title: A domain in the N-terminal extension of class IIb eukaryotic aminoacyl-tRNA synthetases is important for tRNA binding.
abstract: Cytoplasmic aspartyl-tRNA synthetase (AspRS) from Saccharomyces cerevisiae is a homodimer of 64 kDa subunits. Previous studies have emphasized the high sensitivity of the N-terminal region to proteolytic cleavage, leading to truncated species that have lost the first 20-70 residues but that retain enzymatic activity and dimeric structure. In this work, we demonstrate that the N-terminal extension in yeast AspRS participates in tRNA binding and we generalize this finding to eukaryotic class IIb aminoacyl-tRNA synthetases. By gel retardation studies and footprinting experiments on yeast tRNA(Asp), we show that the extension, connected to the anticodon-binding module of the synthetase, contacts tRNA on the minor groove side of its anticodon stem. Sequence comparison of eukaryotic class IIb synthetases identifies a lysine-rich 11 residue sequence ((29)LSKKALKKLQK(39) in yeast AspRS with the consensus xSKxxLKKxxK in class IIb synthetases) that is important for this binding. Direct proof of the role of this sequence comes from a mutagenesis analysis and from binding studies using the isolated peptide.
authors.affiliation: Département 'Mécanismes et Macromolécules de la Synthèse Protéique et Cristallogenèse', UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 rue René Descartes, 67084 Strasbourg Cedex, France.
authors.forename: M
authors.initials: M
authors.lastname: Frugier
authors.forename: L
authors.initials: L
authors.lastname: Moulinier
authors.forename: R
authors.initials: R
authors.lastname: Giegé
mesh_terms: D000595:Amino Acid Sequence; D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry / Q000378:metabolism*; D001226:Aspartate-tRNA Ligase / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D012331:RNA, Fungal / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: D012331:RNA, Fungal; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases; D001226:Aspartate-tRNA Ligase