Recombinant P. heparinus Heparinase II Protein, CF

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Heparinase II digestion of Heparin Sulfate (200 μg)is assessed in a 5-minute kinetic assay by monitoring absorbance at 232 nm. R&D Systems P. heparinus Heparinase I (catalog #6336-GH) exhibits activity at 2595.6 ...read more

Product Details

Summary
Reactivity PhSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

Order Details

Recombinant P. heparinus Heparinase II Protein, CF Summary

Additional Information
Get 50% off R&D Systems® Heparinase I, II, and III! Use Promo Code: Heparinase50
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Details of Functionality
Measured by its ability to liberate oligosaccharides from heparin. The specific activity is >750 pmol/min/μg, as measured under the described conditions.
Source
E. coli-derived p. heparinus Heparinase II protein
Ala26-Arg772, with an N-terminal Met and 6-His tag
Accession #
N-terminal Sequence
Met
Protein/Peptide Type
Recombinant Enzymes
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
86 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
66-75 kDa, reducing conditions
Publications
Read Publication using
6336-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, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.
Buffer
Supplied as a 0.2 μm filtered solution in PBS.
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Assay Procedure
  • Assay Buffer: 100 mM Tris, pH 7.5
  • Recombinant P. heparinus Heparinase II (rPhHeparinase II) (Catalog # 6336-GH)
  • Substrate: Heparin (Tocris, Catalog # 2812), 20 mg/mL stock in deionized water
  • 96 well clear UV-transparent microplate (Corning, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rPhHeparinase II to 20 ng/µL in Assay Buffer.
  2. Dilute Substrate to 3.0 mg/mL in Assay Buffer.
  3. Load into a plate 50 µL of the diluted rPhHeparinase II, and start the reaction by adding 50 µL of 3.0 mg/mL Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 3.0 mg/mL Substrate.
  4. Read in kinetic mode for 5 minutes at an absorbance of 232 nm.
  5. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank 
     **Using the extinction coefficient 3800 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rPhHeparinase II: 1.0 µg
  • Substrate: 1.5 mg/mL

Notes

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

Alternate Names for Recombinant P. heparinus Heparinase II Protein, CF

  • Heparinase II

Background

Heparan sulfate is a sulfated glycosaminoglycan with the repeating disaccharide units of ‑4HexA1,4GlcNAc beta 1-. It is usually attached to the protein cores of proteoglycans found on cell membrane and extracellular matrix, where it binds to a variety of protein ligands and regulates a wide range of biological activities, including developmental processes, angiogenesis, blood coagulation and tumor metastasis (1, 2). Heparan sulfate has a domain structure containing sulfated regions interspaced with less or non-sulfated regions (3, 4). Heparin shares the backbone structure with heparan sulfate but contains no non-sulfated regions. Heparinases are a family of lyases that release unsaturated oligosaccharides from heparin and heparan sulfate upon digestion (5). Heparinase I recognizes highly sulfated regions and is more specific for heparin. Heparinase II digests both heparin and heparan sulfate. Heparinase III prefers less-sulfated regions and is active only on heparan sulfate (6, 7).
  1. MacArthur, J. M. et al. (2007) J. Clin. Invest. 117:153.
  2. Esko, J. D. and Selleck, S. B. (2002) Annu. Rev. Biochem. 71:435.
  3. Maccarana, M. et al. (1996) J. Biol. Chem. 271:17804.
  4. Linker, A. and Hovingh, P. (1975) Biochim. Biophys. Acta. 385:324.
  5. Linker, A. and Hovingh, P. (1965) J. Biol. Chem. 240:3724.
  6. Su, H. et al. (1996) Appl. Environ. Microbiol. 62:2723.
  7. Hovingh, P. and Linker, A. (1970) J. Biol. Chem. 245:6170.

Publications for Heparinase II (6336-GH)(1)

We have publications tested in 1 confirmed species: Bovine.

We have publications tested in 1 application: Enzyme Assay.


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