Array-based fluorescence assay for serine/threonine kinases using specific chemical reaction
Array-based fluorescence assay for serine/threonine kinases using specific chemical reaction
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DOI：
10.1016/j.bmc.2008.07.007
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发表时间：
2008-08-15
影响因子：
3.5
通讯作者：
Higuchi, Tsunehiko
中科院分区：
文献类型：
--
作者：
Akita, Shoji;Umezawa, Naoki;Higuchi, Tsunehiko
我们在此报告的发展，一个有效的荧光测定丝氨酸/苏氨酸激酶使用肽阵列。我们的方法是基于特定的磷酸丝氨酸和磷酸苏氨酸残基的化学反应，即，碱介导的β-消除磷酸基团和随后的迈克尔加成的含巯基的荧光试剂。该过程使得能够共价引入a。在一些实施方案中，所述多肽包含磷酸化肽中的不饱和部分。小说为此目的设计并合成了荧光试剂。用这些试剂，蛋白激酶A（PKA）和Akt-1的活性很容易检测。我们的方法也可用于测量激酶抑制剂的活性。该方法有望在激酶研究中得到广泛应用。(c)2008年由Elsevier Ltd.出版
We report herein the development of an efficient fluorescence assay for serine/threonine kinases using a peptide array. Our approach is based on chemical reactions specific to phosphoserine and phosphothreonine residues, that is, base-mediated beta-elimination of the phosphate group and subsequent Michael addition of a thiol-containing fluorescent reagent. This procedure enables the covalent introduction of a. uorescent moiety into the phosphorylated peptide. Novel. uorescent reagents were designed for this purpose and synthesized. With these reagents, protein kinase A (PKA) and Akt-1 activities were readily detected. Our method can also be used to measure the activity of kinase inhibitors. This assay is expected to be widely applicable in kinase research. (c) 2008 Published by Elsevier Ltd.