蚯蚓纤溶酶的纯化及pH值、温度和胰蛋白酶对其纤溶活性的影响

目的纯化并鉴定蚯蚓纤溶酶 ,研究pH、温度和胰蛋白酶对其纤溶活性的影响。方法以赤子爱胜蚓为材料 ,采用硫酸铵盐析、DEAESepharoseCL 6B离子交换色谱、SephadexG 75凝胶过滤和制备电泳等分离技术纯化蚯蚓纤溶酶 ;利用纤维平板和聚丙烯酰胺凝胶电泳研究pH、温度和胰蛋白酶对蚯蚓纤溶酶纤溶活性的影响。结果分离得到纤溶酶 4种单一组分 :EFE 1、EFE 2、EFE 3、EFE 4 ;分子量分别为 2 70 0 0、2 80 0 0、2 30 0 0、330 0 0 ;4种组分对温度的适应范围较广 ;pH值对其纤溶活性的影响不尽相同 ,但都在pH低于 1.5时纤溶活性完全丧失 ;胰蛋白酶对纤溶酶 4种组分均无降解作用 ,对其纤溶活性无影响。结论纤溶酶口服药制剂宜制成肠溶型

Purification and effect of pH value, temperature and trypsin on the activities of fibrinolytic enzymes from earthworm Eisenia foelide

PurposeTo purify,identify fibrinolytic enzymes from Earthworm and to study on effects of pH value, temperature and trypsin on the activities of EFE.MethodsThe procedure of purification included ammonium sulphate precipitation, ion-exchange chromatography on DEAE-Sepharose CL-6B, Sephadex G-75 gel filtration and preparative PAGE electropheresis. Effects of pH value, temperature and trypsin on the activities of four components of EFE were studied with fibrinous plate and PAGE.ResultsFour components of fibrinolytic enzyme (EFE-1,EFE-2,EFE-3,EFE-4) were isolated and purified from earthworm Eisenia foelide. Four purified components which all showed fibrinolytic activities on fibrinous plate were proved to be homogeneous in SDS-PAGE electropheresis with molecular weights 27 000,28 000,23 000 and 33 000 respectively; The effects of pH value on the activities of four components of EFE were different. The fibrinolytic activities of four components were lost completely when pH value was below 1.5. The activities of four components of EFE were stable within -25℃～50℃; trypsin had no effect on activities of four components of EFE.ConclusionMedicine made from EFE should be dissolved in intestines.