ANALYSIS OF THE HEAT SHOCK RESPONSE IN E. COLI

大肠杆菌的热激反应分析

• 批准号: 3130942
• 负责人: Costa Georgopoulos
• 金额: $ 21.37万
• 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/3130942
• 关键词: Escherichia coli; bacterial genetics; bacterial proteins; environmental stressor; gene expression; genetic library; genetic mapping; microorganism growth; molecular cloning; mutant; nucleic acid sequence; 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.

这项建议的目标是了解热休克的生理学。 (HS)在大肠杆菌中的反应。我们将研究其发生机制。 反应的诱导和回归，以及确切的作用 Hs蛋白在正常细菌生长和保护 细胞在高温下。因为极大地保护了 在生活世界各地的房协反应中，预计许多 拟议中的对大肠杆菌的研究得出的结论将有助于 增加我们对这项规定的认识和对 HS在所有生物体中都有反应。 我们将讨论几个具体的研究领域。(A)我们会 通过克隆鉴定编码未知hs蛋白的基因， 绘制并分离它们中的突变。这些基因在E. 首先将通过分析大肠杆菌的表型来研究大肠杆菌的生理学 在正常和hs条件下的突变体。基因产品将是 经过生化提纯和分析。(二)我们会研究规管 通过研究hs在分子水平上的作用来研究hs的反应 HTpR、dNAK等hs蛋白参与hs反应的调节。我们 将通过分离许多改变了基因的突变体来做到这一点 兴趣，检查他们的各种表型，并分离 原始突变的基因间抑制因子。我们的生化方法 将提纯野生型和突变型蛋白，并研究hs 使用标准转录或转录技术进行基因体外表达 将转录和翻译结合起来。(C)噬菌体感染 在缺乏A的情况下诱导大肠杆菌的hs反应的结果 温度变化。我们将鉴定负责的噬菌体基因(S) 为了达到这个效果，通过分析突变体，提纯它们的产物，分析 上述蛋白质在体外系统中的作用。这 系统可以提供对动物病毒、 如腺病毒、多瘤和SV40，可诱导某些宿主合成 感染过程中的HS蛋白。