BIOCHEMICAL AND IMMUNOLOGICAL STUDIES ON ALPHA-FIBRINOGENASE FROM NAJA NIGRICOLLIS VENOM

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dc.contributor.authorABDULLAHI, Hadiza Lawal
dc.date.accessioned2015-06-11T09:14:06Z
dc.date.available2015-06-11T09:14:06Z
dc.date.issued2014-12
dc.description.abstractA fibrinogenolytic enzyme, Naja nigricollis metalloprotease, was purified from Naja nigricollis venom using Diethylaminoethyl (DEAE) Cellulose, Sephadex G-75 and Heparin-Agarose column chromatography to apparent homogeneity. The purified Naja nigricollis metalloprotease migrated as a single protein band on analytical polyacrylamide gel electrophoresis with an apparent molecular mass of 65 KDa, under reducing conditions in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The Naja nigricollis metalloprotease, displayed fibrinogenase activity, which was about 9-folds higher than that of the crude venom. The enzyme was proteolytically active against human fibrinogen. The enzyme cleaved preferentially the α-chain, leaving the β-chain and γ-chain of fibrinogen intact. The fibrinogenolytic activity of the enzyme was inhibited by the chelating agent EDTA and 1, 10-phenanthroline and slightly inhibited by phenylmethylsulfonyl fluoride, aprotinin and beta-mercaptoethanol, indicating it is a metalloprotease. The inhibition by EDTA could be blocked by Zn2+, but not by Ca2+ or Mg2+ and Co2+. The optimum pH and temperature for the enzyme activity were found to be 7.5 and 45°C respectively. The enzyme was stable at pH 6.5-7.8 and at a temperature range of 20-60°C with activation energy (Ea) of 0.031 kJ/mol/K. The kinetic parameters, Km and Vmax were found to be 0.091 mg/ml and 0.00711 μmol/min respectively. The purified N. nigricollis fibrinogenase has haemorrhagic activity and the polyclonal antibodies raised against the enzyme were capable of neutralizing the haemorrhagic effects of the enzyme in laboratory animals. The kinetic properties and alpha-fibrinogenolytic effect of the enzyme indicates its suitability as a possible prototype for development of defibrinogenating agents and development of diagnostic reagents for detection of fibrinogen. The anti-haemorrhagic effect of the antibodies suggests its suitability also as possible vaccine candidate.en_US
dc.identifier.urihttp://hdl.handle.net/123456789/6644
dc.language.isoenen_US
dc.subjectALPHA-FIBRINOGENASE FROM NAJA NIGRICOLLIS VENOMen_US
dc.subjectALPHA-FIBRINOGENASE FROM NAJA NIGRICOLLIS VENOMen_US
dc.titleBIOCHEMICAL AND IMMUNOLOGICAL STUDIES ON ALPHA-FIBRINOGENASE FROM NAJA NIGRICOLLIS VENOMen_US
dc.typeThesisen_US
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