5ON3

Quaternary complex of mutant T252A of E. coli leucyl-tRNA synthetase with tRNA(leu), leucyl-adenylate analogue, and post-transfer editing analogue of leucine in the aminoacylation conformation

5ON3 の概要

• エントリーDOI: 10.2210/pdb5on3/pdb
• 関連するPDBエントリー: 5OMW 5ON2 5ON3
• 分子名称: Leucine--tRNA ligase, tRNA(leu), ZINC ION, ... (5 entities in total)
• 機能のキーワード: reaction catalysed: atp + l-leucine + trna(leu) = amp + diphosphate + l-leucyl-trna(leu) protein translation, aminoacyl-trna activity, leucine-trna ligase, class ia, translation, ligase
• 由来する生物種: Escherichia coli K-12; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 256820.48

構造登録者

Palencia, A.,Cusack, S. (登録日: 2017-08-02, 公開日: 2017-11-15, 最終更新日: 2024-01-17)

主引用文献

Dulic, M.,Cvetesic, N.,Zivkovic, I.,Palencia, A.,Cusack, S.,Bertosa, B.,Gruic-Sovulj, I.
Kinetic Origin of Substrate Specificity in Post-Transfer Editing by Leucyl-tRNA Synthetase.
J. Mol. Biol., 430:1-16, 2018

PubMed Abstract: The intrinsic editing capacities of aminoacyl-tRNA synthetases ensure a high-fidelity translation of the amino acids that possess effective non-cognate aminoacylation surrogates. The dominant error-correction pathway comprises deacylation of misaminoacylated tRNA within the aminoacyl-tRNA synthetase editing site. To assess the origin of specificity of Escherichia coli leucyl-tRNA synthetase (LeuRS) against the cognate aminoacylation product in editing, we followed binding and catalysis independently using cognate leucyl- and non-cognate norvalyl-tRNA and their non-hydrolyzable analogues. We found that the amino acid part (leucine versus norvaline) of (mis)aminoacyl-tRNAs can contribute approximately 10-fold to ground-state discrimination at the editing site. In sharp contrast, the rate of deacylation of leucyl- and norvalyl-tRNA differed by about 10-fold. We further established the critical role for the A76 3'-OH group of the tRNA in post-transfer editing, which supports the substrate-assisted deacylation mechanism. Interestingly, the abrogation of the LeuRS specificity determinant threonine 252 did not improve the affinity of the editing site for the cognate leucine as expected, but instead substantially enhanced the rate of leucyl-tRNA hydrolysis. In line with that, molecular dynamics simulations revealed that the wild-type enzyme, but not the T252A mutant, enforced leucine to adopt the side-chain conformation that promotes the steric exclusion of a putative catalytic water. Our data demonstrated that the LeuRS editing site exhibits amino acid specificity of kinetic origin, arguing against the anticipated prominent role of steric exclusion in the rejection of leucine. This feature distinguishes editing from the synthetic site, which relies on ground-state discrimination in amino acid selection.

PubMed: 29111343
DOI: 10.1016/j.jmb.2017.10.024

実験手法

X-RAY DIFFRACTION (3.1 Å)