GENETIC CONTROL OF NITRATE RESPIRATION IN E COLI

肠杆菌使用多种化合物作为末端电子受体 用于呼吸。氧优先于所有其他受体使用，并且 硝酸盐是优选的厌氧受体。分级调控 确保适当的呼吸酶系统的合成， 对电子受体可用性变化的反应。因此，在 厌氧生长，硝酸盐诱导合成甲酸酯酶-N， 硝酸还原酶，并抑制呼吸酶的合成， 代谢不太优选的电子受体，如亚硝酸盐， 富马酸盐。 该资助支持的先前工作已经确定了两个相互作用的 控制转录调节的组分调节系统 呼吸酶合成对硝酸盐及其还原的响应 产品，亚硝酸盐。双传感器（NARX和NARQ蛋白）监测 硝酸盐和亚硝酸盐的有效性，以及磷酸化或脱磷酸化 双重反应调节因子（NARL和NARP蛋白）。 本研究的长期目标是了解 这些平行的调节系统协调基因表达， 对硝酸盐和亚硝酸盐可用性的反应。顺式作用调节 NARL激活的narG和fdnG操纵子的位点先前已经被 表征了这项研究将表征顺式作用位点， NARP激活操纵子，aeg-46.5。与NARL激活的比较 调控序列将揭示DNA中的相似性和差异 通过同源反应调节因子NARL和NARP结合。突变 传感器NARX和NARQ的生物化学分析， 监管机构NARL和NARP，将有助于定义传感器响应调节器 相互作用涉及积极和消极的调节。 另外，该分析将对传感器对信号的识别进行探索 分子（硝酸盐和亚硝酸盐），以及响应调节器识别 特异性DNA结合序列。NARL和NARP蛋白将被 在体外分析特定的蛋白质-DNA相互作用。 最后，将解剖硝酸盐和亚硝酸盐信号的生理学 通过监测靶操纵子诱导或阻遏的速率， 响应于信号分子的不同浓度和比率， 硝酸盐和亚硝酸盐。 总的来说，这项研究将促进我们对厌氧和 生理学和代谢以及细菌信号转导途径。

Enterobacteria use a variety of compounds as terminal electron acceptors for respiration. Oxygen is used in preference to all other acceptors, and nitrate is the preferred anaerobic acceptor. Hierarchical regulation ensures that appropriate respiratory enzyme systems are synthesized in response to changing availability of electron acceptors. Thus, during anaerobic growth, nitrate induces synthesis of formate dehydrogenase-N and nitrate reductase, and represses synthesis of respiratory enzymes that metabolize less preferred electron acceptors, such as nitrite and fumarate. Previous work supported by this grant has identified interacting two component regulatory systems that control transcriptional regulation of respiratory enzyme synthesis in response to nitrate and its reduction product, nitrite. Dual sensors (the NARX and NARQ proteins) monitor the availability of nitrate and nitrite, and phosphorylate or dephosphorylate dual response regulators (the NARL and NARP proteins) accordingly. The long-term objective of this research is to understand the action of these parallel regulatory systems to coordinate gene expression in response to nitrate and nitrite availability. The cis-acting regulatory sites for the NARL-activated narG and fdnG operons have been previously characterized. This research will characterize the cis-acting sites for a NARP-activated operon, aeg-46.5. Comparison with the NARL-activated regulatory sequences will reveal similarities and differences in DNA binding by the homologous response regulators, NARL and NARP. Mutational and biochemical analysis of the sensors NARX and NARQ, and of the response regulators NARL and NARP, will help define sensor-response regulator interactions involved in both positive and negative regulation. Additionally, this analysis will probe sensor recognition of signal molecules (nitrate and nitrite), and response regulator recognition of specific DNA binding sequences. The NARL and NARP proteins will be analyzed in vitro with respect to specific protein-DNA interactions. Finally, the physiology of nitrate and nitrite signaling will be dissected by monitoring the rate of target operon induction or repression in response to varying concentrations and ratios of the signal molecules, nitrate and nitrite. Overall, this research will advance our understanding of both anaerobic physiology and metabolism, and of bacterial signal transduction pathways.