Screening for proteolytic activities in snake venom by means of a multiplexing electrospray ionization mass spectrometry assay scheme
Screening for proteolytic activities in snake venom by means of a multiplexing electrospray ionization mass spectrometry assay scheme
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DOI：
10.1002/rcm.2136
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发表时间：
2005-01-01
影响因子：
2
通讯作者：
Karst, U
中科院分区：
文献类型：
--
作者：
Liesener, A;Perchuc, AM;Karst, U
建立了一种同时测定5种不同底物/产物对的多重质谱学分析方法，作为一种筛选蛇毒组分蛋白分解活性的工具。用该方法对8种模型酶进行了功能鉴定。生成了时间分辨的反应曲线，并确定了每个时间点的相对反应进度。它们被用来半定量地将每种酶对各自底物的催化活性分成六类。由此得到的活性模式作为每种酶的活性指纹。然后将多重分析方法应用于从布氏杆菌预分离的蛇毒部分中筛选蛋白水解酶活性。生成每个组分的活性模式，并使用该模式将组分分类为三种不同的活性类别。通过比较蛇毒部分和模型酶的指纹活性模式，发现了一种具有类似活性蛋白C的蛋白分解特性的化合物。版权所有(C)2005 John Wiley&Sons，Ltd.
A multiplexed mass spectrometry based assay scheme for the simultaneous determination of five different substrate/product pairs was developed as a tool for screening of proteolytic activities in snake venom fractions from Bothrops moojeni. The assay scheme was employed in the functional characterization of eight model proteases. Time-resolved reaction profiles were generated and the relative reaction progress at each time point was determined. These were used to semi-quantitatively sort the catalytic activities of each enzyme towards the respective substrates into six classes. The resulting activity pattern served as an activity fingerprint for each enzyme. The multiplex assay scheme was then applied to a screening for proteolytic activities in fractions of the pre-separated venom from B. moojeni. Activity patterns of each fraction were generated and used to sort the fractions into three different categories of activity. By comparison of the fingerprint activity patterns of the venom fractions and the model enzymes, a compound with proteolytic properties similar to activated protein C was detected. Copyright (C) 2005 John Wiley & Sons, Ltd.