Trans-editing of mischarged tRNAs

Abstract

Aminoacyl-tRNA synthetases (aaRSs) are multidomain proteins that specifically attach amino acids to their cognate tRNAs. Their most conserved, and presumably evolutionarily oldest, domains are the catalytic cores, which activate amino acids and transfer them to the 3 ends of tRNAs. Additional domains appended to or inserted in the body of aaRSs increase efficiency and specificity of the aminoacylation process, either by providing additional tRNA contacts, or by hydrolyzing noncognate amino acid products (cisediting). Here, we report specific tRNA-dependent trans-editing by aaRS-like proteins that reciprocate the editing domains of aaRSs, but not the remainder of the corresponding enzyme. A freestanding homologue of the prolyl-tRNA synthetase-editing domain, the PrdX protein from Clostridium sticklandii, efficiently and specifically hydrolyzes Ala-tRNA Pro . Similarly, autonomous alanyl-tRNA synthetase-editing domain homologues (AlaX proteins) from Methanosarcina barkeri and Sulfolobus solfataricus hydrolyze SertRNA Ala and Gly-tRNA Ala substrates. The discovery of autonomous editing proteins efficient in hydrolyzing misacylated products provides a direct link between ancestral aaRSs consisting solely of the catalytic core and extant enzymes to which functionally independent modules are appended.

Molecular basis of alanine discrimination in editing site

“…The AlaX͞AlaRS editing domains are evolutionarily related to ThrRS-N2 and share the characteristic HXXXH and CXXXH motifs ( Fig. 1) (4,8), which were also shown to be important for the deacylation activity of AlaRS (9). The inclusion of noncognate serine and glycine in the AlaRS activation site was reported in ref.…”

“…Therefore, during evolution, an additional editing domain that specifically hydrolyzes mischarged tRNAs has assembled with the catalytic domain to comprise contemporary aaRSs (2). In accordance with this model, the genes that autonomously encode an editing domain were distributed in many organisms (3)(4)(5), and, indeed, some of them are shown to be responsible for the transediting activity of mischarged tRNAs (4,5). AlaX is the one such protein that shows homology to the class II alanyl-tRNA synthetase (AlaRS) editing domain ( Fig.…”

“…1) and is widely scattered among all three kingdoms of life (3). The specific activities of the archaeal AlaXs from Methanosarcina barkeri and Sulfolobus solfataricus have recently been shown to specifically hydrolyze mischarged Ser-and Gly-tRNA Ala in vitro (4).…”

A truncated aminoacyl–tRNA synthetase modifies RNA

“…The function of aaRS paralogs is not restricted to amino acid biosynthesis. Truncated alanyl-, prolyl-, and threonyl-tRNA synthetase-like proteins were shown to possess esterase function and be capable of specific hydrolysis of misacylated tRNA in trans (11,12), a mechanism that should increase the fidelity of the translation process.…”

Turning tRNA upside down: When aminoacylation is not a prerequisite to protein synthesis

“…Functions have been deduced for only a small fraction of these proteins, and, in most cases, it is the catalytic core domains that have been recruited for a variety of new roles. These include roles in amino acid biosynthesis (4,5), DNA replication processivity (6), and protein synthesis quality control (7,8). The paper by Salazar et al (9) in this issue of PNAS adds to this list by describing how the glutamyl-tRNA synthetase (GluRS) paralog YadB attaches glutamate to queuosine, generating a hypermodified nucleoside at the first anticodon position of tRNA Asp .…”