MECHANISM OF INDUCTION OF VIRAL AND CELLULAR FUNCTIONS

病毒和细胞功能的诱导机制

• 批准号: 3272359
• 负责人: DAVID W MOUNT
• 金额: $ 19.65万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-01-01 至 1990-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3272359
• 关键词: DNA repair; DNA replication; Escherichia coli; bacterial genetics; bacteriophage lambda; cell growth regulation; gene expression; gene mutation; genetic manipulation; genetic transcription; microorganism metabolism; mutant; peptidases; protein sequence; regulatory gene; ultraviolet radiation

The goal is to analyze in greater detail the molecular interactions which regulate the SOS response in E. coli. First, specific contacts between amino acids in LexA protein and basepairs in the recA operator will be determined by isolating and sequencing mutant repressors which recognize a symmetrically altered, mutant operator sequence but not the wildtype sequence. Additionally, the role of various basepaires in the two binding sites in the Co1E1 operator on the binding of repressor will be analyzed using a rapid method for isolating and sequencing operator mutants in phage M13mp8/P. Second, we will attempt to obtain evidence for an interaction between LexA and RecA proteins that triggers repressor cleavage. A battery of mutant repressors which is liikely to be deficient in such an interaction is becoming available. An attempt will be made to isolate mutant RecA proteins which promote cleavage of these mutant repressors. The sequence changes in both the lexA and recA mutants would be determined to reveal amino acid residues in the proteins that interact and other possible compensating effects. Third, the direct role which activated Rec A protein plays in SOS mutagenesis of phage Lambda will be analysed through measurement of mutagenesis in various recA mutants and attempted isolation of new mutants based upon several current hypotheses. The molecular mechanism of mutagenesis will be analyzed by direct biophysical methods to determine if DNA synthesis plays a role. Fourth, the importance of various functions in the SOS response will be studied through analysis of mutant LexA proteins which allow constitutive expression of some SOS genes while continuing to repress the others. Finally, mutants of E. coli, which show either defective or constituteive induction of SOS functions will be isolated and analyzed to assist in identifying signal molecules and regions in the RecA protein which are involved in recognizing the inducing signal.

目标是更详细地分析分子相互作用 调节大肠杆菌中的 SOS 反应。 一、具体联系方式 LexA蛋白中的氨基酸和recA算子中的碱基对将是 通过分离和测序突变阻遏物来确定 对称改变的突变操纵子序列，但野生型则不然 顺序。 此外，各种碱基对在两种结合中的作用 将分析 Co1E1 操纵子中阻遏物结合位点 使用快速方法分离噬菌体中的操纵子突变体并对其进行测序 M13mp8/P。 其次，我们将尝试获取互动的证据 LexA 和 RecA 蛋白之间触发阻遏蛋白裂解。 一块电池 突变阻遏物很可能缺乏这样的 互动变得可用。 将尝试隔离 突变的 RecA 蛋白促进这些突变阻遏物的裂解。 lexA和recA突变体的序列变化将被确定 揭示相互作用的蛋白质中的氨基酸残基和其他 可能的补偿效应。 三、激活Rec的直接作用 将通过以下方式分析在噬菌体 Lambda 的 SOS 诱变中发挥作用的蛋白质 测量各种recA突变体的诱变并尝试分离 基于几个当前假设的新突变体。 分子 将通过直接生物物理方法分析诱变机制 确定 DNA 合成是否发挥作用。 四、各种的重要性 将通过突变体分析来研究 SOS 反应中的功能 LexA 蛋白允许一些 SOS 基因的组成型表达，同时 继续压制其他人。 最后，大肠杆菌的突变体，显示 SOS 功能的缺陷或构成性诱导将是 分离和分析以协助识别信号分子和区域 RecA 蛋白中参与识别诱导信号。