Purification and properties of the main coagulant and anticoagulant principles of Vipera russellii snake venom.
Vipera russellii venom was separated into thirteen fractions by means of DEAE-Sephadex A-50 column chromatography. Fraction III possessed anticoagulant and phospholipase A activities and Fraction XI possessed procoagulant and caseinolytic activities, both were further purified by gel filtration on Sephacryl S-200 column. Purified procoagulant (Component II) was a two-chain protein with molecular weight of 86 000 consisting of A-chain (Mr 66 000) and B-chain (Mr 20 000). It was a glycoprotein containing 7.8% neutral sugar and 715 amino-acid residues. The procoagulant activity was 10-times that of the crude venom. It was an acidic proteinase with isoelectric point of pH 4.2. Upon heat treatment at 60 degrees C, Component II was stable at pH 5.5 and 7.2 for 3 h, but was destroyed completely after 30 min at pH 8.9. It was devoid of esterase or amidase activity. Purified anticoagulant (Component I) was a single peptide chain with molecular weight of 16 000. It was carbohydrate free and contained 136 amino-acid residues. It was a basic protein with an isoelectric point of larger than pH 10. It was a potent phospholipase A with an enzymatic activity of 510 +/- 30 mumol/min per mg using phosphatidylcholine as substrate, and 1 microgram/ml was sufficient to cause 100% hemolysis by the indirect hemolytic method. Upon heat treatment at 90 degrees C, Component I was heat stable at pH 5.5 for more than 3 h, but was destroyed completely after 2 h at pH 7.2 and 8.9. The anticoagulant activity of Component I could be neutralized by platelet factor 3, tissue thromboplastin and cephalin.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000925:Anticoagulants / Q000302:isolation & purification*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003029:Coagulants / Q000302:isolation & purification*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D010314:Partial Thromboplastin Time; D010974:Platelet Aggregation / Q000187:drug effects; D011817:Rabbits; D013916:Thrombelastography; D014757:Viper Venoms / Q000032:analysis*
D000596:Amino Acids; D000925:Anticoagulants; D003029:Coagulants; D014757:Viper Venoms