PMID:2129559
| Citation |
Eriani, G, Dirheimer, G and Gangloff, J (1990) Aspartyl-tRNA synthetase from Escherichia coli: cloning and characterisation of the gene, homologies of its translated amino acid sequence with asparaginyl- and lysyl-tRNA synthetases. Nucleic Acids Res. 18:7109-18 |
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| Abstract |
By screening of an Escherichia coli plasmidic library using antibodies against aspartyl-tRNA synthetase (AspRS) several clones were obtained containing aspS, the gene coding for AspRS. We report here the nucleotide sequence of aspS and the corresponding primary structure of the aspartyl-tRNA synthetase, a protein of 590 amino acid residues with a Mr 65,913, a value in close agreement with that observed for the purified protein. Primer extension analysis of the aspS mRNA using reverse transcriptase located its 5'-end at 94 nucleotides upstream of the translation initiation AUG; nuclease S1 analysis located the 3'-end at 126 nucleotides downstream of the stop codon UGA. Comparison of the DNA-derived protein sequence with known aminoacyl-tRNA sequences revealed important homologies with asparaginyl- and lysyl-tRNA synthetases from E.coli; more than 25% of their amino acid residues are identical, the homologies being distributed preferencially in the first part and the carboxy-terminal end of the molecule. Mutagenesis directed towards a consensus tetrapeptide (Gly-Leu-Asp-Arg) and the carboxy-terminal end showed that both domains could be implicated in catalysis as well as in ATP binding. |
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| Keywords |
Amino Acid Sequence; Amino Acyl-tRNA Synthetases/genetics; Amino Acyl-tRNA Synthetases/metabolism; Aspartate-tRNA Ligase/genetics; Aspartate-tRNA Ligase/isolation & purification; Aspartate-tRNA Ligase/metabolism; Base Sequence; Cloning, Molecular; Codon; Escherichia coli/enzymology; Escherichia coli/genetics; Genes, Bacterial; Lysine-tRNA Ligase/genetics; Lysine-tRNA Ligase/metabolism; Molecular Sequence Data; Mutagenesis, Site-Directed; Protein Biosynthesis; RNA, Transfer, Amino Acyl; Restriction Mapping; Sequence Alignment; Sequence Homology, Nucleic Acid |
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