Substitution of the Anticodon Loop of Yeast Tyrosine Transfer Ribonucleic Acid (Rna)

Procedures for replacing residues 33-35 in the anticodon loop of yeast tRNA('Tyr) with any desired oligonucleotide have been developed. Half-molecules of tRNA('Tyr) from partial digestions with ribonuclease A or T(,1) were paired to create a gap in the anticodon loop. An oligonucleotide was inserted into the gap using RNA ligase, polynucleotide kinase, and Pset 1 polynucleotide kinase. The rate of aminoacylation of anticodon loop substituted tRNA('TYR)s by yeast tyrosyl-tRNA synthetase was found to depend upon the sequence of the oligonucleotide inserted. This suggests that the nucleotides in the anticodon loop of yeast tRNA('Tyr) are required for optimal aminoacylation. In addition, tRNA('Tyr) modified to have a phenylalanine anticodon was shown to be misacylated by yeast phenylalanyl-tRNA synthetase at a rate at least 10 times faster than unmodified tRNA('Tyr). Substitution of the anticodon of yeast tRNA('Phe) with a tyrosine anticodon was shown to stimulate the level of misacylation by tyrosyl-tRNA synthetase. Thus, the anticodon is used by phenylalanyl-tRNA synthetase and by the tyrosyl-tRNA synthetase to distinguish between tRNAs.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biochemistry

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2014

Author dc:creator

• Bare, Lance Alan

• Biology, General

Identifier

(UMI)AAI8600122

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/70546