Recombinant Human Hexosaminidase A/HEXA Protein, CF

R&D Systems | Catalog # 6237-GH

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Key Product Details

  • R&D Systems Sf 21 (baculovirus)-derived Recombinant Human Hexosaminidase A/HEXA Protein (6237-GH)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

Sf 21 (baculovirus)

Accession Number

P06865

Applications

Enzyme Activity
View all Hexosaminidase A/HEXA Proteins and Enzymes »

Product Specifications

Source

Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Hexosaminidase A/HEXA protein
Met1-Thr529, with a C-terminal 6-His tag

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.

N-terminal Sequence Analysis

Leu23

Predicted Molecular Mass

59 kDa

SDS-PAGE

57-61 kDa, reducing conditions

Activity

Measured by its ability to hydrolyze 4-methylumbelliferyl-N-acetyl-beta -D-glucosaminide (4-MU-GlcNAc)
The specific activity is >1,250 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

6237-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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Background: Hexosaminidase A/HEXA

beta -hexosaminidases are enzymes involved in the hydrolysis of terminal N-acetyl-D-hexosamine residues in GM2 gangliosides and globo-sphingolipids in lysosomes
(1‑4). The enzymes are composed of two alpha and/or beta  subunits, which are coded by HEXA and HEXB genes, respectively. Different association of the alpha and beta subunits gives rise to beta ‑hexosaminidase isoforms A, B and S (Hex A, B and S) (5), which have the composition of alpha beta, beta beta and alpha alpha, respectively. Our recombinant HEXA is presumably isoform Hex S, because only alpha subunit was expressed. Hex S is suggested to releases non‑reducing end N-acetylgalactosamine residues from dermatan sulfate, chondroitin sulfate and sulfated glycolipid SM2 (6). Recombinant HEXA is also highly active on 4-methylumbelliferyl-N-acetyl-beta -D-glucosaminide (6). Mutations in HEXA and HEXB genes cause lysosomal lipid storage disorders. Specifically, mutations of HEXA cause Tay-Sachs disease, manifested by the harmful accumulation of ganglioside GM2 in tissues and nerve cells in the brain (7‑10). Children with this disease usually die by age 4.

References

  1. Gilbert, F. et al. (1975) Proc. Natl. Acad. Sci. USA 72:263.
  2. Myerowitz, R. et al. (1985) Proc. Natl. Acad. Sci. USA 82:7830.
  3. Korneluk, R.G. et al. (1986) J. Biol. Chem. 261:8407.
  4. Mark, B.L. et al. (2003) J. Mol. Biol. 327:1093.
  5. Mahuran, D.J. et al. (1988) J. Biol. Chem. 263:4612.
  6. Hepbildikler, S.T. et al. (2002) J. Biol. Chem. 277:2562.
  7. Mahuran, D.J. (1991) Biochim. Biophys. Acta 1096:87.
  8. Mencarelli, S. et al. (2005) FEBS Lett. 579:5501.
  9. Neufeld, E.F. (1989) J. Biol. Chem. 264:10927.
  10. Ohno, K. et al. (2008) Mol. Genet. Metab. 94:462.

Alternate Names

HEXA, TSD

Entrez Gene IDs

3073 (Human)

Gene Symbol

HEXA

UniProt

P06865

Additional Hexosaminidase A/HEXA Products

Product Documents for Recombinant Human Hexosaminidase A/HEXA Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant Human Hexosaminidase A/HEXA Protein, CF

For research use only

Citations for Recombinant Human Hexosaminidase A/HEXA Protein, CF

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Protocols

View specific protocols for Recombinant Human Hexosaminidase A/HEXA Protein, CF (6237-GH):

Materials
  • Assay Buffer: 100 mM Sodium Citrate, 250 mM NaCl, pH 4.5
  • Recombinant Human Hexosaminidase A/HEXA (rhHEXA) (Catalog # 6237-GH)
  • Substrate: 4-Methylumbelliferyl-N-Acetyl-beta -D-glucosaminide (4-MU-GlcNAc) (Sigma, Catalog # M2133), 50 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhHEXA to 5 ng/μL in Assay Buffer.
  2. Dilute Substrate to 800 μM in Assay Buffer.
  3. Load into a plate 50 μL of 5 ng/μL rhHEXA, and start the reaction by adding 50 μL of 800 μM Substrate. For Substrate Blanks, load 50 μL of Assay Buffer and 50 μL of 800 μM Substrate.
  4. Read plate at excitation and emission wavelengths of 365 nm and 445 nm, respectively, in kinetic mode for 5 minutes.
  5. 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).

Per Well:
  • rhHEXA: 0.250 μg
  • Substrate: 400 μM

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