Characterization of the bacterial Arc system.

说明书(申请人提供)大肠杆菌的弧形双组分信号转导系统根据生长的氧化还原条件调节许多操纵子的表达。该系统包括ArcB作为传感蛋白，ArcA作为反应调节因子。在还原条件下，二聚体ArcB经历依赖于ATP的自动磷酸化，并通过His-Asp-His-Asp磷酸继电器催化ArcA的磷酸化。磷酸化的ArcA抑制大量参与有氧呼吸的操纵子的转录，并激活少量参与发酵的操纵子。D-乳酸、丙酮酸和醋酸盐可刺激ArcB的自磷酸化。在氧化条件下，积累的泛醌或菜醌抑制ArcB的自动磷酸化，从而允许ArcB通过Asp-His-Asp反向继电器催化Arca-P的净去磷酸化。Arc系统的表征将加深我们对细菌如何优化其能量代谢策略的理解。拟议的项目有三个主要目标。第一种方法是在ArcB中定位对苯二酚的接收位置，并定义信号的作用方式。第二个是确定ArcB的两个亚基之间发生了分子间的信号传递和信号衰减的哪一个磷酸转移步骤。最后一个目标是探索胞质PAS结构域在ArcB中的真实作用，该结构域被错误地报道为氧化还原信号的受体部位。这项研究将主要在墨西哥进行，作为NIH赠款#R01 GM40993的延伸。

DESCRIPTION (provided by applicant) The Arc two-component signal transduction system of Escherichia coil modulates the expression of numerous operons according to the redox condition of growth. This system comprises ArcB as the sensor kinase and ArcA as the response regulator. Under reducing conditions, dimeric ArcB undergoes ATP-dependent autophosphorylation and catalyzes the phosphorylation of ArcA by a His-Asp-His-Asp phosphorelay. Phosphorylated ArcA represses transcription of numerous operons involved in aerobic respiration and activates a small number of operons involved in fermentation. D-lactate, pyruvate, and acetate stimulate ArcB autophosphorylation. Under oxidizing conditions, the accumulating ubiquinone or menaquinone inhibits ArcB autophosphorylation, thereby allowing ArcB to catalyze net dephosphorylation of ArcA-P via an Asp-His-Asp reverse relay. Characterization of the Arc system will deepen our understanding of how bacteria optimize their strategy for energy metabolism.The proposed project has three main aims. The first is to locate the quinone reception site in ArcB and define the mode of action of the signal. The second is to determine which phosphotransfer step in signal transmission and signal decay occurs intermolecularly between the two subunits of ArcB. The last objective is to explore the true role of the cytosolic PAS domain in ArcB, erroneously reported to be the receptor site for the redox signal.This research will be done primarily in Mexico as an extension of NIH grant #R01 GM40993.