REGULATORY MECHANISMS OF THE E COLI ARABINOSE OPERON

我们的长期目标是促进对 基因调控中最重要的基本原则， AraC蛋白调节L-阿拉伯糖操纵子的转录 大肠埃希菌 工作将在基因调控的三个核心领域进行。一是 将研究调节蛋白AraC的结构和功能， 本身将开发和应用遗传和物理方法， 确定和研究蛋白质内的结构域， 连接结构域，并识别与DNA接触的氨基酸。的 AraC的结构域和来自相关蛋白的同源结构域将被 纯化，并进行研究，目的是确定其结构。 阿拉伯地区DNA循环和解循环现象的物理基础 系统将寻求通过研究域，灵活的连接器，和绑定 在存在和不存在阿拉伯糖的情况下蛋白质的位点偏好。 我们的第二个研究领域是确定调节蛋白 影响RNA聚合酶转录。为此，我们将使用遗传学， 体内和体外转录起始测量，以及指导 AraC蛋白和RNA聚合酶之间相互作用的测量。 我们将测试AraC家族中的其他成员， 超过20种，使用相同的DNA结合和转录机制 activation. 我们的第三个研究领域将是对监管的整合， ara基因进入细胞的整体生理。我们将测量 在E. 并将差异与个体差异联系起来 启动子和调控区。我们还将构建和测试 使用阿拉伯糖成分的新调节系统的行为 操纵子及其亲属 从我们的工作中获得的知识应该有助于构建系统 具有指定的监管属性，并使其更容易管理 天然基因系统在细菌、植物、动物中的表达， 病毒

Our long-term objectives are to contribute to the understanding of the basic principles most important in gene regulation by learning how the AraC protein regulates transcription of the L-arabinose operon in Escherichia coli. Work will be done in three areas central to gene regulation. First, we will study the structure and function of the regulatory protein, AraC, itself. Genetic and physical approaches will be developed and applied for determining and studying domains within proteins, linker regions that connect domains, and identifying the amino acids that contact DNA. The domains of AraC and homologous domains from related proteins will be purified, and studied with the objective of determining their structures. The physical basis for DNA looping and unlooping phenomena in the ara system will be sought by studying the domain, flexible linker, and binding site preferences of the protein in the presence and absence of arabinose. Our second research area will be determining how regulatory proteins influence transcription by RNA polymerase. For this, we will use genetics, in vivo and in vitro transcription initiation measurements, and direct measurements of the interaction between AraC protein and RNA polymerase. We will test whether other members in the AraC family, which now numbers more than 20, use the same mechanisms for DNA binding and transcription activation. Our third research area will be on the integration of regulation of the ara genes into the overall physiology of cells. We will measure the in vivo regulatory properties of the five arabinose-regulated promoters in E. coli and relate the differences to the differences in the individual promoters and regulatory regions. We will also construct and test the behavior of new regulatory systems that use components from the arabinose operon and its relatives. The knowledge gained from our work should facilitate constructing systems with specified regulatory properties, and make it easier to manage the expression of natural gene systems in bacteria, plants, animals and viruses.