Quality Control of Protein Translation

蛋白质翻译的质量控制

基本信息

批准号：
8113893
负责人：
A. WALI KARZAI
金额：
$ 30.47万
依托单位：
STATE UNIVERSITY NEW YORK STONY BROOK
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-01 至 2013-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8113893
关键词：
Affinity Alanine-Specific tRNA Alanine-tRNA Ligase Amino Acids Anti-Bacterial Agents Anti-Infective Agents Antibiotics Anticodon Binding Biochemical Bioinformatics Biological Process C-terminal Cell physiology Charge Codon Nucleotides Complex DNA Data Detection Development Distal EEF1A1 gene Effectiveness Elements Ensure Escherichia coli Event Exoribonucleases Family Gene Expression Genetic Goals Guanosine Triphosphate Guanosine Triphosphate Phosphohydrolases Health Hydrolysis Kinetics Laboratories Life Light Link Measures Mediating Messenger RNA Molecular Molecular Genetics Organism Pathway interactions Peptides Phosphodiesterase I Physiological Play Positioning Attribute Process Prokaryotic Cells Property Protein Binding Protein Biochemistry Protein Fragment Proteins Proteolysis Quality Control RNA RNA-Protein Interaction Reading Frames Relative (related person)Research Ribosomes Role Site SmpB protein Staging Structure Surface System Tail Transcript Transfer RNA Translation Process Translations Variant Virulence base design fascinate insight knowledge base mRNA Decay mRNA Stability member microorganism novel pathogenic bacteria programs protein degradation quality assurance ribonuclease R stem tmRNA

项目摘要

DESCRIPTION (provided by applicant): The accurate flow of genetic information from DNA to RNA to protein is essential for all living organisms. An astonishing array of quality-assurance mechanisms have evolved to ensure that high degree of fidelity is maintained at every stage of this process. One of the most fascinating quality control mechanisms involves tmRNA, also known as SsrA or 10Sa RNA. tmRNA is a versatile and highly conserved bacterial molecule endowed with the combined structural and functional properties of both a tRNA and an mRNA. Our previous studies have shown that all known activities of tmRNA require SmpB, a small protein that binds tmRNA specifically and with high affinity to promote its association with stalled ribosomes. The SmpB-tmRNA system orchestrates three key biological functions: 1) recognition and rescue of ribosomes stalled on aberrant mRNAs, 2) disposal of the causative defective mRNAs, and 3) addition of a degradation tag to the incomplete protein fragments for directed proteolysis. Although not essential in E. coli, tmRNA activity is essential for bacterial survival under adverse conditions and for virulence in some, and perhaps all, pathogenic bacteria. Recent evidence from our laboratory suggests that in addition to its quality control function the tmRNA system might also play a key regulatory role in certain physiological pathways. Moreover, because the SmpB and tmRNA are found only in prokaryotes, involves novel RNA and protein factors, and is essential for the survival of pathogenic bacteria, a deeper mechanistic understanding of this system might allow the design of highly specific new anti-bacterial agents. The molecular basis for the formation of the SmpB-tmRNA complex and the subsequent recognition of stalled ribosomes are not well understood. The objective of this research program is to use a combination of molecular genetics, protein biochemistry, bioinformatics, and structural approaches to elucidate the mechanism of action of the SmpB-tmRNA quality control system. The emphasis is on the molecular characterization of how SmpB-tmRNA complex recognizes stalled ribosomes and promotes the detection and selective decay of the causative defective mRNA by the 3'-5' exonuclease RNase R. Specifically, through these studies we wish to understand the biochemical and structural basis for the interactions of SmpB and RNase R with tmRNA and the ribosome; i.e. what amino acid residues are involved, what base specific contacts are made, what structural features contribute to the formation of the tmRNA-associated SmpB and RNase R complexes and their interaction with stalled ribosome. PUBLIC HEALTH RELEVANCE: As currently available antibiotics lose their effectiveness the need for new counter measure becomes ever more urgent. The genetic, biochemical, and structural studies outlined here offer the opportunity to gain novel insights into and a deeper mechanistic understanding of a unique bacterial surveillance system mediated by the versatile tmRNA and its essential protein partner, SmpB. A thorough understanding of this extraordinary bacterial system, essential for survival and virulence of many pathogenic bacteria, should pave the way for development of knowledge-based new anti-infective agents that exclusively target pathogenic microorganisms. Ultimately, these insights will have implications for a better understanding of a variety of cellular processes, including control of gene expression, synthesis and degradation of proteins, and the targeted decay defective mRNAs.

描述（由申请人提供）：遗传信息从 DNA 到 RNA 再到蛋白质的准确流动对于所有生物体至关重要。一系列令人惊叹的质量保证机制已经发展起来，以确保在此过程的每个阶段都保持高度的保真度。最令人着迷的质量控制机制之一涉及 tmRNA，也称为 SsrA 或 10Sa RNA。 tmRNA 是一种多功能且高度保守的细菌分子，具有 tRNA 和 mRNA 的结构和功能特性。我们之前的研究表明，所有已知的 tmRNA 活性都需要 SmpB，这是一种小蛋白，能够特异性地结合 tmRNA，并具有高亲和力，以促进其与停滞核糖体的结合。 SmpB-tmRNA 系统协调三个关键的生物学功能：1) 识别和拯救在异常 mRNA 上停滞的核糖体，2) 处理导致缺陷的 mRNA，3) 在不完整的蛋白质片段中添加降解标签以进行定向蛋白水解。尽管 tmRNA 活性在大肠杆菌中不是必需的，但它对于细菌在不利条件下的生存以及某些（也许是所有）病原菌的毒力至关重要。我们实验室的最新证据表明，除了质量控制功能外，tmRNA 系统还可能在某些生理途径中发挥关键的调节作用。此外，由于SmpB和tmRNA仅存在于原核生物中，涉及新型RNA和蛋白质因子，并且对于病原菌的生存至关重要，因此对该系统更深入的机制了解可能有助于设计高度特异性的新型抗菌剂。 SmpB-tmRNA 复合物形成的分子基础以及随后对停滞核糖体的识别尚不清楚。该研究计划的目标是结合分子遗传学、蛋白质生物化学、生物信息学和结构方法来阐明 SmpB-tmRNA 质量控制系统的作用机制。重点是 SmpB-tmRNA 复合物如何识别停滞核糖体并促进 3'-5' 核酸外切酶 RNase R 检测和选择性降解致病缺陷 mRNA 的分子特征。具体来说，通过这些研究，我们希望了解SmpB 和 RNase R 与 tmRNA 和核糖体相互作用的结构基础；即涉及哪些氨基酸残基、进行哪些碱基特异性接触、哪些结构特征有助于 tmRNA 相关的 SmpB 和 RNase R 复合物的形成及其与停滞核糖体的相互作用。公共健康相关性：随着当前可用的抗生素失去效力，对新对策的需求变得更加迫切。本文概述的遗传、生化和结构研究提供了机会，让我们有机会获得对由多功能 tmRNA 及其重要蛋白质伙伴 SmpB 介导的独特细菌监测系统的新见解和更深入的机制理解。对这种非凡的细菌系统的彻底了解对于许多病原菌的生存和毒力至关重要，应该为开发专门针对病原微生物的基于知识的新型抗感染药物铺平道路。最终，这些见解将对更好地理解各种细胞过程产生影响，包括基因表达的控制、蛋白质的合成和降解，以及目标衰变缺陷 mRNA。