ANALYSIS OF THE HEAT SHOCK RESPONSE IN E. COLI

大肠杆菌的热激反应分析

• 批准号: 3130940
• 负责人: Costa Georgopoulos
• 金额: $ 19.63万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-05-01 至 1994-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3130940
• 关键词: Escherichia coli; bacterial genetics; bacterial proteins; environmental stressor; gene expression; genetic library; genetic mapping; microorganism growth; molecular cloning; mutant; stress proteins; temperature; transposon /insertion element

The goal of this proposal is to understand the physiology of the heat shock (hs) response in Escherichia coli. We will investigate the mechanism of induction and regression of the response and in addition, the exact roles of the hs proteins both in normal bacterial growth and in protection of the cells at high temperature. Because of the tremendous conservation of the hs response throughout the living world, it is expected that many of the conclusions reached by the proposed studies on E. coli will contribute to increasing our understanding of the regulation and the significance of the hs response in all organisms. We will address several specific areas of research. (a) We will characterize the genes coding for the unidentified hs proteins by cloning, mapping and isolating mutations in them. The roles of these genes in E. coli physiology will be studied first by analyzing the phenotypes of mutants in both normal and hs conditions. The gene products will be purified and analyzed biochemically. (b) We will study the regulation of the hs response at the molecular level by investigating the roles of the htpR, dnaK, and other hs proteins in the modulation of the hs response. We will do this by the isolation of many mutants altered in the genes of interest, examination of their various phenotypes, and isolation of intergenic suppressors of the original mutations. Our biochemical approach will be to purify the wild-type and mutant proteins and also to study hs gene expression in vitro using standard techniques for transcription or coupled transcription and translation. (c) Bacteriophage lambda infection results in the induction of the hs response of E. coli in the absence of a temperature shift. We will identify the bacteriophage gene(s) responsible for this effect by analysis of mutants, purify their products, analyze the effects of the proteins in the in vitro system described above. This system may provide insight into the mechanisms by which animal viruses, such as adenovirus, polyoma, and SV40, induce the synthesis of certain host hs proteins during infection.

这个建议的目的是了解热休克的生理学 (hs)在大肠杆菌中的反应。 我们将研究 诱导和回归的反应，此外，确切的作用 的hs蛋白在正常细菌生长和保护的 高温下的细胞 由于大量的自然保护， HS的反应在整个生活世界，预计许多 本文对E.大肠杆菌将有助于 增加我们对法规的理解和 所有生物体的反应。 我们将讨论几个具体的研究领域。 (a)我们将 通过克隆表征编码未鉴定的hs蛋白的基因， 绘制并分离其中的突变。 这些基因在E. 大肠杆菌生理学将首先通过分析 在正常和HS条件下的突变体。 基因产物将 进行了纯化和生化分析 (b)我们会研究规管 在分子水平上的hs反应，通过调查的作用， htpR、dnaK和其他hs蛋白在调节hs反应中的作用。 我们 将通过分离许多突变体来做到这一点，这些突变体在基因中发生了改变， 感兴趣，检查其各种表型，并分离 原始突变的基因间抑制因子。 我们的生化方法 将纯化野生型和突变蛋白，并研究hs 使用用于转录的标准技术进行体外基因表达，或 结合转录和翻译。 (c)λ噬菌体感染 诱导E.大肠杆菌中， 温度变化 我们将确定负责的噬菌体基因 通过分析突变体，纯化其产物，分析 蛋白质在上述体外系统中的作用。 这 该系统可以提供对动物病毒， 如腺病毒、多瘤病毒和SV40，诱导某些宿主合成 HS蛋白在感染过程中的作用