An aminoacyl-tRNA synthetase with a defunct editing site

Stanley W. Lue, Shana O. Kelley*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

64 Scopus citations

Abstract

Many aminoacyl-tRNA synthetases (aaRSs) contain two active sites, a synthetic site catalyzing aminoacyl-adenylate formation and tRNA aminoacylation and a second editing or proofreading site that hydrolyzes misactivated adenylates or mischarged tRNAs. The combined activities of these two sites lead to rigorous accuracy in tRNA aminoacylation, and both activities are essential to LeuRS and other aaRSs. Here, we describe studies of the human mitochondrial (hs mt) LeuRS indicating that the two active sites of this enzyme have undergone functional changes that impact how accurate aminoacylation is achieved. The sequence of the hs mt LeuRS closely resembles a bacterial LeuRS overall but displays significant variability in regions of the editing site. Studies comparing Escherichia coli and hs mt LeuRS reveal that the proofreading activity of the mt enzyme is disrupted by these sequence changes, as significant levels of Ile-tRNALeu are formed in the presence of high concentrations of the noncognate amino acid. Experiments monitoring deacylation of Ile-tRNA Leu and misactivated adenylate turnover revealed that the editing active site is not operational. However, hs mt LeuRS has weaker binding affinities for both cognate and noncognate amino acids relative to the E. coli enzyme and an elevated discrimination ratio. Therefore, the enzyme achieves fidelity using a more specific synthetic active site that is not prone to errors under physiological conditions. This enhanced specificity must compensate for the presence of a defunct editing site and ensures translational accuracy.

Original languageEnglish (US)
Pages (from-to)3010-3016
Number of pages7
JournalBiochemistry
Volume44
Issue number8
DOIs
StatePublished - Mar 1 2005
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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