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TRANSCRIPTION ACTIVATION AT THE RHABAD OPERON

RHABAD 操纵子的转录激活

基本信息

批准号：
6386644
负责人：
SUSAN M EGAN
金额：
$ 10.89万
依托单位：
UNIVERSITY OF KANSAS LAWRENCE
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-08-01 至 2003-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6386644
关键词：
DNA binding protein DNA directed RNA polymerase Escherichia coli bacterial genetics cytoskeletal proteins gene expression gene induction /repression genetic promoter element genetic transcription molecular cloning mutant nucleic acid sequence operon protein protein interaction protein sequence protein structure function site directed mutagenesis transcription factor

项目摘要

A model for transcription activation has recently been developed based on the structure of E. Coli RNA polymerase, and analysis of activation by the CRP protein. In this model, the activator protein directly contacts the RNA polymerase alpha or subunit, resulting in increased transcription initiation. The long range goal of this research is to understand the molecular mechanisms of transcription activation, especially where multiple activator proteins simultaneously regulate expression of a gene(s). The objective of this proposal is to investigate the details of transcription activation, utilizing the E. coli L-rhamnose rhaBAD genes as a model. Full transcription of rhaBAD requires two activator proteins, RhaS and CRP. The central hypothesis is that while activator-RNA polymerase interactions likely contribute to transcription activation, activator-activator and activator-DNA interactions may also be important. In order to understand how transcription activation occurs, it is essential to determine the nature of the protein-protein and protein-DNA interactions that are required. Three specific aims will be pursued to accomplish the objectives of this proposal: 1) characterize DNA binding by RhaS; 2) characterize transcription activation by RhaS; and 3) characterize transcription activation by CRP. The role of protein-DNA interactions will be investigated by isolating mutations which identify the amino acid-base contacts between RhaS and DNA. To understand the role of protein-protein interactions in activation, mutations will be isolated that identify the specific amino acids in RhaS and CRP that are responsible for transcription activation. Once activation defective mutations are identified in RhaS and CRP, second site suppressors will be isolated. The second site suppressor mutations will identify the protein-protein interactions that are important for transcription activation. The expectation is that this research will identify a variety of interactions that contribute to transcription activation. This work is significant because it will contribute to a broader understanding of the mechanisms used for activation, especially with multiple activators.

最近开发了转录激活模型基于大肠杆菌RNA聚合酶的结构，并分析由CRP蛋白激活。在该模型中，激活蛋白直接接触RNA聚合酶α或亚基，导致增加转录起始。本次活动的长远目标研究是为了了解转录的分子机制激活，特别是在多个激活蛋白的情况下同时调节基因的表达。的目标这个提案是为了研究转录激活的细节，利用大肠杆菌 L-鼠李糖 rhaBAD 基因作为模型。满的 rhaBAD 的转录需要两种激活蛋白：RhaS 和 CRP。中心假设是，虽然激活剂-RNA 聚合酶相互作用可能有助于转录激活，激活剂-激活剂和激活剂-DNA相互作用也可能是重要的。为了了解转录激活如何发生这种情况时，必须确定蛋白质-蛋白质的性质以及所需的蛋白质-DNA 相互作用。为实现以下目标，将努力实现三个具体目标该提案：1) 表征 RhaS 与 DNA 的结合； 2）表征 RhaS 的转录激活； 3) 表征 CRP 的转录激活。蛋白质-DNA的作用将通过分离突变来研究相互作用识别 RhaS 和 DNA 之间的氨基酸碱基接触。到了解蛋白质-蛋白质相互作用在激活中的作用，将分离出识别特定氨基酸的突变 RhaS 和 CRP 负责转录激活。一旦在 RhaS 中鉴定出激活缺陷突变， CRP、第二位点抑制子将被分离。第二个站点抑制突变将识别蛋白质-蛋白质相互作用这对于转录激活很重要。期望是这项研究将确定各种相互作用有助于转录激活。这项工作意义重大因为这将有助于更广泛地了解用于激活的机制，尤其是多个激活器。

项目成果

期刊论文数量（13）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The AraC/XylS family activator RhaS negatively autoregulates rhaSR expression by preventing cyclic AMP receptor protein activation.

AraC/XylS 家族激活剂 RhaS 通过阻止环 AMP 受体蛋白激活来负向自动调节 rhaSR 表达。

DOI：
10.1128/jb.00829-08
发表时间：
2010
期刊：
Journal of bacteriology
影响因子：
3.2
作者：
Wickstrum,JasonR;Skredenske,JeffM;Balasubramaniam,Vinitha;Jones,Kyle;Egan,SusanM
通讯作者：
Egan,SusanM

Functional consequences of exchanging domains between LacI and PurR are mediated by the intervening linker sequence.

LacI 和 PurR 之间交换结构域的功能后果是由插入的接头序列介导的。