短尾蝮蛇毒磷脂酶A_2的分离纯化及抗血小板聚集作用

Purification of Phospholipase A_2 from Agkistrodon halys Brevicaudus Venom and Its Inhibition on Platelet Aggregation

目的研究短尾蝮蛇毒磷脂酶A2的分离纯化及其抗血小板聚集作用。方法磷脂酶A2的分离纯化采用CM-Sephadex C-25、DEAE-Sepharose CL-6B、Sephacryl S-200、Sephadex G-75柱层析法,用SDS-聚丙烯酰胺凝胶电泳测定其蛋白分子质量,以磷脂酶A2测定方法测定其酶活性,用比浊法测定其对二磷酸腺苷(ADP)引起的血小板聚集的影响。结果从短尾蝮蛇毒中纯化所得磷脂酶A2的相对分子质量为16.0×103(非还原)、17.6×103(还原),它具有磷脂酶A2活性,能明显抑制ADP引起的血小板聚集并呈剂量-效应关系。结论此方法成功地从短尾蝮蛇毒中分离纯化出磷脂酶A2,并能抑制血小板聚集。

Objective .To purify of Phospholipase A2 from Agkistrodon halys Brevicaudus Venom and study its inhibition on platelet aggregation. Methods The isolation and purification of phospholipase A2 was done by chromatography on CM-Sephadex C-25,DEAE-Sepharose CL-6B,Sephacryl S-200 and Sephadex G-75 column. Its characterization researches were done by SDS-PAGE electrophoresis and phospholipase A2 activity analysis. The experiment of platelet aggregation by ADP was measured by turbidimetric method. Results The results showed that the protein purified from Agkistrodon halys Brevicaudus Venom was Phospholipase A2. Its molecular weights was 16.0 ×10^3 （non-reduction）, 17. 6×10^3（reduction）. The activities of phospholipase A2 showed strongly and it could dose-dependently inhibit platelet aggregation induced by ADP. Conclusion. It is a successful method to purify the Phospholipase A2 from Agkistrodon halys Brevicaudus Venom and the protein can inhibit platelet aggregation.