Structural basis for non-cognate amino acid discrimination by the valyl-tRNA synthetase editing domain

Ryuya Fukunaga, Shigeyuki Yokoyama

Research output: Contribution to journalArticle

Abstract

The editing domain of valyl-tRNA synthetase (ValRS) is known to deacylate, or edit, misformed Thr-tRNAVal (post-transfer editing). Here, we determined the 1.7-Å-resolution crystal structure of the Thermus thermophilus ValRS editing domain. A comparison of the structure with the previously reported tRNA complex structure revealed conformational changes of the editing domain upon accommodation of the terminal A76; the "GTG loop" moves to expand the pocket, and the side chain of Phe-264 on the GTG loop rotates to interact with the A76 adenine ring. If these conformational changes did not occur, then C75 and A76 of the tRNA would clash with Phe-264. To elucidate the mechanism of the threonine side-chain recognition, we determined the crystal structure of the editing domain bound with [N-(L-threonyl)- sulfamoyl]adenosine at 1.7-Å resolution. The γ-OH of the threonyl moiety is recognized by the Lys-270, Thr-272, and Asp-279 side chains, which may reject the cognate valyl moiety. Accordingly, ValRS mutants with an Ala substitution for Lys-270 or Asp-279 synthesized significant amounts of Thr-tRNAVal. The misproduced Thr-tRNAVal was hydrolyzed efficiently by the wild-type ValRS, but this post-transfer editing activity was drastically impaired by the Ala substitutions for Lys-270 and Asp-279 and was also decreased by those for Arg-216, Phe-264, and Thr-272. These results indicate that the threonyl moiety and A76 of Thr-tRNAVal are recognized by the Lys-270, Thr-272, and Asp-279 side chains and by the Phe-264 side chain, respectively, of the ValRS editing domain.

Original languageEnglish (US)
Pages (from-to)29937-29945
Number of pages9
JournalJournal of Biological Chemistry
Volume280
Issue number33
DOIs
StatePublished - Aug 19 2005
Externally publishedYes

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Valine-tRNA Ligase
RNA, Transfer, Val
Amino Acids
Transfer RNA
Substitution reactions
Crystal structure
Thermus thermophilus
Adenine
Threonine
Adenosine

ASJC Scopus subject areas

  • Biochemistry

Cite this

Structural basis for non-cognate amino acid discrimination by the valyl-tRNA synthetase editing domain. / Fukunaga, Ryuya; Yokoyama, Shigeyuki.

In: Journal of Biological Chemistry, Vol. 280, No. 33, 19.08.2005, p. 29937-29945.

Research output: Contribution to journalArticle

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AB - The editing domain of valyl-tRNA synthetase (ValRS) is known to deacylate, or edit, misformed Thr-tRNAVal (post-transfer editing). Here, we determined the 1.7-Å-resolution crystal structure of the Thermus thermophilus ValRS editing domain. A comparison of the structure with the previously reported tRNA complex structure revealed conformational changes of the editing domain upon accommodation of the terminal A76; the "GTG loop" moves to expand the pocket, and the side chain of Phe-264 on the GTG loop rotates to interact with the A76 adenine ring. If these conformational changes did not occur, then C75 and A76 of the tRNA would clash with Phe-264. To elucidate the mechanism of the threonine side-chain recognition, we determined the crystal structure of the editing domain bound with [N-(L-threonyl)- sulfamoyl]adenosine at 1.7-Å resolution. The γ-OH of the threonyl moiety is recognized by the Lys-270, Thr-272, and Asp-279 side chains, which may reject the cognate valyl moiety. Accordingly, ValRS mutants with an Ala substitution for Lys-270 or Asp-279 synthesized significant amounts of Thr-tRNAVal. The misproduced Thr-tRNAVal was hydrolyzed efficiently by the wild-type ValRS, but this post-transfer editing activity was drastically impaired by the Ala substitutions for Lys-270 and Asp-279 and was also decreased by those for Arg-216, Phe-264, and Thr-272. These results indicate that the threonyl moiety and A76 of Thr-tRNAVal are recognized by the Lys-270, Thr-272, and Asp-279 side chains and by the Phe-264 side chain, respectively, of the ValRS editing domain.

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