Isolation, partial purification and characterization of phospholipase A2 from Naja Katiensis venom

Abstract

The most effective and acceptable therapy for snakebite victims is the immediate administration of antivenin which is limited by problems of hypersensitivity reactions in some individuals and its inability to resolve the local effects of the venom. The aim of this study was to isolate, partially purify and characterize phospholipase A2 from Naja Katiensis venom. Phospholipase A2 was partially purified via a two-step process: gel filtration on Sephadex G-75 and ion exchange chromatography using CM Sephadex, and subjected to SDS-PAGE analysis. From the results, the specific activity of the partially purified PLA2 decreased from 0.67μmol/min/mg in crude venom to 0.29μmol/min/mg after ion exchange chromatography with a yield of 5% and purification fold of 0.43. The optimum temperature of the purified PLA2 was found to be 35ºC and optimum p.H of 7. velocity studies for the determination of kinetic constants using L-a-lecithin as substrate revealed a K_m  of 1.47mg/ml and V_max  of 3.32μ moles/min/mg. The sodium dodecyl sulphate polyacrylamide gel electrophoresis of the purified PLA₂ showed a distinct band with molecular weight estimated to be 14KDa. In conclusion, the present study shows that phospholipase A₂ was isolated, purified and characterized. This may serve as a promising candidate for future development of a novel anti-venin drug.