Item Details

title

Investigation into the importnace of a tRNA modifying enzyme and its role in decoding

author

Nilsson, Emil Mikael

abstract

Life as we know it exists because of intrinsic ability to faithfully translate genetic material into functional proteins. Cells have evolved intricate and energetically costly strategies to minimize mistakes during cellular information transfer. One such strategy is the incorporation of post-transcriptional modification on the transfer RNA (tRNA). tRNA modifications regulate many aspects essential to protein biosynthesis including elongation rates, aminoacyl-tRNA synthetase interactions, and codon preference. Some modifications are found in all domains of life while others are unique to one domain; some modifications occurs invariably across tRNAs while others are unique to a single tRNA isoascceptor. Lysidine (L/k2C) is a bacterial-exclusive modification that occurs on a single tRNA in the bacterial cell, the minor isoleucine tRNA acceptor (tRNAIle2) destined to decode the isoleucine codon AUA. Despite the very isolated occurrence of lysidine, the gene encoding the enzyme responsible for its formation, tRNA Ile lysidine synthetase (TilS), has been shown to be essential in Escherichia coli. TilS conservation, 93% of bacteria, suggests that the essentiality of TilS should expand beyond E. coli. However, here we report the natural evolution of single nucleotide polymorphisms (SNP) targeting the tilS gene in Burkholderia cenocepacia (HI2424). We show that each of the SNP is associated with a loss-of-function phenotype in the resulting protein, demonstrated in vitro and in vivo. Despite this the isogenic mutants show an increased ability to propagate under nutrient starvation conditions. We provide results explaining why tilS might be essential in E. coli but not B. cenocepacia, and show that the tilS SNPs have a unique functional effect in B. cenocepacia TilS.

subject

Isoleucine

Lysidine

TilS

Translational fidelity

tRNA

tRNAIle lysidine synthetase

contributor

Alexander, Rebecca W (committee chair)

Lyles, Douglas S (committee member)

Comstock-Ferguson, Lindsay R (committee member)

Donati, George L (committee member)

Stich, Troy A (committee member)

date

2019-05-24T08:35:43Z (accessioned)

2021-05-23T08:30:11Z (available)

2019 (issued)

degree

Chemistry (discipline)

embargo

2021-05-23 (terms)

identifier

http://hdl.handle.net/10339/93950 (uri)

language

en (iso)

publisher

Wake Forest University

type

Dissertation