It hydrolyzes the 6-sulfate group of the N-acetyl-D-glucosamine 6-sulfate units of the GAG. GNS deficiency results in mucopolysaccharidosis type IIID (MPS IIID or Sanfilippo D Syndrome), an inborn error leading to lysosomal accumulation of heparan sulfate. MPS IIID has profound mental deterioration, hyperactivity, and relatively mild somatic manifestations. The deduced amino acid sequence of human GNS consists of a signal peptide (residues 1-36) and a mature chain (residues 37-552) that may be further processed into N-terminal and C-terminal fragments (3). rhGNS corresponds to the single chain and has sulfatase activity described in Activity Assay Protocol.
- Parenti, G. et al. (1997) Curr. Opin. Genet. & Dev. 7:386.
- Neufeld, E.F. and J. Muenzer (2001) in The Metabolic and Molecular Basis of Inherited Disease, Scriver, C.R. et al. (eds.) pp. 3421, New York, McGraw-Hill.
- Robertson, D.A. et al. (1992) Biochem. J. 288:539.
Citation for Recombinant Human Glucosamine (N-acetyl)-6-Sulfatase, 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.
1 Citation: Showing 1 - 1
Oligosaccharides mapping of nitrous acid degraded heparin through UHPLC-HILIC/WAX-MS
Authors: T Zhang, S Xie, Z Wang, R Zhang, Q Sun, X Liu, L Chi, JP Li, H Li, T Tan
Carbohydr Polym, 2020;231(0):115695.
Sample Types: Whole Cells
Applications: Cell Culture
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Reviews for Recombinant Human Glucosamine (N-acetyl)-6-Sulfatase, CF
Average Rating: 5 (Based on 1 Review)
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Reason for Rating: Provides very reproducible activity measurements along with long-term stability - serves as an ideal positive control for both activity assays and SDS-PAGE applications. Did not demonstrate in-vivo intracellular uptake.