Molecular Mechanism of Translation

翻译分子机制

• 批准号: 7628610
• 负责人: SIMPSON JOSEPH
• 金额: $ 30.36万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7628610
• 关键词: Affect; Amino Acyl Transfer RNA; Anthrax disease; Antibiotic Resistance; Antibiotics; Anticodon; Bacteria; Biochemical; Biogenesis; Biological Assay; Bioterrorism; Catalysis; Codon Nucleotides; Complex; Drug resistance; Fluorescence; Goals; Infection; Kinetics; Life; Malignant Neoplasms; Messenger RNA; Methods; Molecular; Neomycin; Neurosciences; Organism; Paromomycin; Phase; Process; Protein Biosynthesis; RNA; Relative (related person); Ribosomes; Shapes; Site-Directed Mutagenesis; Streptomycin; Structure; Tetracyclines; Transfer RNA; Translations; Tuberculosis; X-Ray Crystallography; base; insight; research study; resistant strain

DESCRIPTION (provided by applicant): Protein synthesis is a fundamental process in all living organisms. During the elongation phase of protein synthesis, the ribosome accurately selects the aminoacyl-tRNA corresponding to the mRNA codon. The long-term objective of this proposal is to study the mechanism of tRNA selection by the ribosome. Structural studies revealed that the ribosome recognizes the shape of the codon-anticodon duplex during tRNA selection. However, the relative contributions of these contacts toward tRNA selection and ribosomal accuracy are not known. Therefore, the major goal of this proposal is to determine the contribution of specific contacts between the ribosome and the mRNA-tRNA complex in tRNA selection. We will also study how miscoding antibiotics affect the process of tRNA selection. We have developed a new, fluorescence-based, pre-steady state kinetic assay for studying tRNA selection. This powerful method will permit us to determine which contacts within the ribosome are most important for tRNA selection. In addition, this proposal will use quench-flow methods, site-directed mutagenesis of critical residues, and biochemical assays to study the mechanism of tRNA selection by the ribosome. These experiments will provide information about the dynamics of tRNA selection that cannot be easily acquired by X-ray crystallography. Ribosomes are the target for inactivation by several classes of antibiotics. Antibiotics such as paromomycin, streptomycin, tetracycline, and neomycin affect tRNA selection by the ribosome. Streptomycin is used to treat tuberculosis and tetracycline is used to treat anthrax infections. Antibiotic-resistant strains of bacteria are on the rise, causing a crisis in the management and treatment of these infections throughout the world. Understanding the mechanism of translation will provide insights for developing more effective antibiotics that target the ribosome of these drug-resistant strains of bacteria and infectious bioterrorism agents. Furthermore, this proposal will have a broad impact on fields such as RNA catalysis, origin of life, RNA structure and function, mechanism of eukaryotic protein synthesis, cancer, and neuroscience.

描述（由申请人提供）：蛋白质合成是所有生物体中的一个基本过程。 在蛋白质合成的伸长阶段，核糖体精确选择了与mRNA密码子相对应的氨基酰基-TRNA。 该提案的长期目标是研究核糖体选择tRNA选择机制。 结构研究表明，核糖体在选择tRNA期间识别密码子 - 抗原双链体的形状。 但是，这些接触对tRNA选择和核糖体精度的相对贡献尚不清楚。 因此，该提案的主要目标是确定核糖体和mRNA-tRNA复合物之间在tRNA选择中的特定接触的贡献。 我们还将研究错误编码的抗生素如何影响tRNA选择过程。 我们开发了一种基于荧光的新的，稳态的状态动力学测定法，用于研究tRNA选择。 这种强大的方法将使我们能够确定核糖体内哪些接触对于tRNA选择最重要。 此外，该提案将使用淬火流量方法，临界残基的定点诱变以及生化测定法来研究核糖体选择tRNA选择的机制。 这些实验将提供有关X射线晶体学很容易获取的tRNA选择动力学的信息。 核糖体是几种抗生素失活的靶标。 抗生素（例如抗生素，链霉素，四环素和新霉素）会影响核糖体的tRNA选择。 链霉素用于治疗结核病，四环素用于治疗炭疽感染。 抗生素的细菌菌株正在上升，在全世界对这些感染的管理和治疗中造成了危机。 了解翻译的机制将为开发更有效的抗生素提供见解，以靶向这些耐药细菌和感染性生物恐怖剂的核糖体。 此外，该建议将对诸如RNA催化，生命的起源，RNA结构和功能，真核蛋白质合成的机制，癌症和神经科学等领域产生广泛的影响。