MECHANISM AND REGULATION OF E COLI RNA POLYMERASE

该项目旨在调查有关 大肠杆菌中转录起始的机制和调控。 这些实验的目的是提供一个基本的生化 理解在识别中涉及的几个复杂现象， E.大肠杆菌启动子。 一系列研究将 描述了“封闭络合物”，一种重要的反应中间体， 启动过程。 在体外进行的实验也将 描述一种突变σ亚基的特性， 在体内改变启动子识别。 DNA超螺旋对 启动子识别将使用新开发的检测方法进行探索， 拓扑异构体选择性 这种新的方法将在一项研究中得到推广。 CAP*cAMP对乳糖和半乳糖操纵子的正调控 发起人。 半乳糖对启动子选择性的调节 还将表征Gal启动子处的阻遏物。 结构性 RNA-RNA配对的决定子也将在以下实例中确定： 反义RNA基因调控 在这种情况下， 然而，主要是在翻译开始阶段， 出现的RNA-RNA配对将适用于基因的许多其他步骤 表达及其调控。 我们提出的问题的答案 将有助于对基因调控的生物化学描述 表情

This project is designed to investigate important questions about the mechanism and regulation of transcription initiation in Escherichia coli. The experiments are designed to provide a fundamental biochemical understanding of several complex phenomena involved in the recognition of E. coli promoters by RNA polymerase. A series of studies will characterize the "closed complex", an important reaction intermediate in the initiation process. Experiments performed in vitro will also characterize the properties of a mutant sigma subunit that displays altered promoter recognition in vivo. The effects of DNA supercoiling on promoter recognition will be explored using a newly-developed assay for topoisomer selectivity. This novel approach will be extended in a study of positive regulation by cap*cAMP at the lactose and galactose operon promoters. The modulation of promoter selectivity by the galactose repressor at the gal promoters will also be characterized. The structural determinants of RNA-RNA pairing will also be determined in an example of antisense RNA gene regulation. In this case the regulation occurs primarily at the translation initiation steps, however, the principles for RNA-RNA pairing that emerge will be applicable to many other steps in gene expression and its regulation. The answers to the questions we have posed will contribute to the biochemical description of the control of gene expression.