Modified uridines with C5-methylene substituents at the first position of the tRNA anticodon stabilize U·G wobble pairing during decoding

Shinya Kurata, Albert Weixlbaumer, Takashi Ohtsuki, Tomomi Shimazaki, Takeshi Wada, Yohei Kirino, Kazuyuki Takai, Kimitsuna Watanabe, V. Ramakrishnan, Tsutomu Suzuki

Research output: Contribution to journalArticle

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Abstract

Post-transcriptional modifications at the first (wobble) position of the tRNA anticodon participate in precise decoding of the genetic code. To decode codons that end in a purine (R) (i.e. NNR), tRNAs frequently utilize 5-methyluridine derivatives (xm5U) at the wobble position. However, the functional properties of the C5-substituents of xm5U in codon recognition remain elusive. We previously found that mitochondrial tRNAs Leu(UUR) with pathogenic point mutations isolated from MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes) patients lacked the 5-taurinomethyluridine (τm5U) modification and caused a decoding defect. Here, we constructed Escherichia coli tRNAsLeu(UUR) with or without xm5U modifications at the wobble position and measured their decoding activities in an in vitro translation as well as by A-site tRNA binding. In addition, the decoding properties of tRNAArg lacking mnm5U modification in a knock-out strain of the modifying enzyme (ΔmnmE) were examined by pulse labeling using reporter constructs with consecutive AGR codons. Our results demonstrate that the xm5U modification plays a critical role in decoding NNG codons by stabilizing U·G pairing at the wobble position. Crystal structures of an anticodon stem-loop containing τm5U interacting with a UUA or UUG codon at the ribosomal A-site revealed that the τm5U·G base pair does not have classical U·G wobble geometry. These structures provide help to explain how the τm5U modification enables efficient decoding of UUG codons.

Original languageEnglish
Pages (from-to)18801-18811
Number of pages11
JournalJournal of Biological Chemistry
Volume283
Issue number27
DOIs
Publication statusPublished - Jul 4 2008

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Anticodon
Uridine
Transfer RNA
Codon
Decoding
RNA, Transfer, Arg
RNA, Transfer, Leu
MELAS Syndrome
Genetic Code
Point Mutation
Base Pairing
Labeling
Escherichia coli
Crystal structure
Binding Sites
Derivatives
Defects
Geometry
Enzymes

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Modified uridines with C5-methylene substituents at the first position of the tRNA anticodon stabilize U·G wobble pairing during decoding. / Kurata, Shinya; Weixlbaumer, Albert; Ohtsuki, Takashi; Shimazaki, Tomomi; Wada, Takeshi; Kirino, Yohei; Takai, Kazuyuki; Watanabe, Kimitsuna; Ramakrishnan, V.; Suzuki, Tsutomu.

In: Journal of Biological Chemistry, Vol. 283, No. 27, 04.07.2008, p. 18801-18811.

Research output: Contribution to journalArticle

Kurata, S, Weixlbaumer, A, Ohtsuki, T, Shimazaki, T, Wada, T, Kirino, Y, Takai, K, Watanabe, K, Ramakrishnan, V & Suzuki, T 2008, 'Modified uridines with C5-methylene substituents at the first position of the tRNA anticodon stabilize U·G wobble pairing during decoding', Journal of Biological Chemistry, vol. 283, no. 27, pp. 18801-18811. https://doi.org/10.1074/jbc.M800233200
Kurata, Shinya ; Weixlbaumer, Albert ; Ohtsuki, Takashi ; Shimazaki, Tomomi ; Wada, Takeshi ; Kirino, Yohei ; Takai, Kazuyuki ; Watanabe, Kimitsuna ; Ramakrishnan, V. ; Suzuki, Tsutomu. / Modified uridines with C5-methylene substituents at the first position of the tRNA anticodon stabilize U·G wobble pairing during decoding. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 27. pp. 18801-18811.
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AU - Ohtsuki, Takashi

AU - Shimazaki, Tomomi

AU - Wada, Takeshi

AU - Kirino, Yohei

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AU - Suzuki, Tsutomu

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