Ribosomes translate messenger RNA (mRNA) three nucleotides at a time until translation is terminated at a stop codon. However, during all translation, frameshifting can occur, leading to the formation of proteins with amino acid sequences that differ from the in- frame product. Spontaneous frameshifting can be harmful to an organism. For instance, antibiotics such as streptomycin inhibit bacterial growth by increasing misreading and frameshifting. However, programmed translational frameshifting (which can induce high levels of frameshifting) can be used in some instances to control the ratio of specific proteins (as seen with the dnaX gene) or to increase the density of genomic information. This study explored the effects of endogenous small molecules on the IS3-frameshift-motif that is found in the transposase genes of many mobile elements. Using a cell-free protein synthesis system and a luminescent frameshift reporter, it was discovered that the addition of a small molecule extract derived from E. coli significantly decreased frameshifting, suggesting that it contains molecules that can alter translational fidelity. These experiments also revealed that the addition of the translation inhibitor chloramphenicol to translation assays at sub-inhibitory concentrations, reduced frameshift efficiency. During the studies, the role of the stability of luminescent protein reporters on the reported frameshifting levels was also explored, which is omitted in many studies. Finally, a method was developed that allows for the isolation of molecules that weakly associate with ribosomes, which opens the door for more detailed investigations of chemicals that alter translational fidelity. In conclusion, these studies provide new insight on the potential modulation of translational frameshifting by endogenous small molecules, and they set the stage to reveal the important players in this important biochemical process.
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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
Lagod, Piotr, "Effects of Endogenous E. coli Molecules on Translational Fidelity" (2021). Electronic Theses and Dissertations, 2020-. 717.