Analysis of E. coli ribonucleases and RNA metabolism

大肠杆菌核糖核酸酶和 RNA 代谢分析

• 批准号: 8837643
• 负责人: Sidney R. Kushner
• 金额: $ 31.93万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-15 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8837643
• 关键词: Antibiotic Resistance; Bacteria; Biochemical; Bioinformatics; Biological; Cells; Drug Targeting; Enzymes; Escherichia coli; Eukaryota; Family; Gene Expression Profile; Gene Expression Regulation; Generations; Genetic; Genomics; Grant; Knowledge; Messenger RNA; Metabolism; Modeling; Molecular; Pathway interactions; Play; Polyribonucleotide Nucleotidyltransferase; Prevalence; Process; Prokaryotic Cells; Proteins; RNA; RNA Decay; RNA Processing; RNase P; RNase Z; Reaction; Ribonuclease III; Ribonucleases; Ribosomal RNA; Role; Series; Transfer RNA; Work; density; design; endonuclease; mRNA Decay; research study; response; ribonuclease E; tRNA Precursor

DESCRIPTION (provided by applicant): Although numerous ribonucleases have been identified over the past 40 years, most have been associated primarily with the biological reaction that was used for their identification. Thus an enzyme like RNase P has been assumed to be strictly involved tRNA processing. Likewise the RNase Z family of enzymes has also been thought to be only involved in tRNA processing, while RNase III type proteins, at least in prokaryotes are considered rRNA maturation enzymes. However, more recent experiments have demonstrated that most of these ribonucleases have multiple functions in the cell. When one carefully looks at what is known about the pathways of rRNA maturation, tRNA processing and mRNA decay, it becomes clear that many of the existing models for these processes are far too simplistic and in some cases probably incorrect. In addition, not much is known regarding the enzymatic overlap among these important pathways. For example, during the current grant period, we have demonstrated the existence of multiple new pathways for tRNA processing that require either RNase E, RNase P or polynucleotide phosphorylase (PNPase) as the first step in processing tRNA precursorsAccordingly, this application describes a series of experiments that will focus on developing a more complete understanding of post-transcriptional RNA metabolism in the model prokaryote, Escherichia coli. Our approach will be to use a combination of unique bacterial strains, high density tiling microarrays as well as other molecular biological, biochemical and bioinformatic approaches. Specific experiments include: 1. Determine the molecular mechanism(s) of 30S rRNA processing; 2. Comprehensive analysis of tRNA processing pathways; and, 3. Transcriptome-wide analysis of the initiation of mRNA processing and decay. With the increasing prevalence of antibiotic resistant bacteria, the need to better understand the overall mechanism of post-transcriptional RNA metabolism is becoming increasingly important. Information gained from this work could be instrumental in the identification of potential new drug targets.

描述（由申请人提供）： 尽管过去 40 年来已经鉴定出许多核糖核酸酶，但大多数核糖核酸酶主要与其鉴定所用的生物反应有关。因此，像 RNase P 这样的酶被认为严格参与 tRNA 加工。同样，RNase Z 酶家族也被认为仅参与 tRNA 加工，而 RNase III 型蛋白（至少在原核生物中）被认为是 rRNA 成熟酶。然而，最近的实验表明，大多数核糖核酸酶在细胞中具有多种功能。当人们仔细研究 rRNA 成熟、tRNA 加工和 mRNA 衰变途径的已知信息时，就会发现这些过程的许多现有模型都过于简单化，在某些情况下可能是不正确的。此外，对于这些重要途径之间的酶促重叠知之甚少。例如，在当前资助期内，我们已经证明存在多种新的 tRNA 加工途径，需要 RNase E、RNase P 或多核苷酸磷酸化酶 (PNPase) 作为加工 tRNA 前体的第一步。因此，本申请描述了一系列实验，重点是对模型原核生物埃希氏菌中的转录后 RNA 代谢进行更全面的了解 大肠杆菌。我们的方法将结合使用独特的细菌菌株、高密度平铺微阵列以及其他分子生物学、生物化学和生物信息学方法。具体实验包括： 1.确定30S rRNA加工的分子机制； 2. tRNA加工途径综合分析； 3. mRNA 加工和衰变起始的全转录组分析。随着抗生素耐药细菌的日益流行，更好地了解转录后 RNA 代谢的整体机制的需求变得越来越重要。从这项工作中获得的信息可能有助于识别潜在的新药物靶点。