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Query: EC:6.1.1.24 (nondiscriminating glutamyl-tRNA synthetase)
 2 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The specificity of most aminoacyl-tRNA synthetases for an amino acid and cognate tRNA pair evolved before the divergence of the three domains of life. Glutaminyl-tRNA synthetase (GlnRS) evolved later and is derived from the archaeal-type nondiscriminating glutamyl-tRNA synthetase (GluRS), an enzyme with relaxed tRNA specificity capable of forming both Glu-tRNA(Glu) and Glu-tRNA(Gln). The archaea lack GlnRS and use a specialized amidotransferase to convert Glu-tRNA(Gln) to Gln-tRNA(Gln) needed for protein synthesis. We show that the Methanothermobacter thermautotrophicus GluRS is active toward tRNA(Glu) and the two tRNA(Gln) isoacceptors the organism encodes, but with a significant catalytic preference for tRNA(Gln2)(CUG). The less active tRNA(Gln1)(UUG) responds to the less common CAA codon for Gln. From a biochemical characterization of M. thermautotrophicus GluRS variants, we found that the evolution of tRNA specificity in GlnRS could be recapitulated by converting the M. thermautotrophicus GluRS to a tRNA(Gln) specific enzyme, solely through the addition of an acceptor stem loop present in bacterial GlnRS. One designed GluRS variant is also highly specific for the tRNA(Gln2)(CUG) isoacceptor, which responds to the CAG codon, and shows no activity toward tRNA(Gln1)(UUG). Because it is now possible to eliminate particular codons from the genome of Escherichia coli, additional codons will become available for genetic code engineering. Isoacceptor-specific aminoacyl-tRNA synthetases will enable the reassignment of more open codons while preserving accurate encoding of the 20 canonical amino acids.
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PMID:Rational design of an evolutionary precursor of glutaminyl-tRNA synthetase. 2215 97

The glutaminyl-tRNA synthetase (GlnRS) enzyme, which pairs glutamine with tRNA(Gln) for protein synthesis, evolved by gene duplication in early eukaryotes from a nondiscriminating glutamyl-tRNA synthetase (GluRS) that aminoacylates both tRNA(Gln) and tRNA(Glu) with glutamate. This ancient GluRS also separately differentiated to exclude tRNA(Gln) as a substrate, and the resulting discriminating GluRS and GlnRS further acquired additional protein domains assisting function in cis (the GlnRS N-terminal Yqey domain) or in trans (the Arc1p protein associating with GluRS). These added domains are absent in contemporary bacterial GlnRS and GluRS. Here, using Saccharomyces cerevisiae enzymes as models, we find that the eukaryote-specific protein domains substantially influence amino acid binding, tRNA binding and aminoacylation efficiency, but they play no role in either specific nucleotide readout or discrimination against noncognate tRNA. Eukaryotic tRNA(Gln) and tRNA(Glu) recognition determinants are found in equivalent positions and are mutually exclusive to a significant degree, with key nucleotides located adjacent to portions of the protein structure that differentiated during the evolution of archaeal nondiscriminating GluRS to GlnRS. These findings provide important corroboration for the evolutionary model and suggest that the added eukaryotic domains arose in response to distinctive selective pressures associated with the greater complexity of the eukaryotic translational apparatus. We also find that the affinity of GluRS for glutamate is significantly increased when Arc1p is not associated with the enzyme. This is consistent with the lower concentration of intracellular glutamate and the dissociation of the Arc1p:GluRS complex upon the diauxic shift to respiratory conditions.
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PMID:Coevolution of specificity determinants in eukaryotic glutamyl- and glutaminyl-tRNA synthetases. 2514 3