Purification and characterization of l-amino acid oxidase from king cobra ( Ophiophagus hannah) venom and its effects on human platelet aggregation

Abstract

Z.-Y. Li, T.-F. Yu and E. C.-Y. Lian. Purification and characterization of l-amino acid oxidase from king cobra ( Ophiophagus hannah) venom and its effects on human platelet aggregation. Toxicon 32, 1349–1358, 1994.—Venoms of several snake species contain large amounts of l-amino acid oxidase but its effects on human plasma coagulation and platelet aggregation have not been explored. We have purified l-amino acid oxidase from king cobra venom through CM-Sephadex C-25, Sephadex G-100 and DEAE Sephadex A-50 chromatographies. The purified enzyme has a mol. wt of 135,000 as determined by gel filtration and 65,000 by SDS-PAGE under non-reducing and reducing conditions. Incubation of plasma with l-amino acid oxidase at 200 μg/ml did not affect prothrombin time, activated partial thromboplastin time, or thrombin time. Upon addition of l-amino acid oxidase, platelets in platelet-rich plasma were aggregated. The enzyme-induced aggregation was abolished by catalase. The aggregation was also inhibited by indomethacin, aspirin, ethylenediaminetetraacetate, sodium nitroprusside, prostaglandin E 1, mepacrine and verapamil, but not by heparin, hirudin, creatine phosphate/creatine phosphokinase or antimycin/2-deoxy- d-glucose. These results suggest that l-amino acid oxidase induces human platelet aggregation through the formation of H 2O 2, and subsequent thromboxane A 2 synthesis requiring Ca 2+ but independent of ADP release. The platelet aggregation caused by l-amino acid oxidase is likely to contribute to toxicity inflicted by cobra venom.