IDENTIFICATION OF KINASE SUBSTRATES BY COVALENT CAPTURE OF PHOSPHOPEPTIDES

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 我们将优化一种快速鉴定蛋白激酶底物的方法。 在该方法中，首先将蛋白激酶工程化以接受ATP类似物，从而允许其用生物正交磷酸盐类似物独特地标记其底物。 然后，我们将使用高度特异性的共价捕获和释放方法来快速纯化来自标记底物蛋白的标记肽。 最初的工作将集中在该技术的优化，然后应用于几个重要途径中的几种激酶，包括PI 3 K mTOR信号通路。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. We will optimize a method for rapid identification of protein kinase substrates. In this method a protein kinase is first engineered to accept an ATP analog that allows it to uniquely label its substrate with a bio-orthogonal phosphate analog. Then we will use a highly specific covalent capture-and-release methodology to rapidly purify tagged peptides derived from labeled substrate proteins. Initial work will focus on the optimization of this technique, followed by application to several kinases in several important pathways including the PI3K mTOR signalling pathway.