The peculiar architectural framework of tRNASec is fully recognized by yeast AspRS.
The wild-type transcript of Escherichia coli tRNASec, characterized by a peculiar core architecture and a large variable region, was shown to be aspartylatable by yeast AspRS. Similar activities were found for tRNASec mutants with methionine, leucine, and tryptophan anticodons. The charging efficiency of these molecules was found comparable to that of a minihelix derived from tRNAAsp and is accounted for by the presence of the discriminator residue G73, which is a major aspartate identity determinant. Introducing the aspartate identity elements from the anticodon loop (G34, U35, C36, C38) into tRNASec transforms this molecule into an aspartate acceptor with kinetic properties identical to tRNAAsp. Expression of the aspartate identity set in tRNASec is independent of the size of its variable region. The functional study was completed by footprinting experiments with four different nucleases as structural probes. Protection patterns by AspRS of transplanted tRNASec and tRNAAsp were found similar. They are modified, particularly in the anticodon loop, upon changing the aspartate anticodon into that of methionine. Altogether, it appears that recognition of a tRNA by AspRS is more governed by the presence of the aspartate identity set than by the structural framework that carries this set.
Rudinger-Thirion
J
J
Unité Propre de Recherche 9002 Structure des Macromolécules Biologiques et Mécanismes de Reconnaissance, Institut de Biologie Moléculaire et Cellulaire du Centre National de la Recherche Scientifique, Strasbourg, France.
Giegé
R
R
D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics* / Q000378:metabolism; D000926:Anticodon / Q000235:genetics; D001224:Aspartic Acid / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D005658:Fungi / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000235:genetics; D012345:RNA, Transfer, Amino Acid-Specific / Q000235:genetics*; D012349:RNA, Transfer, Asp / Q000235:genetics; D017279:Selenocysteine / Q000235:genetics / Q000378:metabolism
D000926:Anticodon; D012329:RNA, Bacterial; D012345:RNA, Transfer, Amino Acid-Specific; D012349:RNA, Transfer, Asp; C036928:tRNA, selenocysteine-; D017279:Selenocysteine; D001224:Aspartic Acid; D000604:Amino Acyl-tRNA Synthetases