Dual targeting of a tRNAAsp requires two different aspartyl-tRNA synthetases in Trypanosoma brucei.
The mitochondrion of the parasitic protozoon Trypanosoma brucei does not encode any tRNAs. This deficiency is compensated for by partial import of nearly all of its cytosolic tRNAs. Most trypanosomal aminoacyl-tRNA synthetases are encoded by single copy genes, suggesting the use of the same enzyme in the cytosol and in the mitochondrion. However, the T. brucei genome encodes two distinct genes for eukaryotic aspartyl-tRNA synthetase (AspRS), although the cell has a single tRNAAsp isoacceptor only. Phylogenetic analysis showed that the two T. brucei AspRSs evolved from a duplication early in kinetoplastid evolution and also revealed that eight other major duplications of AspRS occurred in the eukaryotic domain. RNA interference analysis established that both Tb-AspRS1 and Tb-AspRS2 are essential for growth and required for cytosolic and mitochondrial Asp-tRNAAsp formation, respectively. In vitro charging assays demonstrated that the mitochondrial Tb-AspRS2 aminoacylates both cytosolic and mitochondrial tRNAAsp, whereas the cytosolic Tb-AspRS1 selectively recognizes cytosolic but not mitochondrial tRNAAsp. This indicates that cytosolic and mitochondrial tRNAAsp, although derived from the same nuclear gene, are physically different, most likely due to a mitochondria-specific nucleotide modification. Mitochondrial Tb-AspRS2 defines a novel group of eukaryotic AspRSs with an expanded substrate specificity that are restricted to trypanosomatids and therefore may be exploited as a novel drug target.
Charrière
Fabien
F
From the Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland.
O'Donoghue
Patrick
P
Department of Molecular Biophysics and Biochemistry and Department of Chemistry, Yale University, New Haven, Connecticut 06520-8114.
Helgadóttir
Sunna
S
Department of Molecular Biophysics and Biochemistry and Department of Chemistry, Yale University, New Haven, Connecticut 06520-8114.
Maréchal-Drouard
Laurence
L
Institut de Biologie Moléculaire des Plantes, Unité Propre de Recherche 2357 du CNRS, University of Strasbourg, 12 Rue du Général Zimmer, 67084 Strasbourg Cedex, France.
Cristodero
Marina
M
From the Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland.
Horn
Elke K
EK
From the Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland.
Söll
Dieter
D
Department of Molecular Biophysics and Biochemistry and Department of Chemistry, Yale University, New Haven, Connecticut 06520-8114.
Schneider
André
A
From the Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland. Electronic address: andre.schneider@ibc.unibe.ch.
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001226:Aspartate-tRNA Ligase / Q000235:genetics* / Q000378:metabolism*; D003600:Cytosol / Q000201:enzymology; D015195:Drug Design; D008928:Mitochondria / Q000201:enzymology; D010802:Phylogeny; D012349:RNA, Transfer, Asp / Q000096:biosynthesis*; D013379:Substrate Specificity; D046249:Transfer RNA Aminoacylation / Q000502:physiology; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development
D012349:RNA, Transfer, Asp; D000255:Adenosine Triphosphate; D001226:Aspartate-tRNA Ligase