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The structural dynamics of ribosomal frameshifting and ribosome rescue

核糖体移码和核糖体拯救的结构动力学

基本信息

批准号：
10377976
负责人：
Ruben L Gonzalez
金额：
$ 39.46万
依托单位：
COLUMBIA UNIV NEW YORK MORNINGSIDE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-05-01 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10377976
关键词：
Address Amino Acids Antibiotics Bacteria Bacterial Model Binding Biochemical Biological Clinical Codon Nucleotides Collaborations Complement Cryoelectron Microscopy Data Development Disease Electron Microscopy Eukaryota Event Fluorescence Frameshift Mutation Gene Expression Genome Goals Health Human In Vitro Investigation Kinetics Label Laboratories Letters Life Link Malignant Neoplasms Mediating Messenger RNA Modeling Molecular Conformation Molecular Target Motivation Nucleotides Organism Play Positioning Attribute Process Production Property Protein Biosynthesis Proteins Reporting Resolution Rest Ribosomal Frameshifting Ribosomal Proteins Ribosomes Role Saccharomyces cerevisiae Series Signal Transduction Specific qualifier value Structure System Techniques Technology Testing Time Transfer RNA Translations Viral Cancer Virus Diseases Work Yeasts base cryogenics experimental study flexibility fluorescence imaging fluorophore human disease next generation polypeptide public health relevance reconstitution single molecule single-molecule FRET small molecule small molecule therapeutics

项目摘要

PROJECT SUMMARY Translation of messenger RNAs (mRNAs) into proteins by the ribosome and the rest of the translation machinery (TM) is a fundamental step in gene expression that is central to life. Because the bacterial TM is a proven target for the development of new antibiotics and because many human diseases have been causally linked to dysregulation of translation, the mechanisms of translation and translational control in bacteria and eukaryotes remain under intense investigation. Over the past two decades, structural studies have revealed the large-scale structural rearrangements the TM undergoes during protein synthesis. Unfortunately, the size, complexity, and conformational flexibility of the TM have greatly impeded studies of these dynamics, significantly limiting our understanding of how these dynamics contribute to the mechanisms of translation and translational control. Nonetheless, using a combination of single-moleucle fluorescence and structural techniques, we and others have been able to characterize the dynamics of the core steps of translation by the bacterial TM. Despite these accomplishments, critical gaps in our understanding remain regarding whether and how the dynamics of these core steps are modulated as part of biomedically important translational control strategies. To fill these gaps, in the first aim of this application, we propose to use a combination of single- molecule fluorescence, structural, and biochemical approaches to investigate how the dynamics of the bacterial TM are modulated in order to drive ribosomal frameshifting. Frameshifting is a translational control strategy in which the TM slips backward or forward by one or more nucleotides on the mRNA to either correct an insertion or deletion ‘frameshift’ mutation that would otherwise result in production of an aberrant or truncated protein or to drive the synthesis of more than one protein product from a single mRNA. These experiments promise to reveal the still-elusive mechanism(s) that underlie frameshifting. In the second aim, we will use analogous approaches to investigate how ribosome rescue factors modulate the dynamics of the bacterial TM as part of the mechanisms through which they recognize and rescue ribosomes that have become translationally compromised. These studies will provide structure-based mechanistic models of bacterial ribosome rescue systems that can be exploited in the development of new antibiotics. In the third aim, we will extend our combination of single-molecule fluorescence and structural techniques to a yeast translation system, enabling us to investigate eukaryotic-specific aspects of the core steps of translation, frameshifting, and ribosome rescue. The results of these studies will reveal the mechanisms that drive and regulate translation in eukaryotes, providing a framework for investigating the role of translational control in human health and disease.

项目总结