Heparinase purified from Flavobacterium heparinum by ammonium sulfate precipitation, ion - exchange -, adsorption - and affinity - chromatography1), catalyzes the eliminative cleavage of heparin at the alpha - glucosaminide linkage to 2 - O - sulfo - L - iduronic acid, in which 2 - deoxy - 2 - sulfoamino - (6 - O - sulfo) - D - glucose participates2), thereby yielding oligosaccharides with delta 4 -hexuronate at the nonreducing ends. This enzyme does not act on typical heparan sulfate*.
*A heparan sulfate including the heparin - like structure (see above) may be cleaved by this enzyme.
- Reaction:
-
- Source:
- Flavobacterium heparinum
- Systematic Name:
- Heparin lyase
- Alternative Name:
- Heparinase, Heparin eliminase
- EC Number:
- EC 4.2.2.7
- Specific Activity:
- More than 50 units/mg protein
- Contaminants:
-
| Substrates |
|
Relative Activity |
| Chondroitin sulfate A (Whale cartilage) |
Lyase |
More than 0.1% |
| Chondroitin sulfate B (Hog skin) |
Lyase |
More than 1.0% |
| Chondroitin sulfate C (Shark cartilage) |
Lyase |
More than 0.1% |
| Hyaluronic acid (Human umbilical cord) |
Lyase |
More than 0.1% |
| Heparan sulfate (Bovine kidney) |
Lyase |
More than 1.0% |
| Delta Di HS-3S |
Hydrolase |
More than 0.1% |
| Delta Di HS-0S |
Hydrolase |
More than 0.1% |
- Appearance:
- Lyophilized powder containing 10mM potassium phosphate buffer, pH 6.8
BSA free, Preservative free
- Reconstitution:
- Dissolve the enzyme in 200µL of 0.1% BSA.
- Specifications:
-
| Optimum pH 3) |
pH 7.0 |
| Recommended Reaction Temperature |
37oC |
| Molecular Weight 3) |
42,700¡Þ1,200 (SDS - PAGE) |
| Isoelectric point 3) |
8.5 |
| Km value 3) |
8.04 x 10-6M (at 0.5 µg protein/mL)3) |
| Activators |
10-3M Ca2+ activates and/or stabilizes the enzyme.
Slight stimulation with Mn2+, Cd2+, Na+ (10-3M)4, 5) |
| Inhibitors |
Hg2+ (Inhibits by 100%)
Cu2+, Fe3+ (Inhibits by 50%)
Co2+ (10-5 M)4, 5) |
- Unit Definition:
- One unit of the enzyme is the amount required for the eliminative cleavage of typical heparin (e.g. Inolex heparin purified by ion exchange chromatography) yielding UV - absorbing materials corresponding to 1µ mole of delta 4 - hexuronate residues/minute at 37oC, pH7.0, as calculated with a value of 5,5006) for the molar absorption coefficient of the products, delta 4 - unsaturated oligosaccharides. Protein is determined according to Lowry7) with bovine serum albumin as a standard.
- Storage:
- Store at or below - 20oC until opened. Following reconstitutions aliquot and freeze (-20oC).
- References:
-
| 1) |
Ototani, N., Kikuch, M. and Yoshizawa, Z.: Carbohyd. Res., 88, 291 (1981) |
| 2) |
Linker, A. and Hovingh, P.: Carbohyd. Res., 127, 75 (1984) |
| 3) |
Yang, V. C., Linhardt, R. J., Bernstein, H., Cooney, C. L. and Langer, R.: J. Biol. Chem., 260, 1849 (1985) |
| 4) |
Hovingh, P. and Linker, A.: J. Biol. Chem., 245, 6170 (1970) |
| 5) |
Dietlich, C. P.: Methods of Biochemical Analysis, 24, 209 (1977) |
| 6) |
Hovingh, P. and Linker, A.: Carbohyd. Res., 37, 181 (1974) |
| 7) |
Lowry, D. H., Rosenbrough, N. J., Farr, A. L. and Randall, R. J.: J. Biol. Chem., 193, 265 (1951) |
| 8) |
Linker, A. and Hovingh, P.: Methods in Enzymol., 28, 902 (1972)
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