title: Two proteomic methodologies for defining N-termini of mature human mitochondrial aminoacyl-tRNA synthetases.
abstract: Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are encoded in the nucleus, synthesized in the cytosol and targeted for importation into mitochondria by a N-terminal mitochondrial targeting sequence. This targeting sequence is presumably cleaved upon entry into the mitochondria, following a process still not fully deciphered in human, despite essential roles for the mitochondrial biogenesis. Maturation processes are indeed essential both for the release of a functional enzyme and to route correctly the protein within mitochondria. The absence of consensus sequences for cleavage sites and the discovery of possible multiple proteolytic steps render predictions of N-termini difficult. Further, the knowledge of the cleavages is key for the design of protein constructions compatible with efficient production in bacterial strains. Finally, full comprehension becomes essential because a growing number of mutations are found in genes coding for mt-aaRS. In the present study, we take advantage of proteomic methodological developments and identified, in mitochondria, three N-termini for the human mitochondrial aspartyl-tRNA synthetase. This first description of the co-existence of different forms opens new perspectives in the biological understanding of this enzyme. Those methods are extended to the whole set of human mt-aaRSs and methodological advice are provided for further investigations.
authors: Laboratoire de Spectrométrie de Masse BioOrganique, Université de Strasbourg, CNRS, IPHC, UMR 7178, F-67000 Strasbourg, France.
authors: Christine
authors: C
authors: Carapito
authors: Plateforme Protéomique Strasbourg - Esplanade, Institut de Biologie Moléculaire et Cellulaire, CNRS, FRC 1589, 15 rue Descartes, F-67084 Strasbourg Cedex, France.
authors: Lauriane
authors: L
authors: Kuhn
authors: Université de Strasbourg, CNRS, Architecture et Réactivité de l'ARN, UPR 9002, F-67000 Strasbourg, France.
authors: Loukmane
authors: L
authors: Karim
authors: Laboratoire de Spectrométrie de Masse BioOrganique, Université de Strasbourg, CNRS, IPHC, UMR 7178, F-67000 Strasbourg, France.
authors: Magali
authors: M
authors: Rompais
authors: Laboratoire de Chimie et Biologie des Métaux, UMR CNRS-CEA-UGA 5249, iRTSV/LCBM, CEA Grenoble, Grenoble, France.
authors: Thierry
authors: T
authors: Rabilloud
authors: Université de Strasbourg, CNRS, Architecture et Réactivité de l'ARN, UPR 9002, F-67000 Strasbourg, France.
authors: Hagen
authors: H
authors: Schwenzer
authors: Université de Strasbourg, CNRS, Architecture et Réactivité de l'ARN, UPR 9002, F-67000 Strasbourg, France. Electronic address: M.Sissler@ibmc-cnrs.unistra.fr.
authors: Marie
authors: M
authors: Sissler
mesh_terms: D000595:Amino Acid Sequence; D000604:Amino Acyl-tRNA Synthetases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D002458:Cell Fractionation / Q000379:methods; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000201:enzymology* / Q000235:genetics; D003600:Cytosol / Q000737:chemistry / Q000201:enzymology*; D057809:HEK293 Cells; D006801:Humans; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics; D009000:Monocytes / Q000166:cytology / Q000201:enzymology; D010446:Peptide Fragments / Q000032:analysis; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals; D021381:Protein Transport; D040901:Proteomics / Q000295:instrumentation / Q000379:methods*
chemical_list: D010446:Peptide Fragments; D011498:Protein Precursors; D021382:Protein Sorting Signals; D000604:Amino Acyl-tRNA Synthetases
keywords: Aminoacyl-tRNA synthetases; Human; Maturation; Mitochondria; Mitochondrial targeting sequence; N-terminomics; Shotgun proteomics