详细说明
- Purity>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
- Endotoxin Level<1.0 EU per 1 μg of the protein by the LAL method.
- ActivityMeasured by its ability to hydrolyze the substrate 4-Nitrocatechol Sulfate (PNCS). The specific activity is >1.0 pmol/min/µg, as measured under the described conditions. See Activity Assay Protocol on .
- SourceMouse myeloma cell line, NS0-derived Thr36-Pro552 with an N-terminal signal sequence and a C-terminal 10-His tag
- Accession #
- N-terminal Sequence
AnalysisThr36 - Predicted Molecular Mass60 kDa
- SDS-PAGE77 kDa, reducing conditions
| 2486-SU | | |
| 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.
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Assay Procedure
Materials
- Assay Buffer: 50 mM NaOAc, pH 5.0
- Recombinant Mouse Iduronate 2‑Sulfatase/IDS (rmIDS) (Catalog # 2486-SU)
- Substrate: 4-Nitrocatechol Sulfate (4-PNCS) (Sigma, Catalog # N-7251)
- Sodium Hydroxide (NaOH) (Sigma, Catalog # S-0899)
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate Reader (Model: Spectra-Max Plus by Molecular Devices) or equivalent
- Dilute rmIDS to 20 µg/mL in Assay Buffer.
- Dilute Substrate to 2 mM in Assay Buffer.
- Combine 150 µL of 20 µg/mL rmIDS and 150 µL of 2 mM Substrate. Include a Substrate Blank containing Assay Buffer and Substrate only.
- Incubate at 37 °C for 24 hours.
- Stop reaction by adding 300 µL of 0.2 M NaOH to reaction tubes.
- Load 200 µL from each reaction tube in duplicate into a plate.
- Read at 510 nm (absorbance) in endpoint mode.
- Calculate specific activity:
| Specific Activity (pmol/min/µg) = | Adjusted Abs* (OD) x Conversion Factor** (pmol/OD) |
| Incubation time (min) x amount of enzyme (µg) |
*Adjusted for Substrate Blank
**Derived using calibration standard P-Nitrocatechol (PNC) (Sigma, Catalog # N15553).
- rmIDS: 1.0 µg
- Substrate: 1 mM
Background: Iduronate 2-Sulfatase/IDS
As a member of the sulfatase family, Iduronate 2-Sulfatase encoded by the IDS gene is required for the lysosomal degradation of the glycosaminoglycans (GAG) heparan sulfate and dermatan sulfate (1, 2). It hydrolyzes the 2-sulfate group of the L-iduronate 2-sulfate units of the GAG. The IDS deficiency results in mucopolysaccharidosis II (MPS II or Hunter syndrome), an X-linked inborn error leading to lysosomal accumulation of the GAG and their excretion in urine. MPS II has a wide spectrum of clinical manifestations ranging from mild to severe. The mouse IDS has sulfatase activity described in the Activity Assay Protocol. In addition, recombinant human IDS is also available (Catalog # ).
- References:
- 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.) p. 3421, New York, McGraw-Hill.
- Entrez Gene IDs:3423 (Human); 15931 (Mouse)
- Alternate Names:Alpha-L-iduronate sulfate sulfatase; EC 3.1.6.13; IDS; iduronate 2-sulfatase 14 kDa chain; iduronate 2-sulfatase 42 kDa chain; iduronate 2-sulfatase; idursulfase; MPS2; S; SIDS
400-6226-992