Editing by a tRNA synthetase: DNA aptamer-induced translocation and hydrolysis of a misactivated amino acid.
Aminoacyl-tRNA synthetases establish the rules of the genetic code by aminoacylation reactions. Occasional activation of the wrong amino acid can lead to errors of protein synthesis. For isoleucyl-tRNA synthetase, these errors are reduced by tRNA-dependent hydrolytic editing reactions that occur at a site 25 A from the active site. These reactions require that the misactivated amino acid be translocated from the active site to the center for editing. One mechanism describes translocation as requiring the mischarging of tRNA followed by a conformational change in the tRNA that moves the amino acid from one site to the other. Here a specific DNA aptamer is investigated. The aptamer can stimulate amino acid-specific editing but cannot be aminoacylated. Although the aptamer could in principle stimulate hydrolysis of a misactivated amino acid by an idiosyncratic mechanism, the aptamer is shown here to induce translocation and hydrolysis of misactivated aminoacyl adenylate at the same site as that seen with the tRNA cofactor. Thus, translocation to the site for editing does not require joining of the amino acid to the nucleic acid. Further experiments demonstrated that aptamer-induced editing is sensitive to aptamer sequence and that the aptamer is directed to a site other than the active site or tRNA binding site of the enzyme.
Farrow
M A
MA
The Skaggs Institute for Chemical Biology, The Scripps Research Institute, Beckman Center, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Schimmel
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D000215:Acylation; D019943:Amino Acid Substitution / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D001692:Biological Transport / Q000235:genetics; D003432:Cross-Linking Reagents / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006868:Hydrolysis; D007533:Isoleucine-tRNA Ligase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D017393:RNA Editing / Q000235:genetics; D012355:RNA, Transfer, Ile / Q000235:genetics / Q000378:metabolism*; D046249:Transfer RNA Aminoacylation / Q000235:genetics; D014633:Valine / Q000235:genetics
D003432:Cross-Linking Reagents; D004269:DNA, Bacterial; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D012355:RNA, Transfer, Ile; D007533:Isoleucine-tRNA Ligase; D014633:Valine