ANALYSES OF SITE SPECIFIC RECOMBINATION

位点特异性重组分析

• 批准号: 2857090
• 负责人: JEFFREY F GARDNER
• 金额: $ 28.13万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-12-01 至 2000-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2857090
• 关键词: DNA binding protein; DNA topoisomerases; bacteriophage lambda; enzyme activity; gel mobility shift assay; gene complementation; gene mutation; genetic recombination; integrase; microorganism genetics; molecular cloning; nucleic acid sequence; protein protein interaction; site directed mutagenesis

Site-specific recombination systems are important in controlling development and gene expression in a diverse array of organisms ranging from bacteria to humans. The long range goal of this project is to understand how bacteriophage X carries-out site-specific recombination. The findings will have a significant impact on other site-specific recombination systems because many of their integrases are related to the lambda Int protein. Biochemical and genetic approaches will be used to characterize the protein-protein and protein-DNA interactions that occur both during the assembly of recombination complexes (intasomes) and the process of strand-cleavage and exchange. The phage-encoded integrase (Int) protein is a central actor in intasome formation and strand exchange. Int mutants will be isolated and characterized in assays that will determine the defects of the individual proteins in the recombination pathway. The host-encoded integration host factor (IHF) also participates in intasome formation by inducing bends in the DNA. We will isolate mutants that have amino acid substitutions of residues that, as suggested by our previous genetic studies, interact with DNA. Such mutants will also provide information that will be useful in interpreting data derived from physical studies on the protein. The phage-encoded excisionase (Xis) and the host- encoded factor for inversion stimulation (FIS) promote excisive recombination. Xis interacts cooperatively with Int and FIS. We will characterize the mechanism(s) of cooperative protein-protein interactions by isolating and characterizing Xis mutants that are defective in interacting with Int or FIS.

位点特异性重组系统在控制中很重要