Structure/function analysis of the E. coli NIR/NRII signal transduction system

我们建议继续我们对NRI-NRII两个基因的遗传学、生理学和生化的研究- 在控制大肠杆菌氮同化中起主要作用的组成信号转导系统。 该系统为通过碳和氮信号调节中枢新陈代谢提供了范例 状态。我们在NRI/NRII系统上的工作也为理解双分量信号提供了一个范例 转导系统。两组分信号转导系统是最常见的信号类型 细菌中的转导系统，也存在于低等真核生物和植物中。在细菌中，这些 系统控制细胞周期、发育、毒力、趋化性、对环境的众多反应 压力，以及新陈代谢的各个方面。我们的工作还将为信号转导提供一个范例 PII蛋白是自然界分布最广泛的信号转导蛋白，我们的研究 研究PLL蛋白如何整合不同的信号和控制信号转导的受体 酵素。因此，拟议的研究将直接调查信号转导机制，这些机制是 广泛存在于自然界，对人类的健康和福祉有重大影响。 拟议的工作将集中于对大肠杆菌NRI和NRII蛋白的结构/功能的研究。这个 方法使用各种遗传、生化和生物物理方法，旨在提供 关于监管机制的详细信息。具体目标包括：(1)确定 PLL调节NRII激酶和磷酸酶活性的机制，以及(2)测定 NRI“自磷酸酶”活性的机制及其受NRII和PLL复合体的调控。在这两个地方 特定的目的，生化方法被用来识别信号蛋白的相互作用表面， 活性的化学，以及导致催化调节的蛋白质结构域的相互作用 活动。遗传方法被用来定义蛋白质的功能，并将特定的功能映射到 蛋白质的区域。生物物理方法被用来确定蛋白质的结构。 鉴于它们在自然界中广泛存在，并与许多公共卫生和农业 问题，了解双组分系统和PLL使用的信号转导机制 信号蛋白是一个重要的目标。我们用模范系统进行的研究应该会有很快的进展。

We propose to continue our study of the genetics, physiology, and biochemistry of the NRI-NRII two- component signal transduction system that plays a major role in controlling nitrogen assimilation in E. coli. This system provides a paradigm for the regulation of central metabolism by signals of carbon and nitrogen status. Our work on the NRI/NRII system also provides a paradigm for understanding two-component signal transduction systems. Two component signal transduction systems are the most common type of signal transduction system in bacteria and are also present in lower eukaryotes and plants. In bacteria, these systems control the cell cycle, development, virulence, chemotaxis, numerous responses to environmental stress, and various aspects of metabolism. Our work will also provide a paradigm for signal transduction by PII proteins, which are the most widely distributed signal transduction proteins in nature, Our studies investigate how Pll proteins integrate distinct signals and control receptors that are signal-transduction enzymes. Thus, the proposed research will directly investigate signal transduction mechanisms that are widely-occuring in nature and have significant impact on human health and well-being. The proposed work will focus on structure/function studies of the NRI and NRII proteins of E. coli. The approaches use a variety of genetic, biochemical, and biophysical methods and are designed to provide detailed information on the mechanisms of regulation. The Specific Aims include: (1) Determining the mechanism of regulation of NRII kinase and phosphatase activities by Pll, and (2) Determining the mechanism of the NRI "autophosphatase" activity, and its control by the complex of NRII and Pll. In both Specific Aims, biochemical approaches are used to identify interacting surfaces of the signalling proteins, the chemistry of the activities, and the interactions of protein domains that result in regulation of the catalytic activities. Genetic approaches are used to define the functions of the proteins, and map specific functions to regions of proteins. Biophysical approaches are used to determine the structures of proteins. Given their widespread occurance in nature and direct link to numerous public health and agricultural issues, understanding the mechanisms of signal transduction used by two-component systems and Pll signalling proteins is an important objective. Our studies with a model system should allow rapid progress.