Combinatorial analysis of loop nucleotides in human mitochondrial tRNA Leu(UUR)

Ekaterina I. Zagryadskaya, Shana O. Kelley*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

A series of disease-related mutations are known to affect the hs mt tRNALeu(UUR) gene, and the molecular-level properties of this tRNA may underlie the effects of pathogenic sequence changes. A combinatorial approach has been used to explore the importance of the D, TΨC, and anticodon loops of hs mt tRNALeu(UUR) in the structure and function of this molecule. A tRNA library was constructed with 20 randomized nucleotides in the loop regions of hs mt tRNALeu(UUR), and tRNA variants that were aminoacylated by hs mt LeuRS were isolated using an in vitro selection approach. Analysis of 26 selected sequences revealed that a stabilized anticodon stem significantly enhances aminoacylation activity. However, anticodon loop nucleotides were not conserved in the active sequences, indicating that this region of hs mt tRNALeu(UUR) is not involved in recognition by LeuRS. Within the D and TΨC loops, only two nucleotides conserved their identities, while new sequences were selected that likely mediate interloop interactions. The results indicate that hs mt tRNALeu(UUR), which is known to have structurally weak D and anticodon stems, benefits functionally from the introduction of stabilizing interactions. However, the locations of individual nucleotides that govern discrimination of this tRNA by hs mt LeuRS still remain obscure.

Original languageEnglish (US)
Pages (from-to)233-242
Number of pages10
JournalBiochemistry
Volume44
Issue number1
DOIs
StatePublished - Jan 11 2005
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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