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Title: Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase

Abstract

Crystals of human mitochondrial tyrosyl-tRNA synthetase lacking the C-terminal S4-like domain diffract to 2.7 Å resolution and are suitable for structure determination. Human mitochondrial tyrosyl-tRNA synthetase and a truncated version with its C-terminal S4-like domain deleted were purified and crystallized. Only the truncated version, which is active in tyrosine activation and Escherichia coli tRNA{sup Tyr} charging, yielded crystals suitable for structure determination. These tetragonal crystals, belonging to space group P4{sub 3}2{sub 1}2, were obtained in the presence of PEG 4000 as a crystallizing agent and diffracted X-rays to 2.7 Å resolution. Complete data sets could be collected and led to structure solution by molecular replacement.

Authors:
; ; ; ; ; ; ;  [1]
  1. Département ‘Machineries Traductionnelles’, Architecture et Réactivité de l’ARN, Université Louis Pasteur de Strasbourg, CNRS, IBMC, 15 Rue René Descartes, 67084 Strasbourg (France)
Publication Date:
OSTI Identifier:
22360303
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F; Journal Volume: 63; Journal Issue: Pt 4; Other Information: PMCID: PMC2330213; PMID: 17401211; PUBLISHER-ID: hc5025; OAI: oai:pubmedcentral.nih.gov:2330213; Copyright (c) International Union of Crystallography 2007; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CRYSTALLIZATION; CRYSTALS; ESCHERICHIA COLI; MATHEMATICAL SOLUTIONS; RESOLUTION; SOLUTIONS; SPACE GROUPS

Citation Formats

Bonnefond, Luc, Frugier, Magali, Touzé, Elodie, Lorber, Bernard, Florentz, Catherine, Giegé, Richard, E-mail: r.giege@ibmc.u-strasbg.fr, Rudinger-Thirion, Joëlle, and Sauter, Claude. Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase. United Kingdom: N. p., 2007. Web. doi:10.1107/S1744309107012481.
Bonnefond, Luc, Frugier, Magali, Touzé, Elodie, Lorber, Bernard, Florentz, Catherine, Giegé, Richard, E-mail: r.giege@ibmc.u-strasbg.fr, Rudinger-Thirion, Joëlle, & Sauter, Claude. Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase. United Kingdom. doi:10.1107/S1744309107012481.
Bonnefond, Luc, Frugier, Magali, Touzé, Elodie, Lorber, Bernard, Florentz, Catherine, Giegé, Richard, E-mail: r.giege@ibmc.u-strasbg.fr, Rudinger-Thirion, Joëlle, and Sauter, Claude. Sun . "Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase". United Kingdom. doi:10.1107/S1744309107012481.
@article{osti_22360303,
title = {Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase},
author = {Bonnefond, Luc and Frugier, Magali and Touzé, Elodie and Lorber, Bernard and Florentz, Catherine and Giegé, Richard, E-mail: r.giege@ibmc.u-strasbg.fr and Rudinger-Thirion, Joëlle and Sauter, Claude},
abstractNote = {Crystals of human mitochondrial tyrosyl-tRNA synthetase lacking the C-terminal S4-like domain diffract to 2.7 Å resolution and are suitable for structure determination. Human mitochondrial tyrosyl-tRNA synthetase and a truncated version with its C-terminal S4-like domain deleted were purified and crystallized. Only the truncated version, which is active in tyrosine activation and Escherichia coli tRNA{sup Tyr} charging, yielded crystals suitable for structure determination. These tetragonal crystals, belonging to space group P4{sub 3}2{sub 1}2, were obtained in the presence of PEG 4000 as a crystallizing agent and diffracted X-rays to 2.7 Å resolution. Complete data sets could be collected and led to structure solution by molecular replacement.},
doi = {10.1107/S1744309107012481},
journal = {Acta Crystallographica. Section F},
number = Pt 4,
volume = 63,
place = {United Kingdom},
year = {Sun Apr 01 00:00:00 EDT 2007},
month = {Sun Apr 01 00:00:00 EDT 2007}
}
  • The expression, purification and crystallization of recombinant human mitochondrial phenylalanyl-tRNA synthetase (mitPheRS) are reported. Diffraction data were collected to 2.2 Å resolution and the mitPheRS structure was solved using the molecular-replacement method. Human monomeric mitochondrial phenylalanyl-tRNA synthetase (mitPheRS) is an enzyme that catalyzes the charging of tRNA with the cognate amino acid phenylalanine. Human mitPheRS is a chimera of the bacterial α-subunit of PheRS and the B8 domain of its β-subunit. Together, the α-subunit and the ‘RNP-domain’ (B8 domain) at the C-terminus form the minimal structural set to construct an enzyme with phenylalanylation activity. The recombinant human mitPheRS was purifiedmore » to homogeneity and crystallized in complex with phenylalanine and ATP. The crystals diffracted to 2.2 Å resolution and belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 55, b = 90, c = 96 Å.« less
  • Tyrosyl-tRNA synthetase from the hyperthermophilic archaeon A. pernix K1 was cloned, purified and crystallized. The crystals belonged to the tetragonal space group P4{sub 3}2{sub 1}2, with unit-cell parameters a = b = 66.1, c = 196.2 Å, and diffracted to beyond 2.15 Å resolution at 100 K. Hyperthermophilic archaeal tyrosyl-tRNA synthetase from Aeropyrum pernix K1 was cloned and overexpressed in Escherichia coli. The expressed protein was purified by Cibacron Blue affinity chromatography following heat treatment at 363 K. Crystals suitable for X-ray diffraction studies were obtained under optimized crystallization conditions in the presence of 1.5 M ammonium sulfate using themore » hanging-drop vapour-diffusion method. The crystals belonged to the tetragonal space group P4{sub 3}2{sub 1}2, with unit-cell parameters a = b = 66.1, c = 196.2 Å, and diffracted to beyond 2.15 Å resolution at 100 K.« less
  • No abstract prepared.
  • Glutamyl-tRNA synthetase and prolyl-tRNA synthetase belong to different classes of aminoacyl-tRNA synthetases that are thought to have evolved along independent evolutionary pathways. However, both enzymes are on one polypeptide chain encoded by a single human gene, the EPRS locus, which is transcribed as one long mRNA. The authors report the structure of the human EPRS gene, which consists of 29 exons spread over at least 90 kb of genomic DNA. The exons, encoding the glutamyl-specific and the prolyl-specific parts of the enzyme, are each clustered in 10-kb sections located at opposite ends of the gene. These two exon clusters aremore » separated by a long intervening DNA section with a number of exons, encoding functions that may be involved in the organization of the mammalian multienzyme synthetase complex. The upstream gene region shows structural features of a regulated gene, and preliminary experiments suggest that the gene is expressed at specific times in growth-stimulated cultured cells. The authors have localized the gene to the distal long arm of human chromosome 1 and to a corresponding site in mouse chromosome 1. 64 refs., 6 figs., 2 tabs.« less