The Unexplored Mechanisms And Regulatory Functions Of Ribosomal Translocation

Other Titles
Abstract

In every cell, protein synthesis is carried out by the the ribosome, a complex macromolecular RNA-protein assembly,. Decades of structural and kinetic studies have increased our understanding of ribosome initiation, decoding, translocation and termination. Yet, the underlying mechanism of these fundamental processes has yet to be fully delineated. Hence, the molecular basis of regulation remains obscure. Here, single-molecule fluorescence methods are applied to decipher the mechanism and regulatory roles of the multi-step process of directional substrate translocation on the ribosome that accompanies every round of protein synthesis. In Chapter 1, single-molecule fluorescence resonance energy transfer (smFRET) is introduced as a tool for studying bacterial ribosome translocation. Chapter 2 details the experimental methods. In Chapter 3, the elongation factor G(EF-G)-catalyzed movement of substrates through the ribosome is examined from several perspectives or signals reporting on various degrees of freedom of ribosome dynamics. Two ribosomal states interconvert in the presence of EF-G(GDP), displaying novel head domain motions, until relocking takes place. In Chapter 4, in order to test if the mentioned fluctuations leading to relocking are correlated to the engagement of the P-site by the peptidyl-tRNA, the translocation of miscoded tRNAs is studied. Severe defects in the relocking stages of translocation reveal the correlation between this new stage of translocation and P-site tRNA engagement.

Journal / Series
Volume & Issue
Description
Sponsorship
Date Issued
2017
Publisher
Keywords
Aminoglycosides; Antibiotic resistance; EF-P; Miscoding; Protein synthesis; Ribosome translocation
Location
Effective Date
Expiration Date
Sector
Employer
Union
Union Local
NAICS
Number of Workers
Committee Chair
Committee Co-Chair
Committee Member
Degree Discipline
Physiology, Biophysics & Systems Biology
Degree Name
Degree Level
Doctor of Philosophy
Related Version
Related DOI
Related To
Related Part
Based on Related Item
Has Other Format(s)
Part of Related Item
Related To
Related Publication(s)
Link(s) to Related Publication(s)
References
Link(s) to Reference(s)
Previously Published As
Government Document
ISBN
ISMN
ISSN
Other Identifiers
Rights
Attribution-NonCommercial-NoDerivatives 4.0 International
Types
dissertation or thesis
Accessibility Feature
Accessibility Hazard
Accessibility Summary
Link(s) to Catalog Record