SENSORY TRANSDUCTION IN BACTERIA

我们的研究重点是大肠杆菌趋化作用的机制 对氧气(趋气性)和相关反应，如氧化还原趋向性 还有甘油出租车。他们的反应与其他人不同 需要功能性电子传递的趋化行为 系统。Aer是一种新的黄素蛋白，TSR是丝氨酸 化学感受器，最近被本实验室鉴定为 氧、氧化还原和甘油(能量)趋向性的传感器。 长期目标是从分子细节上理解 由传感器进行信号处理。具体目标包括： 1)检验黄素腺嘌呤二核苷酸(FAD)在 AER Senses Redox Change发布日期：5/07/1997打印日期： 1997年10月22日，在电子传输系统中，TSR探测到质子 动力。2)研究结构与结构之间的关系 Aer和TSR在氧化还原电位和质子传感中的作用 动力。3)检验质子动机的假设 力/氧化还原电位是恐空症和 在大肠杆菌中的嗜氧性反应，引导细菌达到最佳 质子动力最大的环境。一个 跨学科方法-结合当代方法 分子生物学、遗传学和生物化学与新颖 在本实验室开发的技术-将用于解决 为每个目标确定的关键研究问题。这个 电子传递系统将被扰乱在大肠杆菌中使用新的 可用构件。Aer中Fad结合的假定残基 将通过定点突变而发生突变，并对 趋空性、FAD结合和中间点还原潜力 下定决心。Aer在膜中的拓扑结构将是 使用三明治Aer融合蛋白确定，在该融合蛋白中PhoA或 LacZ被插入Aer的N端域内的帧中。 半胱氨酸扫描H328，精选组氨酸残基 K1盘状结构域周围的胞质结构域和残基 将被用来识别Ph受体及其对 TSR质子动力(氧化还原)传感。假设 趋氧性将细菌引导到支持 最高的质子推动力将用同步 氧气浓度和膜电位的测量， 在氧气浓度可以很紧的情况下 控制住了。阐明这些机制将提供重要的 对趋化的原始途径的洞察，以及对 氧气传感的一般原则(其中一些可能是 适用于人类颈动脉小体化学感受器)。

Our studies focus of the Escherichia coli mechanism of chemotaxis to oxygen (aerotaxis) and related responses, such as redox taxis and glycerol taxis. The responses are different from other chemotactic behaviors in requiring a functional electron transport system. Aer, a novel flavoprotein, and Tsr, the serine chemoreceptor, have been recently identified by this laboratory as the transducers for oxygen, redox and glycerol (energy) taxis. The long term goal is to understand, in molecular detail, the signal processing by thee transducers. The specific aims include: 1) Test the hypothesis that flavin adenine dinucleotide (FAD) in Aer senses redox changes Date Released:05/07/1997 Date Printed: 10/22/1997 in the electron transport system, and Tsr senses proton motive force. 2) Investigate the relationship between structure and function of Aer and Tsr in sensing redox potential and proton motive force. 3) Test the hypothesis that proton motive force/redox potential is a signal that is common to aerophobic and aerophilic responses in E. Coli, guiding bacteria to an optimal environment where the proton motive force is maximal. An interdisciplinary approach - combining contemporary methods of molecular biology, genetics and biochemistry with novel techniques developed in this laboratory - will be used to address the critical research questions identified for each aim. The electron transport system will be perturbed in E. Coli using newly available constructs. Putative residues for FAD binding in Aer will be mutated by site directed mutagenesis and the effect on aerotaxis, FAD binding and mid-point reduction potential determined. The topology of Aer in the membrane will be determined using a sandwich Aer fusion protein in which PhoA or LacZ is inserted in frame within the N-terminal domain of Aer. Cysteine scanning of H328, selected histidine residues in the periplasmic domain and residues around the K1 coiled coil domain of Tsr will be used to identify the Ph receptors and their effect on proton motive force (redox) sensing by Tsr. The hypothesis that aerotaxis guides bacteria to the oxygen concentration that supports the highest proton motive force will be tested using simultaneous measurement of oxygen concentration and membrane potential, under conditions where the oxygen concentration can be tightly controlled. Elucidating these mechanisms will provide important insights into a primordial pathway for chemotaxis, and also into general principles of oxygen sensing (some of which may be applicable to carotid body chemoreceptors in humans).