The molecular mechanism of post-peptidyl quality control on the ribosome

核糖体肽基后质量控制的分子机制

基本信息

批准号：
8725512
负责人：
Hani Zaher
金额：
$ 24.49万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8725512
关键词：
Affect Amino Acids Amino Acyl Transfer RNA Amino Acyl-tRNA Synthetases Antibiotics Biochemical Biological Assay Biology Catalytic RNA Chemicals Code Codon Nucleotides Complex Cues DNA-Directed DNA Polymerase DNA-Directed RNA Polymerase Defect Ensure Escherichia coli Eukaryota Exhibits Genomics Goals Growth Investigation Kinetics Knock-out Life Light Mass Spectrum Analysis Messenger RNA Molecular Monitor Mutate Mutation Pathway interactions Peptides Phenotype Process Protein Biosynthesis Proteins Quality Control Reaction Recombinant Proteins Reporter Reporter Genes Research Resolution Ribosomes Role Sense Codon Signal Transduction Site Specificity Structure System Techniques Technology Terminator Codon Therapeutic Therapeutic Agents Thermodynamics Time Transfer RNA Translations Trees Two-Dimensional Gel Electrophoresis Variant Work Yeasts analytical tool base interest mutant polypeptide premature programs protein aminoacid sequence release factor release factor 3 research study ribosome releasing factor stopped-flow fluorescence

项目摘要

Project Summary: Organismal growth and viability is dependent on the faithful and fast decoding of its genomic information into functional peptide sequences. High-accuracy protein synthesis ensures that errant polypeptides, which are more prone to misfold and hence may have undesirable toxic consequences, are not produced. The overall fidelity of protein synthesis appears to be limited by the action of the ribosome, which is the two-subunit macromolecular machine responsible for the decoding of the messenger RNA into protein in all domains of life. During each cycle of elongation, the ribosome carefully selects the appropriate aminoacyl-tRNA (aa-tRNA) that matches the codon in the decoding center from a large-pool of competing aa-tRNAs. In addition to this, we have recently uncovered a quality control mechanism on the ribosome that takes place after peptide-bond formation, which contributes to high-fidelity protein synthesis. Akin to the proofreading strategies enjoyed by DNA and RNA polymerases and tRNA synthetases, the newly discovered ribosome-based mechanism is in place to monitor the quality of the just completed chemical step. During the elongation cycle, the incorporation of an incorrect amino acid was found to have dramatic effects on the specificity of the subsequent reaction. This iterated accumulation of errors results in the abortive termination of protein synthesis by release factors, which under normal conditions rarely decode sense codons. The long term goal of our work is to gain a thorough understanding of the molecular mechanisms underlying this process. Our immediate goal is to find out how the signal is communicated from a perturbed mRNA-tRNA interaction to the decoding center, which ultimately leads to low-fidelity protein synthesis. We are also interested in how the activity of release factors is modulated on sense codons in the presence of a perturbed mRNA- tRNA interaction, and the structural cues that are responsible for this activity. These goals are built around pre-steady state kinetics approaches in the context of mutated translation components, and low-resolution structural probing techniques. As a third goal we are interested in exploring a previously unknown role for release factor 3 in the quality control mechanism and its utility in cellular viability. Finally we are interested in finding whether this system exists in eukaryotes, and identifying other factors, if any, that might be involved during this process.

项目摘要：有机生长和生存能力取决于其基因组的忠实和快速解码信息到功能性肽序列中。高准确蛋白合成可确保错误多肽更容易折叠，因此可能会带来不良的毒性后果，不是生产。蛋白质合成的总体保真度似乎受到了限制核糖体，这是负责解码的两支大分子机器信使RNA成为生命所有领域的蛋白质。在每个伸长周期中，核糖体仔细选择与解码中的密码子匹配的适当氨基酰基tRNA（AA-tRNA）来自大型竞争的AA-Trnas的中心。除此之外，我们最近发现了核糖体的质量控制机制发生在肽键形成之后的核糖体上，有助于高保真蛋白的合成。类似于DNA和DNA享受的校对策略 RNA聚合酶和tRNA合成酶，新发现的基于核糖体的机制已经存在监视刚刚完成的化学步骤的质量。在伸长周期中，掺入发现不正确的氨基酸对随后的特异性具有巨大影响反应。这种迭代的错误积累导致蛋白质合成的流产释放因子在正常条件下很少解码感官密码子。我们的长期目标工作是要彻底了解该过程的分子机制。我们的直接目标是找出如何从扰动的mRNA-tRNA相互作用中传达信号到达解码中心，最终导致低保真蛋白的合成。我们也对在存在扰动的mRNA-的情况下，如何调节释放因子的活性。 tRNA相互作用以及负责此活动的结构提示。这些目标是建立的在突变的翻译组件中，围绕稳态的状态动力学方法，以及低分辨率结构探测技术。作为第三个目标，我们有兴趣探索以前的释放因子3在质量控制机制中的不明作用及其在细胞活力中的效用。最后，我们有兴趣发现该系统是否存在于真核生物中，并确定其他在此过程中可能涉及的因素（如果有）。