Chemical approaches for detecting S-nitrosothiols

描述（由申请人提供）：一氧化氮是一种细胞信号分子，参与许多生理和病理生理过程。一氧化氮对半胱氨酸残基的修饰，即S-亚硝化，改变了多种蛋白质的功能。这个反应代表了一个重要的翻译后修饰，转导一氧化氮依赖的信号。然而，生物样品中s -亚硝化的检测和定量仍然是一个挑战，因为s -亚硝化产物：s -亚硝基硫醇（RSNOs）的不稳定性。我们小组最近开发了一系列新的RSNOs反应，包括双结扎、还原性结扎和还原性消除（形成脱氢丙氨酸）。这些反应选择性地靶向RSNOs并将其转化为稳定/可检测的产物。在对照实验的基础上，我们假设这些反应可以用于设计检测rsno的新方法。在本项目中，我们计划实现以下四个具体目标：(1)研究基于双连接的蛋白质s-亚硝化检测方法；(2)研究基于还原连接的蛋白质s -亚硝化检测方法；(3)研究S-亚硝基半胱氨酸脱氢丙氨酸的形成及其在蛋白质S-亚硝化检测中的应用；(4)验证新的RSNO检测方法在生物系统中的应用。这些研究将促进对RSNOs的化学和生物学特性的理解，并允许在复杂系统中研究s -亚硝化相关的一氧化氮信号转导。

DESCRIPTION (provided by applicant): Nitric oxide is a cell-signaling molecule involved in a number of physiological and pathophysiological processes. Modification of cysteine residues by nitric oxide, i.e. S- nitrosation, changes the function of a broad spectrum of proteins. This reaction represents an important post-translational modification that transduces nitric oxide- dependent signals. However, the detection and quantification of S-nitrosation in biological samples remains a challenge because of the lability of the S-nitrosation products: S-nitrosothiols (RSNOs). Our group recently developed a series of new reactions for RSNOs, which include bis-ligation, reductive ligation, and reductive elimination (to form dehydroalanine). These reactions selectively target on RSNOs and convert them to stable/detectable products. Based on control experiments, we hypothesize these reactions can be used to design new methods for the detection of RSNOs. In this project, we plan to pursue the following four Specific Aims: (1) to study bis-ligation based methods for detecting protein S-nitrosation; (2) to study reductive ligation based methods for detecting protein S-nitrosation; (3) to study dehydroalanine formation from S-nitrosocysteines and the applications in the detection of protein S- nitrosation; and (4) to validate the applications of new RSNO detection methods in biological systems. These studies will advance the understanding of the chemical and biological properties of RSNOs and permit studies of S-nitrosation-related nitric oxide signal transducation in complex systems.