Assemblying an Understanding of Ribosome Biosynthesis

理解核糖体生物合成

• 批准号: 7362185
• 负责人: Gloria M Culver
• 金额: $ 8.22万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7362185
• 关键词: Escherichia coli; SDS polyacrylamide gel electrophoresis; bacterial RNA; bacterial proteins; biosynthesis; chemical aggregate; electrospray ionization mass spectrometry; gene deletion mutation; intermolecular interaction; microorganism culture; nucleic acid reconstitution; nucleic acid structure; protein folding; protein reconstitution; recombinant proteins; ribosomal RNA; ribosomal proteins; ribosomes

DESCRIPTION (provided by applicant): Approximately 40% of the total energy production of E. coli is consumed to synthesize the large number of ribosomal components: 3 RNAs (comprised of more than 400 nucleotides) and greater than 50 proteins. This observation and the direct correlation between the rates of E. coli growth and ribosome biogenesis indicate that an accurately assembled and functional ribosome is of utmost importance for cell viability. Our long term goal is to gain a detailed understanding of ribosome biogenesis and thus reveal the impact of this dynamic process on fundamental aspects of cell physiology, such as growth regulation. This proposal details studies of E. coli small (SOS) ribosomal subunit biogenesis. The purpose of this work is to gain an in-depth understanding of conformational changes and the factors that modulate these changes during the course of functional SOS subunit assembly. Three specific aims have been proposed to address these questions. First, conformational changes in 16S rRNA and the ribosomal proteins that facilitate these changes during different stages of assembly will be identified using a panel of chemical probing strategies. Second, using a combination of genetics and biochemistry, novel SOS subunit assembly intermediates formed in vivo will be identified and characterized. Two ribosomal protein mutant strains, including a chromosomal deletion of ribosomal protein S15, are in hand for this analysis. Third, extra- ribosomal factors that facilitate SOS subunit assembly will be analyzed. Factors identified in the current funding period will be explored in detail and new methods will lead to the identification and characterization of additional factors. Our progress during the current funding cycle and our preliminary data, both biochemical and genetic, indicate that all three specific aims will be successful and thus will directly impact our understanding of SOS subunit biogenesis. Such an understanding is critical for the identification of novel targets for attenuating bacterial growth and therefore for the development of antimicrobials.

描述（由申请人提供）：约40%的总能量生产的E。大肠杆菌被消耗来合成大量的核糖体成分：3个RNA（由400多个核苷酸组成）和50多个蛋白质。这一观察结果与E.大肠杆菌生长和核糖体生物发生表明，一个准确组装和功能性的核糖体是细胞活力的最重要的。我们的长期目标是详细了解核糖体生物发生，从而揭示这种动态过程对细胞生理学基本方面的影响，如生长调节。本文详细介绍了E.大肠杆菌核糖体小亚基（SOS）的生物合成。这项工作的目的是深入了解的构象变化和调节这些变化的因素，在功能SOS亚基组装过程中。为解决这些问题提出了三个具体目标。首先，构象变化的16S rRNA和核糖体蛋白，促进这些变化在不同阶段的装配将被确定使用一组化学探测策略。第二，使用遗传学和生物化学的组合，新的SOS亚基组装体在体内形成的中间体将被识别和表征。两个核糖体蛋白突变株，包括核糖体蛋白S15的染色体缺失，是在手进行这项分析。第三，我们将分析促进SOS亚基组装的核糖体外因子.将详细探讨在本供资期间查明的因素，并采用新的方法查明其他因素并确定其特征。我们在目前的资金周期中取得的进展以及我们的初步数据，包括生物化学和遗传学，表明所有这三个具体目标都将取得成功，因此将直接影响我们对SOS亚基生物起源的理解。这种理解对于鉴定用于减弱细菌生长的新靶标以及因此对于开发抗菌剂至关重要。