A new assay for tRNA aminoacylation kinetics

Alexey D. Wolfson, Jeffrey A. Pleiss, Olke C. Uhlenbeck*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

42 Scopus citations

Abstract

An improved quantitative assay for tRNA aminoacylation is presented based on charging of a nicked tRNA followed by separation of an aminoacylated 3'-fragment on an acidic denaturing polyacrylamide gel. Kinetic parameters of tRNA aminoacylation by Escherichia coli AlaRS obtained by the new method are in excellent agreement with those measured by the conventional method. This assay provides several advantages over the traditional methods of measuring tRNA aminoacylation: (1) the fraction of aminoacyl-tRNA is measured directly; (2) data can be obtained at saturating amino acid concentrations; and (3) the assay is significantly more sensitive.

Original languageEnglish (US)
Pages (from-to)1019-1023
Number of pages5
JournalRNA
Volume4
Issue number8
DOIs
StatePublished - Aug 1998

Keywords

  • Acid gel electrophoresis
  • Alanine
  • Aminoacyl-tRNA synthetase
  • tRNA, kinetics

ASJC Scopus subject areas

  • Molecular Biology

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