Chemical Approaches to Characterizing Kinase-catalyzed Modifications

描述(申请人提供)：磷酸化是一种普遍存在的翻译后蛋白质修饰，在各种细胞功能中发挥关键作用。事实上，针对磷酸化蛋白底物的激酶的各种药物已经开发出来，用于治疗从癌症到阿尔茨海默病的各种疾病。因此，监测细胞中的激酶及其磷酸蛋白底物是表征健康和疾病状态以及开发新药的重要目标。不幸的是，现有的监测细胞磷酸化的方法是具有挑战性的。为了充分了解细胞信号和疾病的形成，有必要采用新的方法来监测和表征激酶及其修饰的底物。这个项目的长期目标是严格表征激酶的活性和激酶催化的修饰。该项目的科学基础是我们最近揭示的，即激酶混杂地接受β-磷酸修饰的ATP类似物作为辅助底物。基于激酶-ATP共底物的混杂，我们正在追求两个重要目标。首先，我们假设，除了磷酸化外，激酶还催化蛋白质的修饰。我们的假设是基于这样一个事实，即几个？-磷酸修饰的ATP类似物是自然产生的，并与细胞应激条件有关，尽管它们的作用机制尚不清楚。与我们的假设一致，我们最近发现了一种新的翻译后蛋白质修饰，称为核苷酸磷酸化。由于它们在细胞信号转导中的关键作用，这些研究对于阐明激酶在细胞生物学中的整体作用至关重要。我们的第二个重要目标集中在用各种功能探针标记激酶底物的实验上，以便能够检测、序列识别和表征激酶催化的修饰。通过使用开发的化学方法来监测和表征激酶在健康和患病组织中的作用，整个项目的重大成果是更好地理解了信号在疾病形成中的作用。由于激酶酶是药物开发的靶点，这些研究将导致创造出更有效的影响蛋白质磷酸化的药物。与公共卫生相关：激酶与各种疾病有关，是多种药物的靶标。磷酸化蛋白底物，使磷蛋白的特性成为健康相关研究的一个重要方面。该申请概述了表征激酶底物和激酶催化修饰的新方法，以帮助了解疾病和开发新的治疗方法。

DESCRIPTION (provided by applicant): Phosphorylation is a ubiquitous post-translational protein modification that plays a critical role in a variety of cellular functions. In fact, various drugs targeting the kinases that phosphorylate protein substrates have been developed to treat diseases from cancers to Alzheimer's disease. As a result, the monitoring of kinases and their phosphoprotein substrates in cells represents an important goal towards characterizing healthy and disease states and development of new drugs. Unfortunately, available methods to monitor cellular phosphorylation are challenging. New methods for monitoring and charactering kinases and their modified substrates are necessary to allow a full understanding of cellular signaling and disease formation. The long-term goal of this project is a rigorous characterization of kinase activity and kinase-catalyzed modifications. The scientific foundation of the project is our recent revelation that kinases promiscuously accept ?-phosphate modified ATP analogs as cosubstrates. Based on kinase ATP cosubstrate promiscuity, we are pursuing two significant aims. First, we hypothesize that kinases catalyze protein modifications in addition to phosphorylation. Our hypothesis is based on the fact that several ?-phosphate modified ATP analogues are naturally occurring and associated with cell stress conditions, although their mechanisms of action are unknown. Consistent with our hypothesis, we have recently discovered a novel post-translational protein modification, termed nucleotide phosphorylation. With their key role in cell signaling, these studies are critical to illuminate the overall role of kinases in cell biology. Our second significant aim centers on experiments to label kinase substrates with various functional probes to allow detection, sequence identification and characterization of the kinase-catalyzed modifications. By using the developed chemical approaches to monitor and characterize the role of kinases in healthy and diseased tissues, the significant outcome of the overall project is a greater understanding of signaling in disease formation. Because kinase enzymes are targeted in drug development, these studies will lead in the creation of more effective drugs influencing protein phosphorylation. PUBLIC HEALTH RELEVANCE: Kinase enzymes are involved in various diseases and are the target of multiple pharmaceutical drugs. Kinases phosphorylate protein substrates, making a characterization of phosphoproteins an important aspect of health-related research. The application outlines new approaches to characterizing kinase substrates and kinase-catalyzed modifications to aid in the understanding of disease and development of new treatments.