Heparinase III
Heparinase III, also known as Heparin Lyase III or heparan sulfate lyase/eliminase or Heparitinase I is an endolytic enzyme. It cleaves selectively and exclusively the heparan sulfate via an elimination mechanism, undersulfated polysaccharide chains or acetylated or unsubstituted 1-4 linkages between hexosamines and glucuronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids which can be detected by UV spectroscopy at 232 nm. The enzyme is active only on heparan sulfate and does not cleave heparin.
Recombinant enzyme homologous to its native counterpart and expressed in the natural host, Pedobacter heparinus (formerly, Flavobacterium heparinum).
- EC Number: 4.2.2.8
- Purity: ≥ 95 % by reversed phase HPLC analysis
- IBEX liquid heparinase III has optimal activity and stability for up to 30 months when stored frozen at -70°C
- IBEX lyophilized heparinase I has optimal activity and stability for up to two years when stored at 2-8oC
- Calcium dependent enzyme and has optimal activity in the presence of 1.5-5.0 mM CaCl2 . Highest activity observed when Ca+2 was at 10 mM
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- Data Sheet - LIQUID
- Safety Data Sheet - LIQUID
- Data Sheet - LYOPHILIZED
- Safety Data Sheet - Lyophilized
Data Sheet - LIQUID
| DATA SHEET | Revised August 2018, R.05 | |
| Heparinase III | Research Grade | PN 50-012 50-012-001 |
| Synonyms | Heparin sulfate eliminase; Heparitin-sulfate lyase | |
| Source | Flavobacterium heparinum (recombinant) | |
| EC Number | 4.2.2.8 | |
| CAS Number | 37290-86-1 | |
| Catalyzed Reaction | The enzyme cleaves selectively, via an elimination mechanism, sulfated polysaccharide chains containing 1-4 linkages between hexosamines and glucuronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids which can be detected by UV spectroscopy at 232 nm. The enzyme is active only towards heparan sulfate and does not cleave heparin or low molecular weight heparins. | |
| Substrate Specificity | Heparan sulfate. Heparinase III cleaves heparan sulfate exclusively, and does not cleave unfractionated heparin or low molecular weight heparins. | |
| Properties | • Molecular weight: 73,202 Da • Isoelectric point: 9.6 – 9.9 • pH optimum for activity: 7 – 8 • pH range for activity: 5.5 – 9 • Optimal testing temperature range: 20 ºC – 37 ºC • Optimal storage temperature: – 70 ºC | |
| Purity | ≥95 % by reversed phase HPLC analysis. | |
 | ||
| Specific Activity | >45 IU/mg. One international unit (IU) is defined as the amount of enzyme that will liberate 1.0 µmole unsaturated oligosaccharides from heparan sulfate per minute at 30 ºC and pH 7.5. | |
| Stability | • PN 50-012 (vial of 0.5 IU): Expiration is 30 months from manufacturing date frozen at -70 ºC in aqueous buffer containing Sodium Chloride, Sodium Phosphate and Sucrose 5%. • PN 50-012-001 (vial of 0.1 IU): Expiration is 30 months from manufacturing date frozen at -70 ºC in aqueous buffer containing Sodium Chloride, Sodium Phosphate and Sucrose 5% | |
| Applications | • As a research reagent (glycosaminoglycan degradation). • For the preparation of disaccharides of heparan sulfate and the preparation of oligosaccharide libraries. | |
| Availability | A proprietary expression system for F. heparinum and the fermentation and isolation processes developed by IBEX Pharmaceuticals allow the production of large quantities of high purity product. | |
| References | • Review: “Enzymatic Degradation of Glycosaminoglycans”. S. Ernst et al. in Critical Reviews in Biochemistry and Molecular Biology (1995), 30(5): 387-444. • “Purification and Characterization of Heparin Lyases from Flavobacterium heparinum”. D.L. Lohse and R.J. Linhardt in J. Biol. Chem. (1992) 267: 24347-24355. • “Purification and Characterization of Heparinase from Flavobacterium heparinum”. V.C.Yang, R.J. Linhardt, H. Bernstein, C.L. Cooney and R. Langer in J. Biol. Chem. (1985) 260(3): 1849-1857. • “Substrate Specificity of the Heparin Lyases from Flavobacterium heparinum”. U.R.Desai, H.Wang and R.J. Linhardt in Archives of Biochemistry and Biophysics (1993) 306(2): 461-468. • “Isolation and Expression in Escherichia coli of hepB and hepC, Genes Coding for the Glycosaminoglycan-Degrading Enzymes Heparinase II and Heparinase III, Respectively, from Flavobacterium heparinum”. HongSheng Su, Françoise Blain, Roy A. Musil, Joseph J.F. Zimmermann, KangFu Gu and D. Clark Bennett, in Applied and Environmental Microbiology (1996): 2723-2734. • “Heparinase III from Flavobacterium heparinum: Cloning and Recombinant Expression in Escherichia coli”. R. Godavarti, M. Davis, G. Venkataraman, C. Cooney, R. Langer and R. Sasisekharan, in Biochemical and Biophysical Research Communications (1996) 225: 751-758. • “Involvement of heparan sulfate and related molecules in sequestration and growth promoting activity of fibroblast growth factor”. I. Vlodavsky, H-Q. Miao, P. Danagher and D. Ron, in Cancer and Metastasis Reviews (1996) 15(2): 177-186. |
Safety Data Sheet - LIQUID
| Safety Data Sheet - Heparinase III | |
| SECTION 1 – IDENTIFICATION | |
| Release Date | May 2019 R. 01 |
| Product Identity | Heparinase III, recombinant in sucrose, Research Grade |
| Part Number | 50-012 |
| Manufacturer | IBEX Pharmaceuticals Inc. 5485 Rue Paré, Suite 100 Montréal, Québec, H4P 1P7 Canada Tel : (514) 344-4004 Fax : (514) 344-8827 |
| Product use | Research |
| SECTION 2 – HAZARD IDENTIFICATION | |
| Emergency overview | Solution in water. Will not support burning. Not explosive. |
| Potential Health Effects | |
| Skin | May cause skin irritation |
| Eyes | May cause irritation or burning sensation |
| Inhalation | May cause irritation to the respiratory tract |
| Swallowed | Not known to cause toxic effects |
| Injection | Not known to cause toxicity with acute exposure, but may produce toxicity with repeated injections. |
| Carcinogenicity | Not determined |
| SECTION 3 – COMPOSITION/INFORMATION ON INGREDIENTS | |
| CAS No. | 37290-86-1 |
| EC No. | 4.2.2.8 |
| Systematic name | Heparin-sulfate lyase |
| Synonyms | Heparin sulfate eliminase Heparinase III |
| Origin | Recombinant, non-pathogenic microorganism |
| Ingredients | No components need to be disclosed according to the applicable regulations. |
| SECTION 4 – FIRST AID MEASURES | |
| Skin contact | Flush with copious amounts of water. |
| Eye contact | Rinse opened eye for several minutes under running water. Then consult a doctor. |
| Inhalation | Supply the victim with fresh air; consult doctor in case of complaints. |
| Swallowing | Rinse mouth with plenty of water. |
| On all of the above | Consult a doctor if symptoms develop. |
| SECTION 5 – FIRE FIGHTING MEASURES | |
| Suitable extinguishing agents | Noncombustible. |
| Protective equipment | No special measures required. Prevent contact with skin and eyes |
| SECTION 6 – ACCIDENTAL RELEASE MEASURES | |
| Person-related safety precautions | Prevent contact with skin and eyes. Ventilate area. |
| Measures for environmental protection | No special measures required. |
| Measures for cleaning/collecting | Spilled material should be carefully wiped up and disposed as biomedical waste as per applicable waste disposal regulations. Wash spill site after material pickup is complete. |
| SECTION 7 – HANDLING AND STORAGE | |
| Handling | |
| Information for safe handling | Prevent contact with skin and eyes and prevent inhalation and swallowing. No other special precautions are necessary if used correctly. |
| Information about protection against explosions and fires | No special measures required. |
| Storage | |
| Requirements to be met by storerooms and receptacles | No special requirements except that product should be stored frozen (preferably at -70 °C). |
| Information about storage in one common storage facilit | Not required. |
| Further information about storage conditions | None |
| Storage class | None assigned |
| Class according to regulation on flammable liquids | Void |
| SECTION 8 – EXPOSURE CONTROLS / PERSONAL PROTECTION | |
| General protective and hygienic measures | The usual precautionary measures for handling chemicals and potentially biohazardous materials should be followed. Use clothing sufficient to avoid skin contact. Safety shower and eye wash should be available in proximity. |
| Protection of hands | Use synthetic water resistant gloves. |
| Eye protection | Use safety glasses. |
| SECTION 9 – PHYSICAL AND CHEMICAL PROPERTIES | |
| Physical form | Frozen, colorless, odorless aqueous solution. (Heparinase III is a protein) |
| Melting point | 0 °C (melting point of aqueous solution) |
| Boiling point | Not applicable |
| Flash point | Not applicable |
| Flammability | Product is not flammable |
| Danger of explosion | Not determined |
| Solubility | Heparinase III and ingredients are water soluble |
| Organic solvents | 0 % |
| Solids content | 0 % |
| SECTION 10 – STABILITY AND REACTIVITY | |
| Thermal decomposition / conditions to be avoided | No decomposition if used according to specifications. |
| Dangerous reactions | No dangerous reactions known. |
| Dangerous products of decomposition | No dangerous decomposition products known. |
| SECTION 11 – TOXICOLOGICAL INFORMATION | |
| Effects of Acute Exposure | Low acute toxicity expected following inhalation, ingestion, injection or skin exposure. The potential eye irritation effects of Heparinase III are unknown and it should be considered to be potentially irritating to the eyes. |
| Effects of Chronic Exposure | Not determined for oral ingestion or skin exposure |
| Development Toxicity | Not determined |
| Reproductive Toxicity | Not determined |
| Carcinogenicity | Not determined |
| SECTION 12 – ECOLOGICAL INFORMATION | |
| Toxicity | No data available |
| Persistence and degradability | No data available |
| Bioaccumulative potential | No data available |
| Mobility in soil | No data available |
| PBT and vPvB assessment | No data available |
| Other adverse effects | No data available |
| SECTION 13 – DISPOSAL INFORMATION | |
| Waste Disposal | Incinerate or bury as a solid in a licensed facility. Do not discharge into waterways or sewer systems without proper authority. |
| Container Disposal | Dispose of in a licensed facility. Recommend crushing or other means to prevent unauthorized reuse. |
| SECTION 14 – TRANSPORT INFORMATION | |
| DOT (US) | Not dangerous goods |
| IMDG | Not dangerous goods |
| IATA | Not dangerous goods |
| SECTION 15 – REGULATORY INFORMATION | |
| Not a dangerous substance or mixture according to the Globally Harmonised System (GHS). | |
| SECTION 16 – OTHER INFORMATION | |
| This information is given without any warranty or representation. It is believed to be correct but does not claim to be all inclusive and shall be used only as a guide. IBEX Pharmaceuticals Inc. shall not be held liable for any damage resulting from handling or contact with the above product. It is offered solely for your consideration, investigation and verification. | |
Data Sheet - LYOPHILIZED
| DATA SHEET | Revised August 8, 2018 | |
| Heparinase III Lyophilized | Research Grade | PN 60-020 60-012-021 |
| Synonyms | Heparin sulfate eliminase; Heparitin-sulfate lyase | |
| Source | Flavobacterium heparinum (recombinant) | |
| EC Number | 4.2.2.8 | |
| CAS Number | 37290-86-1 | |
| Catalyzed Reaction | The enzyme cleaves selectively, via an elimination mechanism, sulfated polysaccharide chains containing 1-4 linkages between hexosamines and glucuronic acid residues. The reaction yields oligosaccharide products (mainly disaccharides) containing unsaturated uronic acids which can be detected by UV spectroscopy at 232 nm. The enzyme is active only towards heparan sulfate and does not cleave heparin or low molecular weight heparins. | |
| Substrate Specificity | Heparan sulfate. Heparinase III cleaves heparan sulfate exclusively, and does not cleave unfractionated heparin or low molecular weight heparins. | |
| Properties | • Lyophilized powder • Molecular weight: 73,202 Da • Isoelectric point: 9.6 – 9.9 • pH optimum for activity: 7 – 8 • pH range for activity: 5.5 – 9 • Optimal testing temperature range: 20 ºC – 37 ºC • Optimal storage temperature: 5 +/-3ºC | |
| Purity | ≥ 95 % by reversed phase HPLC analysis, made from Heparinase III (PN 50-003). | |
| Enzymatic Activity | One international unit (IU) is defined as the amount of enzyme that will liberate 1.0 µmole unsaturated oligosaccharides from heparan sulfate per minute at 30 ºC and pH 7.5. | |
| Reconstitution | Add 250 µL of water to reconstitute to its original formulation. | |
| Stability | Expiration is 24 months from manufacturing date when stored at 5 +/- 3ºC. | |
| Applications | • As a research reagent (glycosaminoglycan degradation). • For the preparation of disaccharides of heparan sulfate and the preparation of oligosaccharide libraries. | |
| Availability | A proprietary expression system for F. heparinum and the fermentation and isolation processes developed by IBEX Pharmaceuticals allow the production of large quantities of high purity product. | |
| References | • Review: “Enzymatic Degradation of Glycosaminoglycans”. S. Ernst et al. in Critical Reviews in Biochemistry and Molecular Biology (1995), 30(5): 387-444. • “Purification and Characterization of Heparin Lyases from Flavobacterium heparinum”. D.L. Lohse and R.J. Linhardt in J. Biol. Chem. (1992) 267: 24347-24355. • “Purification and Characterization of Heparinase from Flavobacterium heparinum”. V.C.Yang, R.J. Linhardt, H. Bernstein, C.L. Cooney and R. Langer in J. Biol. Chem. (1985) 260(3): 1849-1857. • “Substrate Specificity of the Heparin Lyases from Flavobacterium heparinum”. U.R.Desai, H.Wang and R.J. Linhardt in Archives of Biochemistry and Biophysics (1993) 306(2): 461-468. • “Isolation and Expression in Escherichia coli of hepB and hepC, Genes Coding for the Glycosaminoglycan-Degrading Enzymes Heparinase II and Heparinase III, Respectively, from Flavobacterium heparinum”. HongSheng Su, Françoise Blain, Roy A. Musil, Joseph J.F. Zimmermann, KangFu Gu and D. Clark Bennett, in Applied and Environmental Microbiology (1996): 2723-2734. • “Heparinase III from Flavobacterium heparinum: Cloning and Recombinant Expression in Escherichia coli”. R. Godavarti, M. Davis, G. Venkataraman, C. Cooney, R. Langer and R. Sasisekharan, in Biochemical and Biophysical Research Communications (1996) 225: 751-758. • “Involvement of heparan sulfate and related molecules in sequestration and growth promoting activity of fibroblast growth factor”. I. Vlodavsky, H-Q. Miao, P. Danagher and D. Ron, in Cancer and Metastasis Reviews (1996) 15(2): 177-186. • “Control of Cell Proliferation by Heparan Sulfate and Heparin-Binding Growth Factors”. I. Vlodavsky, H-Q. Maio, R. Atzmon, E. Levi, J. Zimmermann, R. Bar-Shavit, T. Peretz and S. Ben-Sasson, in Thrombosis and Haemostasis (1995) 74(1): 534-540. • “Heparinase III Exerts Endothelial and Cardioprotective Effects in Feline Myocardial Ischemia-Reperfusion Injury”. R. Hayward, T.O. Nossuli and A.M. Lefer, in J. Pharmacology and Experimental Therapeutics (1997) 283: 1032-1038. • “IBT 9302 (Heparinase III): a novel enzyme for the management of reperfusion injury-related vascular damage, restenosis and wound healing”. P. Silver, in Exp. Opin. Invest. Drugs (1998) 7(6): 1003-1014 • “Cellular mechanisms of heparinase III protection in rat traumatic shock”. R. Hayward, R. Scalia, B. Hopper, J. Appel III and A. Lefer, in Am. J. Physiol. (1998) 275 (Heart Circ. Physiol. 44): H23-H30. • “Heparinase III limits rat arterial smooth muscle cell proliferation in vitro and in vivo”. P. Silver, J-P. Moreau, E. Denholm, Y.Q. Lin, L. Nguyen, C. Bennett, A. Recktenwald, D. DeBlois, S. Baker, S. Ranger in Euro. J. Pharmacol. (1998) 351:79-83. |
Safety Data Sheet - Lyophilized
| Safety Data Sheet - Heparinase III - Lyophilized | |
| SECTION 1 – IDENTIFICATION | |
| Release Date | August 7, 2018 |
| Product Identity | Heparinase III, Lyophilized |
| Part Number | 60-020, 60-021 |
| Manufacturer | IBEX Pharmaceuticals Inc. 5485 Rue Paré, Suite 100 Montréal, Québec, H4P 1P7 Canada Tel : (514) 344-4004 Fax : (514) 344-8827 |
| Product use | Research |
| SECTION 2 – HAZARD IDENTIFICATION | |
| Emergency overview | Powder. Will not support burning. Not explosive. |
| Potential Health Effects | |
| Skin | May cause skin irritation |
| Eyes | May cause irritation or burning sensation |
| Inhalation | May cause irritation to the respiratory tract |
| Swallowed | Not known to cause toxic effects |
| Injection | Not known to cause toxicity with acute exposure, but may produce toxicity with repeated injections. |
| Carcinogenicity | Not determined |
| SECTION 3 – COMPOSITION/INFORMATION ON INGREDIENTS | |
| CAS No. | 37290-86-1 |
| EC No. | 4.2.2.8 |
| Systematic name | Heparin-sulfate lyase |
| Synonyms | Heparin sulfate eliminase Heparinase III |
| Origin | Recombinant, non-pathogenic microorganism |
| Ingredients | No components need to be disclosed according to the applicable regulations. |
| SECTION 4 – FIRST AID MEASURES | |
| Skin contact | Flush with copious amounts of water. |
| Eye contact | Rinse opened eye for several minutes under running water. Then consult a doctor. |
| Inhalation | Supply the victim with fresh air; consult doctor in case of complaints. |
| Swallowing | Rinse mouth with plenty of water. |
| On all of the above | Consult a doctor if symptoms develop. |
| SECTION 5 – FIRE FIGHTING MEASURES | |
| Suitable extinguishing agents | Noncombustible. |
| Protective equipment | No special measures required. Prevent contact with skin and eyes |
| SECTION 6 – ACCIDENTAL RELEASE MEASURES | |
| Person-related safety precautions | Prevent contact with skin and eyes. Ventilate area. |
| Measures for environmental protection | No special measures required. |
| Measures for cleaning/collecting | Spilled material should be carefully wiped up and disposed as biomedical waste as per applicable waste disposal regulations. Wash spill site after material pickup is complete. |
| SECTION 7 – HANDLING AND STORAGE | |
| Handling | |
| Information for safe handling | Prevent contact with skin and eyes and prevent inhalation and swallowing. No other special precautions are necessary if used correctly. |
| Information about protection against explosions and fires | No special measures required. |
| Storage | |
| Requirements to be met by storerooms and receptacles | No special requirements except that product should be stored refrigerated 2 - 8 °C |
| Information about storage in one common storage facilit | Not required. |
| Further information about storage conditions | None |
| Storage class | None assigned |
| Class according to regulation on flammable liquids | Void |
| SECTION 8 – EXPOSURE CONTROLS / PERSONAL PROTECTION | |
| General protective and hygienic measures | The usual precautionary measures for handling chemicals and potentially biohazardous materials should be followed. Use clothing sufficient to avoid skin contact. Safety shower and eye wash should be available in proximity. |
| Protection of hands | Use synthetic water resistant gloves. |
| Eye protection | Use safety glasses. |
| SECTION 9 – PHYSICAL AND CHEMICAL PROPERTIES | |
| Physical form | Lyophilized powder in amber glass vial closed with cap covered with aluminum seal. |
| Melting point | 0 °C (melting point of aqueous solution) |
| Boiling point | Not applicable |
| Flash point | Not applicable |
| Flammability | Product is not flammable |
| Danger of explosion | Not determined |
| Solubility | Heparinase III and ingredients are water soluble |
| Organic solvents | 0 % |
| Solids content | 100 % |
| SECTION 10 – STABILITY AND REACTIVITY | |
| Thermal decomposition / conditions to be avoided | No decomposition if used according to specifications. |
| Dangerous reactions | No dangerous reactions known. |
| Dangerous products of decomposition | No dangerous decomposition products known. |
| SECTION 11 – TOXICOLOGICAL INFORMATION | |
| Effects of Acute Exposure | Low acute toxicity expected following inhalation, ingestion, injection or skin exposure. The potential eye irritation effects of Heparinase III are unknown and it should be considered to be potentially irritating to the eyes. |
| Effects of Chronic Exposure | Not determined for oral ingestion or skin exposure |
| Development Toxicity | Not determined |
| Reproductive Toxicity | Not determined |
| Carcinogenicity | Not determined |
| SECTION 12 – ECOLOGICAL INFORMATION | |
| Toxicity | No data available |
| Persistence and degradability | No data available |
| Bioaccumulative potential | No data available |
| Mobility in soil | No data available |
| PBT and vPvB assessment | No data available |
| Other adverse effects | No data available |
| SECTION 13 – DISPOSAL INFORMATION | |
| Waste Disposal | Incinerate or bury as a solid in a licensed facility. Do not discharge into waterways or sewer systems without proper authority. |
| Container Disposal | Dispose of in a licensed facility. Recommend crushing or other means to prevent unauthorized reuse. |
| SECTION 14 – TRANSPORT INFORMATION | |
| DOT (US) | Not dangerous goods |
| IMDG | Not dangerous goods |
| IATA | Not dangerous goods |
| SECTION 15 – REGULATORY INFORMATION | |
| Not a dangerous substance or mixture according to the Globally Harmonised System (GHS). | |
| SECTION 16 – OTHER INFORMATION | |
| This information is given without any warranty or representation. It is believed to be correct but does not claim to be all inclusive and shall be used only as a guide. IBEX Pharmaceuticals Inc. shall not be held liable for any damage resulting from handling or contact with the above product. It is offered solely for your consideration, investigation and verification. | |
| Date of revision | August 7, 2018 |
Research Papers using IBEX Heparinase III
IBEX heparinase III (50-012-001) used to elucidate the mechanotransduction pathways for the shear-induced nitric oxide production and to improve the understanding of the cardiovascular disease progression. This will enable the design of future therapeutics to rescue endothelial dysfunction.
Heparan sulfate proteoglycan glypican‑1 and PECAM‑1 cooperate in shear‑induced endothelial nitric oxide production.
Abstract
This study aimed to clarify the role of glypican-1 and PECAM-1 in shear-induced nitric oxide production in endothelial cells. Atomic force microscopy pulling was used to apply force to glypican-1 and PECAM-1 on the surface of human umbilical vein endothelial cells and nitric oxide was measured using a fuorescent reporter dye. Glypican-1 pulling for 30 min stimulated nitric oxide production while PECAM-1 pulling did not. However, PECAM-1 downstream activation was necessary for the glypican-1 force-induced response. Glypican-1 knockout mice exhibited impaired fow-induced phosphorylation of eNOS without changes to PECAM-1 expression. A cooperation mechanism for the mechanotransduction of fuid shear stress to nitric oxide production was elucidated in which glypican-1 senses fow and phosphorylates PECAM-1 leading to endothelial nitric oxide synthase phosphorylation and nitric oxide production
Click the scientific reports logo above to read the full paper.
Click the scientific reports logo above to read the full paper.