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

描述（由申请人提供）：我们建议继续我们的遗传学，生理学和生物化学的NRI-NRII双组分信号转导系统，在控制氮同化在大肠杆菌中起着重要作用的研究。杆菌该系统提供了一个通过碳和氮状态信号调节中枢代谢的范例。我们对NRI/NRII系统的研究也为理解双组分信号转导系统提供了一个范例。双组分信号转导系统是细菌中最常见的信号转导系统，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、对环境应激的众多反应以及代谢的各个方面。我们的工作也将为自然界中分布最广泛的信号转导蛋白PII蛋白的信号转导提供一个范例。我们的研究探讨了PII蛋白如何整合不同的信号并控制信号转导酶受体。因此，拟议的研究将直接研究自然界中广泛存在的信号转导机制，并对人类健康和福祉产生重大影响。 本论文的主要工作是对大肠杆菌NRI和NRII蛋白的结构和功能进行研究。杆菌这些方法使用各种遗传、生物化学和生物物理方法，旨在提供有关调控机制的详细信息。具体目标包括：（1）确定PII调节NRII激酶和磷酸酶活性的机制，和（2）确定NRI“自磷酸酶”活性的机制，以及NRII和PII的复合物对其的控制。在两个特定目标中，生物化学方法用于鉴定信号蛋白的相互作用表面、活性的化学性质以及导致催化活性调节的蛋白质结构域的相互作用。遗传学方法用于定义蛋白质的功能，并将特定功能映射到蛋白质的区域。生物物理方法用于确定蛋白质的结构。鉴于它们在自然界中的广泛存在以及与许多公共卫生和农业问题的直接联系，了解双组分系统和PII信号蛋白所使用的信号转导机制是一个重要的目标。我们的研究与模型系统应该允许快速进展。

DESCRIPTION (provided by applicant): 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 PII 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.