Recombinant F. heparinum Heparinase I Protein, CF

R&D Systems | Catalog # 7897-GH

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Key Product Details

  • R&D Systems E. coli-derived Recombinant F. heparinum Heparinase I Protein (7897-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

Q05819

Applications

Enzyme Activity
View all Heparinase I Proteins and Enzymes »

Product Specifications

Source

E. coli-derived f. heparinum Heparinase I protein
Gln22-Arg384, 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

42 kDa

SDS-PAGE

40-42 kDa, reducing conditions

Activity

Measured by its ability to liberate oligosaccharides from heparin.
The specific activity is >24,500 pmol/min/μg, as measured under the described conditions.

Scientific Data Images for Recombinant F. heparinum Heparinase I Protein, CF

Recombinant F. heparinum Heparinase I Protein Enzyme Activity

Recombinant F. heparinum Heparinase I Protein Enzyme Activity

Heparinase I digestion of Heparin Sulfate (200 μg) is assessed in a 5-minute kinetic assay at room temperature by monitoring absorbance at 232 nm. R&D Systems RecombinantF. heparinumHeparinase I (Catalog # 6145-GH) (orange) exhibits greater activity than theBacteroidesHeparinase I from the competition (green).

Formulation, Preparation, and Storage

7897-GH
Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and DTT.
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 I

Heparin and heparan sulfate are sulfated glycosaminoglycans that share basic carbohydrate backbone structure with alternating uronic acid and N-acetylglucosamine residues (1, 2). Heparin is found in mast cells and has strong anticoagulation properties. Heparan sulfate is found on cell membrane and extracellular matrix and is involved in various biological events from cell growth, adhesion and migration to lipid metabolism. Heparin has a much higher degree of sulfation than heparan sulfate, which can be considered as a polysaccharide with regions similar to heparin interspaced with much less sulfated regions. Both heparin and heparan sulfate can be digested by heparinases, a group of bacterial lyases that are widely used as tools for processing and analyze these polysaccharides. Heparinases degrade heparin and heparan sulfate glycosaminoglycans through an eliminative mechanism (3). Heparinase I from Flavobacterium heparinum is highly active on heparin and has no activity against chondroitin sulfate and keratan sulfate (4). The enzyme readily releases highly sulfated oligosaccharides from heparin (5).

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. Linhardt, R. J. et al. (1986) Appl. Biochem. Biotech. 12:135.
  4. Wu, Z.L. et al. (2011) Glycobiology 21:625.
  5. Ernst, S. et al. (1996) Biochem. J. 315:589.

Entrez Gene IDs

1073402 (B. thetaiotaomicron)

Gene Symbol

BT_RS23555

UniProt

Q05819

Additional Heparinase I Products

Product Documents for Recombinant F. heparinum Heparinase I Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant F. heparinum Heparinase I Protein, CF

For research use only

Citations for Recombinant F. heparinum Heparinase I Protein, CF

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Protocols

View specific protocols for Recombinant F. heparinum Heparinase I Protein, CF (7897-GH):

Materials
  • Assay Buffer: 50 mM Tris, 100 mM NaCl, 2 mM CaCl2, pH 7.5
  • Recombinant F. heparinum Heparinase I (rFhHeparinase I) (Catalog # 7897-GH)
  • Substrate: Heparin (Tocris, Catalog # 2812), 20 mg/mL stock in deionized water
  • 96-well Clear UV Plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rFhHeparinase I to 0.5 µg/mL in Assay Buffer.
  2. Dilute Substrate to 0.75 mg/mL in Assay Buffer.
  3. Load 100 µL of diluted rFhHeparinase I into a UV plate, and start the reaction by adding 200 µL of 0.75 mg/mL Substrate. Include a Substrate Blank containing 100 µL of Assay Buffer and 200 µL of 0.75 mg/mL Substrate.
  4. Read plate 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 0.92 cm
     Note: the output of many spectrophotometers is in mOD Per Reaction:

  • rFhHeparinase I: 0.05 µg
  • Substrate: 0.5 mg/mL

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