Purification And Characterization Of L-amino Acid Oxidase From Guangxi Cobra Venom And Its Antiplatelet Effect On Human And Rabbit

Objective :To purify L-amino acid oxidase from Guangxi cobra venom and study the physical and biochemical characterization,as well as its effect on inhibitting the platelet aggregation in vitro and in vivo .Methods :By using of Sephacryls-200 gel filtration,DEAE- Sepharose CL-6B ion-exchange chromatography and CM- Sepharose CL-6B ion-exchange chromatography,a protein with L-amino acid oxidase activity was purified from Guangxi cobra venom and its properties were characterized .SDS- polyacrylamid gel electrphoresis(SDS-PAGE) was used to analyze its molecular weight at both reducing and non-reducing conditions.The isoelectric point was estimated by the isoelectricfocusing electrophoresis .The platelet aggregation induced by ADP ,thrombin,collagen and AA was measured by turbidinmetric method .The level of GMP-140 was measured by immunoradiometry .The ultrastructures of platelet were observed by the electron microscopy as well. Result:The purified L-amino acid oxidase showed to be a single band on the SDS-PAGE with a molecular weight of 55.2KD.The isoelectric point of the purified L-amino acid oxidase was PH 7.6 .It was devoid of phospholipase A2, fibrinogenolytic, fibrinolytic , esterase , haemorrhagenic activities.The puried L-amino acid oxidase dose-dependently inhibited the aggregation of Human platelet induced by ADP , thrombin , collagen and AA in vitro and dose-dependently inhibited the aggregation of rabbit platelet induced by ADP in vivo. The puried L-amino acid oxidase could decrease the level of GMP-140 in plasm ,inhibit the platelet...