Widespread Distribution of Cell Defense against d-Aminoacyl-tRNAs

Abstract

Several L-aminoacyl-tRNA synthetases can transfer a D-amino acid onto their cognate tRNA(s). This harmful reaction is counteracted by the enzyme D-aminoacyl-tRNA deacylase. Two distinct deacylases were already identified in bacteria (DTD1) and in archaea (DTD2), respectively. Evidence was given that DTD1 homologs also exist in nearly all eukaryotes, whereas DTD2 homologs occur in plants. On the other hand, several bacteria, including most cyanobacteria, lack genes encoding a DTD1 homolog. Here we show that Synechocystis sp. PCC6803 produces a third type of deacylase (DTD3). Inactivation of the corresponding gene (dtd3) renders the growth of Synechocystis sp. hypersensitive to the presence of D-tyrosine. Based on the available genomes, DTD3-like proteins are predicted to occur in all cyanobacteria. Moreover, one or several dtd3-like genes can be recognized in all cellular types, arguing in favor of the nearubiquity of an enzymatic function involved in the defense of translational systems against invasion by D-amino acids.Although they are detected in various living organisms (reviewed in Ref. 1), D-amino acids are thought not to be incorporated into proteins, because of the stereospecificity of aminoacyl-tRNA synthetases and of the translational machinery, including EF-Tu and the ribosome (2). However, the discrimination between L-and D-amino acids by aminoacyl-tRNA synthetases is not equal to 100%. Significant D-aminoacylation of their cognate tRNAs by Escherichia coli tyrosyl-, tryptophanyl-, aspartyl-, lysyl-, and histidyl-tRNA synthetases has been characterized in vitro (3-9). Recently, using a bacterium, transfer of D-tyrosine onto tRNA Tyr was shown to occur in vivo (10). With such misacylation reactions, the resulting D-aminoacyl-tRNAs form a pool of metabolically inactive molecules, at best. At worst, D-aminoacylated tRNAs infiltrate the protein synthesis machinery. Although the latter harmful possibility has not yet been firmly established, several cells were shown to possess a D-tyrosyl-tRNA deacylase, or DTD, that should help them counteract the accumulation of D-aminoacyl-tRNAs. This enzyme shows a broad specificity, being able to remove various D-aminoacyl moieties from the 3Ј-end of a tRNA (4 -6, 11). Such a function makes the deacylase a member of the family of enzymes capable of editing in trans mis-aminoacylated tRNAs. This family includes several homologs of aminoacyl-tRNA synthetase editing domains (12), as well as peptidyl-tRNA hydrolase (13,14).Two distinct deacylases have already been discovered. The first one, called DTD1, is predicted to occur in most bacteria and eukaryotes (see Table 1). Inactivation of the gene of this deacylase in E. coli (dtd) or in Saccharomyces cerevisiae (DTD1) exacerbates cell growth inhibition by several D-amino acids, including D-tyrosine (6). In fact, in an E. coli ⌬dtd strain grown in the presence of 2.4 mM D-tyrosine, as much as 40% of the cellular tRNATyr pool becomes esterified with D-tyrosine (10). Homologs of dtd/DTD1 are not found in the available archae...

The ribosome can discriminate the chirality of amino acids within its peptidyl-transferase center

“…Second, there is growing evidence that ribosomes may have to contend with D-aa-tRNAs in vivo: D-amino acids are synthesized by racemase enzymes (6) and can be found at high concentrations in cells (7), and a growing number of aa-tRNA synthetase (aaRS) enzymes exhibit the ability to misacylate tRNAs with D-amino acids (8,9). Consistent with this possibility, the D-aa-tRNA deacylase (DTD) enzyme is nearly universally conserved (10) and functions to remove D-amino acids that have been misacylated onto tRNA (8,10).…”

Origin, Microbiology, Nutrition, and Pharmacology of D‐Amino Acids

“…This finding suggests that tRNA aminoacylation, a process by which amino acids first encounter RNA, may have been a critical step in determining LAA homochirality. However, it should be noted that cells contain defenses against daminoacyl-tRNAs [10].…”

Initiator tRNA genes template the 3′ CCA end at high frequencies in bacteria

“…Tyrosine residues are critically important for both catalysis and stability of RuBisCo [43], one of the most abundant proteins in Cyanobacteria [44]. It is remarkable that there exists a D-Tyr-tRNA Tyr deacylase that is conserved and apparently unique to Cyanobacteria [45], which helps maintain the accuracy of tRNA Tyr charging. Competition experiments that model biologically relevant conditions with Cyanobacterial strains with or without CCA-templating for tRNA Tyr , as well as biochemical assays, could test this hypothesis.…”