Diced Electrophoresis Gel Assay for Screening Enzymes with Specified Activities
Diced Electrophoresis Gel Assay for Screening Enzymes with Specified Activities
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DOI：
10.1021/ja401792d
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发表时间：
2013-04-24
影响因子：
15
通讯作者：
Nagano, Tetsuo
中科院分区：
文献类型：
--
作者：
Komatsu, Tom;Hanaoka, Kenjiro;Nagano, Tetsuo
我们已经建立了切割电泳凝胶（DEG）测定作为蛋白质组范围内的筛选工具，以确定酶的活动感兴趣的使用营业额为基础的荧光底物。该方法利用非变性聚丙烯酰胺凝胶电泳（PAGE）与基于多孔板的荧光测定的组合，以发现具有指定活性的蛋白质点。通过开发模拟中性粒细胞化学引诱物结构的荧光底物，我们可以鉴定参与化学引诱物代谢失活的酶。
We have established the diced electrophoresis gel (DEG) assay as a proteome-wide screening tool to identify enzymes with activities of interest using turnover-based fluorescent substrates. The method utilizes the combination of native polyacrylamide gel electrophoresis (PAGE) with a multiwell-plate-based fluorometric assay to find protein spots with the specified activity. By developing fluorescent substrates that mimic the structure of neutrophil chemoattractants, we could identify enzymes involved in metabolic inactivation of the chemoattractants.