Recombinant P. heparinus Chondroitinase B Protein, CF

R&D Systems | Catalog # 6974-GH

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Key Product Details

  • R&D Systems E. coli-derived Recombinant P. heparinus Chondroitinase B Protein (6974-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

Q46079

Applications

Enzyme Activity
View all Chondroitin B Lyase/Chondroitinase B Proteins and Enzymes »

Product Specifications

Source

E. coli-derived p. heparinus Chondroitin B Lyase/Chondroitinase B protein
Gln26-His506, with an N-terminal Met and 6-His tag

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Inconclusive results, Met predicted. Protein identity confirmed by MS analysis of tryptic fragments.

Predicted Molecular Mass

55 kDa

SDS-PAGE

52-56 kDa, reducing conditions

Activity

Measured by its ability to hydrolyze chondroitin sulfate B.
The specific activity is >90,000 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

6974-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: Chondroitin B Lyase/Chondroitinase B

Chondroitinase B from P. heparinus is a depolymerizing lyase that is especially active on the sulfated polysaccharide dermatan sulfate (DS) (1, 2). DS is a sulfated polysaccharide that is abundantly located in the skin, but is also present in the blood vessels, heart valves, tendons, and lungs. DS may play roles in coagulation, cardiovascular disease, carcinogenesis, infection, wound repair, and fibrosis (3). DS with a repeating disaccharide unit of IdoA alpha 1-3GlcNAc is most closely related to chondroitin sulfate (CS) that has a repeating disaccharide unit of GlcA beta 1-3GlcNAc. Due to this similarity, DS was originally referred to as chondroitin sulfate B, and the DS-depolymerizing enzyme was named Chondroitinase B. DS contains more sulfate than CS at levels comparable to the anticoagulant drug heparin, which is the reason that purified commercial heparin is more likely to be contaminated by DS. Chondroitinase B can be used to specifically degrade and distinguish DS in glycosaminoglycan preparations (4, 5, 6).

References

  1. Oike, Y. et al. (1980) Biochem. J. 191:193.
  2. Prabhakar, V. et al. (2009) J. Biol. Chem. 284:974.
  3. Trowbridge, J.M. and Gallo, R.L. (2002) Glycobiology 12:117R.
  4. Gu, K. et al. (1995) Biochem. J. 312:356.
  5. Tkalec, A.L. et al. (2000) Appl. Environ. Microbiol. 66:29.
  6. Wu, Z.L. et al. (2011) Glycobiology 21:625.

Alternate Names

Chondroitinase B

Entrez Gene IDs

8251878 (P. heparinus)

Gene Symbol

PHEP_0789

UniProt

Q46079

Additional Chondroitin B Lyase/Chondroitinase B Products

Product Documents for Recombinant P. heparinus Chondroitinase B Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Recombinant P. heparinus Chondroitinase B Protein, CF

Coomassie is a registered trademark of Imperial Chemical Industries Ltd.

For research use only

Citations for Recombinant P. heparinus Chondroitinase B Protein, CF

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Protocols

View specific protocols for Recombinant P. heparinus Chondroitinase B Protein, CF (6974-GH):

Materials
  • Assay Buffer: 50 mM Tris, 5 mM CaCl2, pH 7.5
  • Recombinant P. heparinus Chondroitin B Lyase/Chondroitinase B (rP. heparinus Chondroitinase B) (6974‑GH)
  • Substrate: Chrondroitin Sulfate B (Sigma, Catalog # C3788), 25 mg/mL stock in deionized water
  • 96 well UV Plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rP. heparinus Chondroitinase B to 0.4 ng/μL in Assay Buffer.
  2. Dilute Substrate to 4 mg/mL in Assay Buffer.
  3. Load 50 μL of 0.4 ng/μL rP. heparinus Chondroitinase B into the plate, and start the reaction by adding 50 μL of 4 mg/mL Substrate. Include a Substrate Blank containing 50 μL of Assay Buffer and 50 μL of 4 mg/mL Substrate.
  4. Read at an absorbance of 232 nm in kinetic mode for 5 minutes.
  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:
  • rP. heparinus Chondroitinase B: 0.020 μg
  • Chondroitin Sulfate B: 200 μg

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