Purification, characterization and crystallization of Jararacussin-I, a fibrinogen-clotting enzyme isolated from the venom of Bothrops jararacussu
| Autor: | Andreimar M. Soares, Leandra Watanabe, R.K. Bortoleto, Raghuvir K. Arni, Mário T. Murakami |
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| Rok vydání: | 2002 |
| Předmět: |
Ion chromatography
Crystallography X-Ray Toxicology Esterase Chromatography Affinity Benzamidine Sepharose chemistry.chemical_compound Fibrinolytic Agents Affinity chromatography Crotalid Venoms Animals Bothrops Protease Inhibitors Chromatography Molecular mass Serine Endopeptidases Esterases Fibrinogen Substrate (chemistry) Chromatography Ion Exchange Benzamidines Molecular Weight Dithiothreitol Isoelectric point chemistry Biochemistry Cattle Electrophoresis Polyacrylamide Gel Crystallization |
| Zdroj: | Toxicon. 40:1307-1312 |
| ISSN: | 0041-0101 |
| Popis: | A fibrinogen-clotting enzyme, Jararacussin-I, was purified from the venom of Bothrops jararacussu by a combination of ion exchange chromatography using Resource 15S resin and affinity chromatography using Benzamidine Sepharose 6B resin. Jararacussin-I displays a molecular mass of 28 kDa as estimated by sodium dodecyl sulphate–PAGE and possesses an isoelectric point of 5.0. The coagulant specific activity of the enzyme was determined to be 45.8 NIH U/mg using bovine fibrinogen as the substrate and the esterase specific activity was determined to be 258.7 U/mg. The protease inhibitors, benzamidine and DTT inhibited the esterase specific activity by 72.4 and 69.7%, respectively. The optimal temperature and pH for the degradation of both chains of fibrinogen and esterase specific activity were determined to be 37 °C and 7.4–8.0, respectively. The enzyme was inactivated at both 4 and 75 °C. Single crystals of Jararacussin-I were obtained and complete three-dimensional X-ray diffraction data was collected at the Brazilian National Synchrotron Source (LNLS) to a resolution of 2.4 A. |
| Databáze: | OpenAIRE |
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