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Characterization of a temperature-sensitive mutation that impairs the function of yeast tRNA nucleotidyltransferase

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Characterization of a temperature-sensitive mutation that impairs the function of yeast tRNA nucleotidyltransferase

Shan, Xunying (2005) Characterization of a temperature-sensitive mutation that impairs the function of yeast tRNA nucleotidyltransferase. Masters thesis, Concordia University.

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Abstract

Mature tRNAs require a 3'-terminal cytidine-cytidine-adenosine sequence to function in protein synthesis. The addition of these nucleotides into tRNAs with incomplete 3'-termini is catalyzed by the enzyme tRNA nucleotidyltransferase. Genetic studies have resulted in the isolation of the gene coding for tRNA nucleotidyltransferase in yeast and in vivo studies identified a temperature-sensitive yeast strain with a mutant CCA1 gene, which can survive at room temperature, but not at 37{493}C. Gene sequencing revealed that the temperature-sensitive phenotype came from a single nucleotide substitution in the CCA1 gene, which led to the substitution of a glutamate residue by a lysine in the protein (D. Kushner, personal communication). In this work, enzymatic characterization showed that this lysine variant had approximately 5% of the activity of the wild-type enzyme and a tertiary structural change as detected by fluorescence spectroscopy at the permissive temperature. Thermal stability monitored by circular dichroism showed that the lysine variant is less thermally stable than the wild-type enzyme with an approximately 6{493}C lower transitional temperature.

Divisions:Concordia University > Faculty of Arts and Science > Chemistry and Biochemistry
Item Type:Thesis (Masters)
Authors:Shan, Xunying
Pagination:xiii, 125 leaves : ill ; 29 cm.
Institution:Concordia University
Degree Name:M. Sc.
Program:Chemistry and Biochemistry
Date:2005
Thesis Supervisor(s):Joyce, Paul
ID Code:8512
Deposited By:Concordia University Libraries
Deposited On:18 Aug 2011 14:27
Last Modified:18 Aug 2011 14:27
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