江浙蝮蛇毒纤溶酶的分离纯化及性质研究

目的:江浙蝮蛇毒纤溶酶的分离纯化及部分生物化学和药理性质的研究.方法:应用DEAE Sepharose CL-6B阴离子交换柱和HiPrep Sephacryl S 100柱分离纯化纤溶酶,以高效液相色谱仪和聚丙烯酰胺凝胶电泳检测纯度、相对分子质量,应用等电聚焦电泳测定等电点,然后测定氨基酸组分及部分药理性质.结果:分离得到相对分子质量为59 100,等电点为4.98, SDS-PAGE为一条带,高效液相色谱分析为单一峰的纤溶酶.该组分属于金属蛋白酶类.氨基酸组成分析表明它含较多的酸性氨基酸.高浓度的纤溶酶对肿瘤细胞CEN2有抑制作用,有微弱的出血毒性.结论:分离纯化得到了一种有微弱出血毒,具有较强抗凝活性的单一组分的酸性金属蛋白酶.

PURIFICATION AND CHARACTERIZATION OF FIBRINOLYTIC ENZYME FROM AGKISTRODON HALYS PALLAS VENOM

Objective:To investigate the purification method of a fibrinolytic enzyme(FLE) from the venom of Agkistrodon blomhoffi brevicaudus and research some biochemistral characterizations.Methods:The fibrinolytic enzyme(FLE) was purified from the venom of Agkistrodon blomhoffi brevicaudus by ion-exhange chromatography on a DEAE Sepharose CL-6B column,followed by gel filtration chromatography on HiPrep Sephacryl S 100.The molecular weight was estimated with SDS-PAGE.The pI was determined through isoelectric focusing.Amino acid composition assay was carried out on a Hitachi 835-50 mould automatic amino acid analyzer.A series of pharmacological experiments were carried out.Result:The purified FLE was shown to be homogeneous in SDS-PAGE,which acted as an anticoagulant in vitro.Its molecular weight was estimated to be 59,100 D and isoelectric point was 4.98.According to the result of amino acid composition assay,it has a number of acidic amino acid,which is consistent with the pI we detected.Through the research of pharmacology,we found it possessed low haemorrhagic activity.FLE belongs to metallic protein and can inhabit the growth of cancer in high concentration.Conclusion:We separate a fibrinolytic enzyme(FLE) which is a acid protein and has low haemorrhagic activity from the venom of Agkistrodon blomhoffi brevicaudus.