详细说明
- Purity>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
- 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 >0.4 pmol/min/µg, as measured under the described conditions. See Activity Assay Protocol on .
- SourceSpodoptera frugiperda, Sf 21 (baculovirus)-derived Arg23-Leu502, with a C-terminal 10-His tag
- Accession #
- N-terminal Sequence
AnalysisArg23 - Predicted Molecular Mass56 kDa
- SDS-PAGE60 kDa, reducing conditions
| 2969-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 Buffer: 50 mM MES, pH 5.5
- Recombinant Mouse Sulfamidase/SGSH (mSGSH) (Catalog # 2969-SU)
- Substrate: 4-Nitrocatechol Sulfate (PNCS) (Sigma, Catalog # N-7251), 100 mM stock in deionized water
- Sodium Hydroxide (NaOH) (Sigma, Catalog # 221465), 2 M stock in deionized water
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute rmSGSH to 40 µg/mL in Assay Buffer.
- Dilute Substrate to 20 mM in Assay Buffer.
- Mix 150 µL of 40 µg/mL of rmSGSH and 150 µL of 20 mM of Substrate. Include a Substrate Blank with 150 µL of Assay Buffer and 150 µL of Substrate.
- Incubate at 37 °C for 72 hours.
- Stop reaction by adding 300 µL of 0.2 M NaOH to the reaction vials.
- Load in a plate 200 µL in duplicate from each reaction vial.
- 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).
Per Well:- rmSGSH: 2 µg
- Substrate: 5 mM
Also known as N-sulfoglucosamine sulfohydrolase and heparan N-sulfatase, Sulfamidase/SGSH is an important member of the sulfatase family involved in the degradation of heparan sulfate (1). The SGSH deficiency results in mucopolysaccharidosis type IIIA (MPS IIIA, Sanfilippo A syndrome), an autosomal recessive lysosomal storage disease characterized by neurological dysfunction but relatively mild somatic manifestations (2). The deduced amino acid sequence of mouse SGSH consists of a signal peptide (residues 1-22) and a mature chain (residues 23-502) (1). Recombinant Mouse Sulfamidase/SGSH corresponds to the mature chain and has sulfatase activity as described in the Activity section.
- References:
- Diez-Roux, G. and Ballabio, A. (2005) Annu. Rev. Genomics Hum. Genet. 6:355.
- Gliddon, B.L. et al. (2004) Mol. Genet. Metab. 83:239.
- Entrez Gene IDs:6448 (Human); 27029 (Mouse)
- Alternate Names:heparan sulfate sulfatase; HSS; HSSN-sulphoglucosamine sulphohydrolase; MPS3A; MPS3Asulphamidase; N-sulfoglucosamine sulfohydrolase; SFMD; SFMDEC 3.10.1.1; SGSH; Sulfamidase; Sulfoglucosamine sulfamidase; Sulphamidase
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