Purification and characterization of a fibrinolytic enzyme from venom of the southern copperhead snake (Agkistrodon contortrix contortrix).
Purification and characterization of a fibrinolytic enzyme from venom of the southern copperhead snake (Agkistrodon contortrix contortrix).
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南方铜头蛇 (Agkistrodon contortrix contortrix) 毒液中纤溶酶的纯化和表征。
DOI：
10.1016/0003-9861(91)90462-r
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发表时间：
1991
影响因子：
3.9
通讯作者：
MarklandJr,FS
中科院分区：
文献类型：
--
作者：
Guan,AL;Retzios,AD;Henderson,GN;MarklandJr,FS
用CM-纤维素层析、SephadexG-100分子筛层析、对氨基苯甲脒-琼脂糖亲和层析和DEAE-cellulose层析，从南方铜斑蛇（Agkistrodon contorrectum contorrectum，Southern copperhead）蛇毒中分离纯化了一种纤溶酶。纤维蛋白酶的分子量为23，000 - 24，000，等电点为pH6.8。它由大约200个氨基酸组成，具有封闭的NH 2末端，含有很少或不含碳水化合物。该酶对一系列显色对硝基苯胺底物无活性，不受二异丙基氟磷酸盐、大豆胰蛋白酶抑制剂、特雷西洛或对氯汞苯甲酸盐的抑制。然而，该酶是一种金属蛋白酶，因为它被EDTA、邻菲咯啉和四亚乙基五胺（一种特定的锌螯合剂）抑制。金属分析显示1摩尔锌/摩尔蛋白质。使用胰岛素的氧化B链的纤溶酶的切割位点偏好的研究揭示，特异性与主要针对X-Leu键的切割的其他蛇毒金属蛋白酶相似。有趣的是，与其他一些毒液纤溶金属蛋白酶不同，纤溶酶几乎没有出血活性。该酶具有直接的纤维蛋白溶解活性，不激活纤溶酶原。体外研究表明，纤维蛋白水解酶溶解由纯化的纤维蛋白原或全血制成的凝块。
A fibrinolytic enzyme present inAgkistrodon contortrix contortrix(southern copperhead) venom has been purified by combination of CM-cellulose chromatography, molecular sieve chromatography on Sephadex G-100,p-aminobenzamidine-agarose affinity chromatography, and DEAE-cellulose chromatography. The enzyme, fibrolase, has a molecular weight of 23,000–24,000 and an isoelectric point of pH 6.8. It is composed of approximately 200 amino acids, possesses a blocked NH2-terminus and contains little or no carbohydrate. The enzyme shows no activity against a series of chromogenicp-nitroanilide substrates and is not inhibited by diisopropylfluorophosphate, soybean trypsin inhibitor, Trasylol, orp-chloromercuribenzoate. However, the enzyme is a metalloproteinase since it is inhibited by EDTA,o-phenanthroline and tetraethylenepentamine (a specific zinc chelator). Metal analysis revealed 1 mol of zinc/mol of protein. Study of cleavage site preference of the fibrinolytic enzyme using the oxidized B chain of insulin revealed that specificity is similar to other snake venom metalloproteinases with cleavage primarily directed to an X-Leu bond. Interestingly, unlike some other venom fibrinolytic metalloproteinases, flbrolase exhibits little if any hemorrhagic activity. The enzyme exhibits direct fibrinolytic activity and does not activate plasminogen.In vitrostudies revealed that flbrolase dissolves clots made either from purified fibrinogen or from whole blood.