The role played by the RNAs and tRNAs in translation

描述(申请人提供)：核糖体是两个亚基的大分子机器，负责将信使RNA解码成编码的多肽--遗传密码的翻译。在履行这一重要的细胞功能时，核糖体执行一系列离散的功能，包括选择合适的氨基酰基tRNAs，催化多肽键的形成，准确地转移mRNA：tRNA复合体，以及释放完整的多肽链。对生命起源的理论考量预测了rRNA在翻译中的核心作用。这一观点得到了RNA的催化、rRNA序列之间的极端保守以及核糖体的原子分辨结构的证实，这些结构主要显示在功能关键区的rRNA。我们工作的长期目标是了解核糖体功能的分子力学。我们特别感兴趣的是了解翻译机制的rRNA和tRNA组件如何对蛋白质合成的整个过程做出贡献。在过去的十年里，来自CryoEM的原子分辨率结构信息和低分辨率信息的出现极大地改变了我们的研究重点。虽然之前的挑战是确定位于功能关键区的核糖体元件，但现在的目标是了解这些从结构上确定的关键组件如何发挥作用。核糖体为解剖分子运动和研究它们如何定义生物学上的基本信号转导通路提供了一个极好的、易处理的系统。所有三个特定的目标都建立在稳定前的动力学方法和分析突变的翻译成分，以在分子水平上剖析核糖体功能。我们特别感兴趣的是核糖体如何在正义密码子上选择同源氨酰-tRNA和在终止密码子上如何选择合适的释放因子，以及这些机制是如何相关的。这项提议的一个主要新焦点是探索在多肽键形成后发生在核糖体上的一种新的质量控制系统的机制特征。 与公共卫生相关：核糖体是许多天然和合成抗生素的靶标，这些抗生素影响翻译周期中的不同步骤。从这里提出的详细分子研究中获得的信息将有助于确定当前抗生素的特征，使其有效，并将有助于设计更有效的治疗方法。

DESCRIPTION (provided by applicant): The ribosome is the two-subunit macromolecular machine responsible for the decoding of the messenger RNA into the encoded polypeptide - the translation of the genetic code. In fulfilling this essential cellular role, ribosomes carry out a number of discrete functions including selecting the appropriate aminoacyl tRNAs, catalyzing peptide bond formation (PT), accurately translocating the mRNA:tRNA complex and releasing the completed polypeptide chain. Theoretical considerations of the origin of life predict a central role for the rRNAs in translation. This view has been strengthened by the demonstration of catalysis by RNA, extreme conservation observed among rRNA sequences and finally by atomic resolution structures of the ribosome displaying primarily rRNA in the functionally critical regions. The long-term goal of our work is to understand the molecular mechanics of ribosome function. We are particularly interested in understanding how the rRNA and tRNA components of the translation machinery contribute to the overall process of protein synthesis. The advent of atomic resolution structural information and lower resolution information from cryoEM has substantially changed the focus of our research during the past ten years. While the previous challenge was to identify elements of the ribosome located in functionally critical regions, the goal now is to understand how these structurally identified critical components function. The ribosome provides an excellent, tractable system for dissecting molecular movements and examining how they define biologically fundamental signal transduction pathways. All three specific aims are built around pre-steady state kinetic approaches and the analysis of mutated translation components to dissect ribosome function at the molecular level. We are particularly interested in the molecular details of how the ribosome selects the cognate aminoacyl-tRNA on sense codons and the appropriate release factor on stop codons, and how these mechanisms are related. A major new focus of this proposal is the exploration of the mechanistic features of a novel quality control system on the ribosome that takes place following peptide bond formation. PUBLIC HEALTH RELEVANCE: The ribosome is the target of many natural and synthetic antibiotics that affect distinct steps in the translation cycle. Information obtained from detailed molecular studies proposed here will help in defining the features of current antibiotics which make them effective and will aid in the design of more effective therapeutics.