الفهرس 
Effect of snake venom on the haemostatic
Anticoagulant proteins with non enzymatic activityOnly 14 pages are availabe for public view
 |  | from | 157 |  |  |
| | | from | 157 | | |
Abstract
The present study includes purification and characterization of a fibrinogenolytic metalloprotease from crude Naja nigricollis venom.
In this work, the fibrinogenolytic metalloprotease was purified by three steps of fractionation.
The first step: gel filtration chromatography on sephadex G-100, using ammonium acetate buffer (pH 4.8, 0.02 M). Three fractions were obtained: I, II and III. Fraction III showed the shortest plasma recalcification time and fibrinogen clotting time, indicating that it has the strongest coagulant activity. But Saad thesis (2005) reported that there is a coagulant component in fraction II by second step of fractionation using ion exchange Chromatography. Therefore in this work, fraction II was fractionated by ion exchange chromatography.
The second step: ion exchange chromatography on DEAE sephadex A-50, eluted two peaks IIa, IIb were obtained. The first peak showed shortening of fibrinogen clotting time indicating its coagulant activity. It was designated as IIa.
The third step: gel filtration chromatography on sephadex G-50,
using ammonium acetate buffer (pH4.8, 0.02M). Two fractions were obtained: IIIa, IIIb, both show shortening of fibrinogen clotting time, but fraction IIIa have large protein content.
Disc gel electrophoresis of fraction IIIa showed sharp band of approximate M.w. which is 24 kDa, optimum pH 8.0 with thermal stability up to 40 ˚C, with minimal caseinolytic activity, and not affected by heparin, but urea caused lyses of the formed clot by fraction IIIa.
Fraction IIIa showed a hemorrhagic fibrinogenolytic activity. Its activity was inhibited by EDTA and totally restored by adding back zinc ions, indicating that it is a zinc metalloprotease. Fraction IIIa also showed very minimal aggregatory effect on platelet aggregation.
Also fraction IIIa hemorrhagic activity was neutralized by the polyvalent antivenin (Egyptian polyvalent antivenin from VACSERA).