ABRB BINDING TO DEVELOPMENTALLY CONTROLLED PROMOTERS

枯草芽孢杆菌的abr B蛋白调控着许多基因， 包括一些孢子形成过程所必需的，表达了 在平衡指数增长和 固定相。它既可以起到正向调节作用，也可以起到负调节作用 转录，取决于特定的目标基因。这是一个 DNA结合蛋白，对其控制的启动子具有特异性。这个 这种特殊性的决定因素在很大程度上是未知的，但似乎是 与可以假设的某种类型的三维DNA结构有关 由各种不同的碱基序列组成。我们建议研究 ABRB-DNA相互作用更详细，目的是了解更多 AbrB如何充分区分其控制的启动子和那些启动子 不会的。这些研究将包括在体外获得详细的 AbrB与DNA结合的足迹并将此信息与 通过使用活体足迹技术的细胞内情况 有可能。我们还将分离突变结合位点并在体外构建 删除和插入变体，以进一步界定 承认。将搜索影响AbrB活性的第二个位点突变 因为为了更全面地了解任何其他影响 ABRB介导的调控。在可能的情况下，我们将分析 其他调节蛋白可能对AbrB的活性有影响。作为一种可能 AbrB介导的调节机制是通过其对 启动子的局域化结构，我们将检测AbrB的作用 结合在DNA弯曲的变化上。我们还将发起一项 AbrB蛋白的结晶学分析，最终目标是 将结构与功能联系起来。

The AbrB protein of Bacillus subtilis regulates a number of genes, including some essential for the sporulation process, that are expressed during the transition state between balanced exponential growth and stationary phase. It can act as either a positive or negative regulator of transcription, depending upon the particular target gene. It is a DNA-binding protein with specificity toward the promoters it controls. The determinants of this specificity are largely unknown but appear to be related to some type of three-dimensional DNA structure that can be assumed by a variety of different base sequences. We propose to examine the AbrB-DNA interaction in greater detail with the goal to understand more fully how AbrB discriminates between the promoters it controls and those it does not. These studies will include obtaining detailed In vitro footprints of AbrB binding to DNA and correlating this information to the intracellular situation by the use of in vivo footprinting techniques where possible. We will also isolate mutant binding sites and construct in vitro deletion and insertion variants to further delimit elements necessary for recognition. Second site mutants affecting AbrB activity will be searched for in order to more fully understand any other factors that impinge upon AbrB-mediated regulation. Where possible, we will assay the effects that other regulatory proteins may have upon AbrB activity. As one possible mechanism for AbrB-mediated regulation is through its effect upon the localized structure of the promoter, we will examine the effects of AbrB binding upon changes in DNA bending. We will also initiate a crystallographic analysis of the AbrB protein with the ultimate goal of relating structure to function.