Date of Award

10-29-2022

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Department of Chemistry

First Advisor

Christopher S Weitzel

Abstract

Aminoacyl-tRNA synthetases (aaRSs) are essential proteins. Their canonical function is to catalyze the attachment of amino acids to cognate tRNA. The organism Sulfolobus islandicus contains two aaRSs for leucine. These leucyl-tRNA synthetase (LeuRS) paralogs were previously studied using a single tRNALeu isoacceptor, tRNALeu-UAG. One leucyl-tRNA synthetase, LeuRS-F, was shown to faithfully perform the necessary function of charging, the process of catalytically attaching leucine to tRNALeu. Furthermore, the essentiality of LeuRS-F was emphasized by an inability to remove its gene from S. islandicus. The paralog, LeuRS-I, was shown to have no aminoacylation activity, compared to LeuRS-F, even though it could activate amino acids and bind tRNA. Despite this, it is required for the organism’s optimal growth. This project focused on vetting LeuRS-F’s and LeuRS-I’s ability to charge tRNALeu using isoacceptors tRNALeu-CAA and tRNALeu-CAG via aminoacylation assays that utilize tritiated leucine. The results further support that LeuRS-F performs the canonical function of an aaRS whereas LeuRS-I does not. Hypothesizing that LeuRS-I might interact with LeuRS-F, mixing assays containing both LeuRS-F and LeuRS-I were performed using ribonucleic acid samples isolated from S. islandicus, representative of the organisms total tRNA repertoire. The results showed an increase in total charging possibly indicating the first tRNA charging activity observed in LeuRS-I.

Comments

Imported from Brewster_ilstu_0092N_12274.pdf

DOI

https://doi.org/10.30707/ETD2023.20230711063200745168.999995

Page Count

134

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