Recombinant Mouse Sulfamidase/SGSH Protein, CF

• Reactivity: Mu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Mouse Sulfamidase/SGSH Protein, CF Summary

• Details of Functionality: Measured 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.
• Source: Spodoptera frugiperda, Sf 21 (baculovirus)-derived mouse Sulfamidase/SGSH protein
  Arg23-Leu502, with a C-terminal 10-His tag
• Accession #: NP_061292
• N-terminal Sequence: Arg23
• Protein/Peptide Type: Recombinant Enzymes
• Gene: Sgsh
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Theoretical MW: 56 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 60 kDa, reducing conditions

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.
• Buffer: Supplied as a 0.2 μm filtered solution in Tris and NaCl.
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain
• Assay Procedure:
  • 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
  1. Dilute rmSGSH to 40 µg/mL in Assay Buffer.
  2. Dilute Substrate to 20 mM in Assay Buffer.
  3. 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.
  4. Incubate at 37 °C for 72 hours.
  5. Stop reaction by adding 300 µL of 0.2 M NaOH to the reaction vials.
  6. Load in a plate 200 µL in duplicate from each reaction vial.
  7. Read at 510 nm (absorbance) in endpoint mode.
  8. 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

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Mouse Sulfamidase/SGSH Protein, CF

• heparan sulfate sulfatase
• HSS
• HSSN-sulphoglucosamine sulphohydrolase
• MPS3A
• MPS3Asulphamidase
• N-sulfoglucosamine sulfohydrolase
• SFMD
• SFMDEC 3.10.1.1
• SGSH
• Sulfamidase
• Sulfoglucosamine sulfamidase
• Sulphamidase

Background

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.

1. Diez-Roux, G. and Ballabio, A. (2005) Annu. Rev. Genomics Hum. Genet. 6:355.
2. Gliddon, B.L. et al. (2004) Mol. Genet. Metab. 83:239.

Publications for Sulfamidase/SGSH (2969-SU)(2)

Publications using 2969-SU

• E Makino, H Klodnitsky, J Leonard, J Lillie, TC Lund, J Marshall, J Nietupski, PJ Orchard, WP Miller, C Phaneuf, D Tietz, ML Varban, M Donovan, A Belenki Fast, sensitive method for trisaccharide biomarker detection in mucopolysaccharidosis type 1 Sci Rep, 2018;8(1):3681. 2018 [PMID: 29487322] (Bioassay, Mouse)
• Lamanna WC, Lawrence R, Sarrazin S, Esko JD Secondary storage of dermatan sulfate in Sanfilippo disease. J. Biol. Chem., 2011;286(9):6955-62. 2011 [PMID: 21193389] (Bioassay, Human)

Bioinformatics

• Gene Symbol: Sgsh
• Uniprot:
  • NP_061292()