Structure-Function Relationships in E. coli Response Regulators NarL and NarP

9630809 Dickerson该项目的目标是了解基因表达的双组分调控系统的分子机制，使用控制细菌中硝酸盐呼吸的NarQ/NarX/NarL/ NarP系统。 在这个系统中，NarQ和NarX蛋白感知周围环境中硝酸盐或亚硝酸盐的存在，并通过磷酸化特定的天冬氨酸来激活反应调节分子NarL或NarP。然后，激活的反应调节因子与DNA上的特定位点结合，适当地打开和关闭基因。 该项目已开始与一个成功的NarL晶体结构分析。 计划开展的其他项目有：（1）在不同的晶体环境中完成NarL的结构，考察晶体环境效应，（2）类似但独立作用的NarP的X-射线结构分析，（3）NarL的DNA结合C-末端结构域与其DNA识别序列的复合物的结构分析，（4）NarL的DNA结合C-末端结构域与其DNA识别序列的复合物的结构分析，（5）NarL的DNA结合C-末端结构域与其DNA识别序列的复合物的结构分析。（4）检测NarL和NarP的组成型突变体，它们能抵抗去磷酸化并与DNA强结合;（5）对完整的NarL/DNA和NarP/DNA复合物进行X射线分析。 * 这项研究的意义在于，它将是第一次在分子水平上对双组分传感器/响应系统进行系统研究，其结果适用于细菌双组分控制系统的广泛进化范围。 它还将揭示高等生物中的多组分遗传控制系统，该系统也通过磷酸化关键效应蛋白发挥作用。 总之，NarQ/NarX/NarL/NarP家族包含复杂但灵活的控制系统，其可以响应于硝酸盐和亚硝酸盐的相对浓度来调节若干基因的相对表达。 这个分子反应网络的四组分例子背后的完整逻辑尚未被理解。 这个项目的主要目标是增加对它如何工作以及为什么它必须如此复杂的理解。 %%%

9630809 Dickerson The goal of this project is to understand the molecular mechanism of two-component regulatory systems of genetic expression, using the NarQ/NarX/NarL/ NarP system that controls nitrate respiration in bacteria. In this system, NarQ and NarX proteins sense the presence of nitrate or nitrite in the surrounding environment, and activate response regulator molecules NarL or NarP by phosphorylating a particular aspartic acid. The activated response regulators then bind to specific sites on DNA, turning genes on and off as appropriate. The project has begun with a successful crystal structure analysis of NarL. Further projects planned are: (1) Completion of the NarL structure in a different crystal environment, to examine crystal environment effect; (2) X-ray structure analysis of the similar but independently-acting NarP; (3) Structure analysis of the DNA-binding C-terminal domain of NarL in a complex with its recognition sequence of DNA; (4) Examination of constitutive mutants of NarL and NarP that resist dephosphorylation and bind strongly to DNA; and (5) X-ray analysis of full NarL/DNA and NarP/DNA complexes. *** The significance of this study is that it will be the first systematic investigation at the molecular level of a two component sensor/response system, with results applicable to the broad evolutionary range of bacterial two-component control systems. It will also shed light on multi- component genetic control systems in higher organisms, that also act by phosphorylating key effector proteins. Taken together the NarQ/NarX/NarL/NarP family comprises a complex but flexible control system that can adjust the relative expression of several genes in response to the relative concentrations of nitrate and nitrite. The full logic behind this four-component example of a molecular response network is not yet understood. An increased understanding of how it works and why it must be so complex is the major goal of this project. %%%