Purification of salmon thrombin and its potential as an alternative to mammalian thrombins in fibrin sealants
Purification of salmon thrombin and its potential as an alternative to mammalian thrombins in fibrin sealants
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DOI：
10.1016/s0049-3848(02)00333-x
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发表时间：
2002-09-01
影响因子：
7.5
通讯作者：
Janmey, PA
中科院分区：
文献类型：
--
作者：
Michaud, SE;Wang, LZ;Janmey, PA
一种从鲑鱼血中生产高纯度凝血酶的方法，一系列的生物化学、细胞生物学和生物物理学试验证明了鲑鱼和人类凝血酶之间的功能相似性和一些差异。比活性大于1000单位/mg总蛋白的盐-非凝血酶可通过用于纯化人凝血酶的方法的改进来制备。使用基于人纤维蛋白原A-α多肽序列的合成底物作为酶活性的指示剂，鲑鱼和人凝血酶制剂含有相似的每质量纯化蛋白的比活性，鲑鱼凝血酶活化人纤维蛋白原并引发纤维蛋白凝块的形成，其结构和流变性质与被人凝血酶凝结的人纤维蛋白的结构和流变性质没有区别，鲑鱼凝血酶也能激活人体血小板。与人凝血酶相比，对于相同的血小板聚集速率，需要大约高10倍的活性，并且相对于人凝血酶的作用，血小板活化的某些方面，最显著的是磷脂酰丝氨酸暴露减少。后一个发现表明鲑鱼凝血酶可能不会激活作为人凝血酶靶点的所有受体，尽管它似乎确实激活了足以产生正常激活和聚集速率的信号，如通过常规聚集测定法所测量的。与最近鲑鱼纤维蛋白原的纯化及其在哺乳动物伤口愈合中的应用一起，鲑鱼凝血酶的可用性允许配制不含任何外源性哺乳动物蛋白的生物密封剂，因此可以帮助设计具有增加的安全性以防止传染病传播的材料。(C)2002爱思唯尔科技有限公司版权所有。
A method to produce highly purified thrombin from salmon blood is described, and a series of biochemical, cell biologic, and biophysical assays demonstrate the functional similarities and some differences between salmon and human thrombins. Salt-non thrombin with specific activity greater than 1000 units/mg total protein can be prepared by modifications of the methods used for purification of human thrombin. Using a synthetic substrate based on the human fibrinogen A-alpha polypeptide sequence as an indicator of enzymatic activity, salmon and human thrombin preparations contain similar specific activities per mass of purified protein, Salmon thrombin activates human fibrinogen and initiates the fort-nation of fibrin clots whose structure and rheologic properties are indistinguishable from those of human fibrin clotted by human thrombin, Salmon thrombin also activates human platelets. Approximately 10 times higher activities are needed for the same rate of platelet aggregation compared to human thrombin, and some aspects of platelet activation, most notably phosphatidylserine exposure, are diminished relative to the effects of human thrombin. This latter finding suggests that salmon thrombin may not activate all of the receptors that are targets of human thrombin, although it does appear to activate signals that are sufficient to produce normal rates of activation and aggregation as measured by conventional aggregometry. Together with the recent purification of salmon Fibrinogen and its application in mammalian wound healing, the availability of salmon thrombin allows the formulation of biological sealants devoid of any exogenous mammalian proteins and so may aid the design of materials with increased safety from infectious disease transmission. (C) 2002 Elsevier Science Ltd. All rights reserved.