Recombinant Human alpha-L-Iduronidase/IDUA Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

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Recombinant Human alpha-L-Iduronidase/IDUA Protein, CF Summary

Details of Functionality
Measured by its ability to cleave a fluorogenic substrate, 4-Methylumbelliferyl  alpha -L-iduronide. The specific activity is >7,500 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human alpha-L-Iduronidase/IDUA protein
Ala26-Pro653 (Ala26Thr), with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Thr26
Protein/Peptide Type
Recombinant Enzymes
Gene
IDUA
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
71 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
83 kDa, reducing conditions
Publications
Read Publications using
4119-GH in the following applications:

Packaging, Storage & Formulations

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.
Buffer
Supplied as a 0.2 μm filtered solution in Sodium Acetate, NaCl and Glycerol.
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Assay Procedure
  • Assay Buffer: 50 mM NaOAc, 150 mM NaCl, 0.02% Brij-35 (w/v) pH 3.5
  • Developing Buffer: 0.1 M Tris, pH 9.0
  • Recombinant Human alpha ‑L‑Iduronidase/IDUA (rhIDUA) (Catalog # 4119-GH)
  • Substrate: 4-methylumberlliferyl-alpha -L-iduronide (Glycosynth, Catalog # 44076), 20 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 rhIDUA to 0.2 µg/mL in Assay Buffer. Minimize the number of dilution steps to obtain the best activity results.
  2. Dilute Substrate to 200 µM in Assay Buffer.
  3. Combine equal volumes of 0.2 µg/mL rhIDUA and 200 µM Substrate. Include a Substrate Blank containing Assay Buffer and Substrate.
  4. Incubate for 10 minutes at room temperature.
  5. Dilute mixtures to 0.005 µg/mL in Developing Buffer.
  6. Load 100 µL of the diluted mixtures into a plate.
  7. Read at excitation and emission wavelengths of 365 nm and 445 nm (top read), respectively in endpoint mode.
  8. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Fluorescence* (RFU) x Conversion Factor** (pmol/RFU)
Incubation time (min) x amount of enzyme (µg)

     *Adjusted for Substrate Blank

     **Derived using calibration standard 4-methylumbelliferone (Sigma, Catalog # M1381).

Per Well:
  • rhIDUA: 0.0005 µg
  • Substrate: 5 µM

Notes

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

Alternate Names for Recombinant Human alpha-L-Iduronidase/IDUA Protein, CF

  • alphaLIduronidase
  • alpha-L-Iduronidase
  • IDA
  • IDUA

Background

alpha -L-Iduronidase encoded by the IDUA gene is an important enzyme required for the lysosomal degradation of glycosaminoglycans (GAGs). It hydrolyzes the non‑reducing terminal alpha -L-iduronic acid residues in GAGs including dermatan sulfate and heparan sulfate. Mutations in IDUA that result in enzymatic deficiency lead to the autosomal recessive disease mucopolysaccharidosis type I (MPS I) (1). MPS I causes progressive cellular, tissue and organ damage, and several clinical studies using enzyme replacement therapy have shown promising benefits (2).
  1. Scott, H.S. et al. (1995) Hum. Mutat. 6:288.
  2. Wraith, J.E. (2005) Expert Opin. Pharmacother. 6:489.

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Bioinformatics

Gene Symbol IDUA
Uniprot
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