Recombinant P. heparinus Heparinase II Protein, CF

R&D Systems | Catalog # 6336-GH

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Key Product Details

  • R&D Systems E. coli-derived Recombinant P. heparinus Heparinase II Protein (6336-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

E. coli

Accession Number

YP_003092674

Applications

Enzyme Activity
View all Heparinase II Proteins and Enzymes »

Product Specifications

Source

E. coli-derived p. heparinus Heparinase II protein
Ala26-Arg772, with an N-terminal Met and 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

Met

Predicted Molecular Mass

86 kDa

SDS-PAGE

66-75 kDa, reducing conditions

Activity

Measured by its ability to liberate oligosaccharides from heparin.
The specific activity is >750 pmol/min/μg, as measured under the described conditions.

Scientific Data Images for Recombinant P. heparinus Heparinase II Protein, CF

Recombinant P. heparinus Heparinase II Protein Enzyme Activity

Recombinant P. heparinus Heparinase II Protein Enzyme Activity

Heparinase II digestion of Heparin Sulfate (200 μg) is assessed in a 5-minute kinetic assay by monitoring absorbance at 232 nm. R&D SystemsP. heparinusHeparinase I (catalog # 6336-GH) exhibits activity at 2595.6 pm/min/μg.

Formulation, Preparation, and Storage

6336-GH
Formulation Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Shipping The product is shipped with dry ice or equivalent. 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, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Background: Heparinase II

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).

References

  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.

UniProt

YP_003092674

Additional Heparinase II Products

Product Documents for Recombinant P. heparinus Heparinase II Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant P. heparinus Heparinase II Protein, CF

For research use only

Citations for Recombinant P. heparinus Heparinase II Protein, CF

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Protocols

View specific protocols for Recombinant P. heparinus Heparinase II Protein, CF (6336-GH):

Materials
  • 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






















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