Recombinant P. heparinus Chondroitinase B Protein, CF Summary
Gln26-His506, with an N-terminal Met and 6-His tag
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
|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.
- 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
- Dilute rP. heparinus Chondroitinase B to 0.4 ng/μL in Assay Buffer.
- Dilute Substrate to 4 mg/mL in Assay Buffer.
- 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.
- Read at an absorbance of 232 nm in kinetic mode for 5 minutes.
- 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
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).
- Oike, Y. et al. (1980) Biochem. J. 191:193.
- Prabhakar, V. et al. (2009) J. Biol. Chem. 284:974.
- Trowbridge, J.M. and Gallo, R.L. (2002) Glycobiology 12:117R.
- Gu, K. et al. (1995) Biochem. J. 312:356.
- Tkalec, A.L. et al. (2000) Appl. Environ. Microbiol. 66:29.
- Wu, Z.L. et al. (2011) Glycobiology 21:625.
Product Specific NoticesCoomassie is a registered trademark of Imperial Chemical Industries Ltd.
Citations for Recombinant P. heparinus Chondroitinase B Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
Citations: Showing 1 - 4
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Glycosaminoglycan Domain Mapping of Cellular Chondroitin/Dermatan Sulfates
Authors: A Persson, E Vorontsov, G Larson, J Nilsson
Sci Rep, 2020;10(1):3506.
Sample Types: Cells
Applications: enzyme activity
LC-MS/MS characterization of xyloside-primed glycosaminoglycans with cytotoxic properties reveals structural diversity and novel glycan modifications
Authors: A Persson, A Gomez Tole, E Vorontsov, W Nasir, D Willén, F Noborn, U Ellervik, K Mani, J Nilsson, G Larson
J. Biol. Chem., 2018;0(0):.
Xyloside-primed chondroitin sulfate/dermatan sulfate from breast carcinoma cells with a defined disaccharide composition has cytotoxic effects in vitro
J Biol Chem, 2016;0(0):.
Sample Types: Complex Sample Type
Musculocontractural Ehlers-Danlos syndrome and neurocristopathies: dermatan sulfate is required for Xenopus neural crest cells to migrate and adhere to fibronectin
Authors: N Gouignard, M Maccarana, I Strate, K von Stedin, A Malmström, EM Pera
Dis Model Mech, 2016;0(0):.
Sample Types: Protein
Applications: Enzyme Assay
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