摘要
以新鲜蚯蚓为原料，经过保温抽提、乙醇沉淀、DEAE-SepharoseFastFlow离子交换层析、Lysine-Sepharose4B亲和层析以及SDS-PAGE制备电泳等纯化步聚，得到一种纯度达95％以上的蚯蚓纤溶酶．该酶具有强烈的溶解纤维蛋白的作用及蛋白酶活性，平板法测得其比活性为90OUK单位／毫克蛋白，TAME法测得其比活性为2500O单位／毫克蛋白．酶学性质研究表明其最适反应温度为65℃，最适反应PH值为8．5．该酶的分子量为33kD，等电点为pH3．5．还对该酶进行了氨基酸组成分析，并测定了其N端部分序列．
From fresh earthworm,an fibrinolytic enzyme was purified by incubation,extraction,alcohol precipitation and chromatographies on DEAE-Sepharose Fast Flow and Lysine-Sepharose 4B. Then a purified enzyme was recovered by SDS-PAGE. The enzyme,with a molecular weight of 33 000 and a PI value of 3. 5,is a single chain protein. The activity and effects of temperature and PH on the activity of the enzyme were analyzed. The amino acid composition of the enzyme shows that it is an acidic protein. Sequence of thirteen amino acids at the N-terminus of the protein was determined as following: Ⅱe-Val-Gly-Gly-Ⅱe-Glu-Ala-Arg-Pro-Tyr-Glu-Phe-Pro.
基金
国家八五科技攻关项目!85-08-05-04
关键词
蚯蚓纤溶酶
分离
纯化
序列测定
Earthworm fibrinolytic enzyme, Purification, Sequence determination
