Translational frameshifting is a process that leads to the production of multiple different proteins from a single mRNA. Programmed translational frameshifting is typically utilized by retroviruses to increase genome coding capacity and to control the ratio between proteins necessary for different infection stages. In E. coli, translational frameshifting regulates the production of several essential proteins, including the replication/clamp loader subunits tau (τ) and gamma(γ), which are products of the gene dnaX. In E. coli, τ is an essential auxiliary factor of DNA polymerase III holoenzyme. τ can dimerize Pol III and interact with the DnaB helicase, which allows synthesis of leading and lagging DNA strands. The role of γ is unknown since it does not share similar properties with τ apart for clamp loading; but, γ is implicated in DNA repair and likely has key role in polymerase switching. In this study, the effects of ribosomal availability in the cell on the dnaX frameshift frequencies (regulation of τ/γ) were investigated. It was found that lower ribosomal availability in the cell resulted in lower frameshift frequencies on reporters containing the dnaX frameshift motif. Also, E. coli strains with tagged genomic dnaX and disrupted ompT were generated. Although, it is generally accepted that τ/γ are produced at equimolar concentrations, it was discovered that endogenous γ was produced at substantially higher concentrations than τ when cells were grown in defined media (EZ MOPS). This finding shows that that the ratio of τ/γ can be regulated. These findings suggest there are yet uncharacterized roles of γ, such as polymerase switching, that may be the reason for its production in many bacteria.
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Master of Science (M.S.)
College of Medicine
Burnett School of Biomedical Sciences
Length of Campus-only Access
Masters Thesis (Campus-only Access)
College of Medicine
Kargazhanov, Akhmetzada, "Investigating Effects of Ribosomal Occupancy on the Production of the Clamp Loader Subunits Tau and Gamma" (2021). Electronic Theses and Dissertations, 2020-. 706.