ASSEMBLING AN UNDERSTANDING OF RIBOSOME BIOSYNTHESIS

加深对核糖体生物合成的理解

• 批准号: 6636561
• 负责人: Gloria M Culver
• 金额: $ 20.7万
• 依托单位: IOWA STATE UNIVERSITY
• 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 2006-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6636561
• 关键词: Escherichia coli; SDS polyacrylamide gel electrophoresis; bacterial RNA; bacterial proteins; biosynthesis; chemical aggregate; intermolecular interaction; microorganism culture; nucleic acid reconstitution; nucleic acid structure; protein folding; protein reconstitution; recombinant proteins; ribosomal RNA; ribosomal proteins; ribosomes

DESCRIPTION (adapted from applicant's abstract): Approximately 40 percent of the total energy production of E. coli is consumed to synthesize the large number of ribosomal components, 3 RNAs totaling over 4000 nucleotides and greater than 50 proteins. In bacteria, the rates of cell growth and ribosome biosynthesis are intimately related and directly correlated. Thus, for cell viability, the synthesis of ribosomal components and their subsequent assembly must be coordinated. These observations suggest that an accurately assembled and functional ribosome is one of the most important cellular organelles and that defects in ribosome assembly would be fatal. Although, years of research have been dedicated to studying ribosomal components and ribosome function, the pathway(s) and dynamics of ribosome assembly remain largely uncharacterized despite general importance to cell physiology. This proposal describes three approaches to study the mechanism, pathway and dynamics of ribosome assembly. The goal of this proposal is to understand what factors are involved in and critical for E. coli 30S ribosomal subunit assembly. First, in vitro reconstitution of 30S subunits using a complete set of recombinant small subunit ribosomal proteins will be used to identify l6S rRNA nucleotides and small subunit ribosomal proteins involved in 30S subunit assembly. Second, a combination of solution chemical probing and small subunit ribosomal protein-directed hydroxyl radical probing will be used to map 16S rRNA folding during different stages of 30S subunit formation. Lastly, in vitro reconstitution and standard biochemical approaches will be used to purify and characterize cellular (extra-ribosomal) factors involved in ribosome assembly. Preliminary results suggest that all three specific aims will add to our understanding of ribosome assembly. Insights gained in these studies will allow us to pursue future experiments studying ribosome biogenesis in vivo. In addition, these studies may identify targets for regulation of bacterial cell growth and perhaps lead to the production of novel antimicrobials.

描述（改编自申请人的摘要）：大约40%的 E.大肠杆菌被用来合成 核糖体成分的数量，3个RNA总计超过4000个核苷酸， 超过50种蛋白质。在细菌中，细胞生长和核糖体 生物合成是密切相关和直接相关的。因此，对于Cell 活性、核糖体成分的合成及其随后的组装 必须协调。这些观察表明，一个精确组装的 而功能性核糖体是最重要的细胞器之一， 核糖体组装的缺陷是致命的。虽然多年的研究 一直致力于研究核糖体成分和核糖体功能， 核糖体组装的途径和动力学在很大程度上仍不清楚 尽管对细胞生理学具有普遍重要性。 本研究从三个方面探讨了细胞凋亡的机制、途径和机制， 核糖体组装的动力学本提案的目的是了解 影响E.大肠杆菌30S核糖体亚基 组装件.首先，使用完整的组体外重建30S亚基， 重组小亚基核糖体蛋白将用于鉴定16S 参与30S亚基的rRNA核苷酸和小亚基核糖体蛋白 组装件.第二，溶液化学探针和小亚基的组合 核糖体蛋白导向的羟基自由基探测将用于绘制16S 30S亚基形成的不同阶段rRNA折叠。最后，体外 重构和标准生物化学方法将用于纯化和 表征参与核糖体组装的细胞（核糖体外）因子。 初步结果表明，所有这三个具体目标将增加我们的 了解核糖体组装。在这些研究中获得的见解将使 我们将继续进行研究体内核糖体生物合成的未来实验。在 此外，这些研究可能会确定细菌细胞调节的靶点， 生长，并可能导致新的抗菌剂的生产。