The C-Ala Domain Brings Together Editing and Aminoacylation Functions on One tRNA

Abstract

Protein synthesis involves the accurate attachment of amino acids to their matching tRNA molecules. Mistranslating the amino acids serine or glycine for alanine is prevented by the function of independent but collaborative aminoacylation and editing domains of alanyl-tRNA synthetases (AlaRSs). Here we show that the C-Ala domain plays a key role in AlaRS function. The C-Ala domain is universally tethered to the editing domain both in AlaRS and in many homologous free-standing, editing proteins. Crystal structure and functional analyses showed that C-Ala forms an ancient single-stranded nucleic acid binding motif that promotes cooperative binding of both aminoacylation and editing domains to tRNAAla. In addition, C-Ala may have played an essential role in the evolution of AlaRSs by coupling aminoacylation to editing to prevent mistranslation. The algorithm of the genetic code is established in the first reaction of protein synthesis. In this reaction, aminoacyl-tRNA synthetases (AARSs) catalyze the attachment of amino acids to their cognate transfer RNAs (tRNAs) that bear the triplet anticodons of the genetic code. When a tRNA is acylated with the wrong amino acid, mistranslation occurs if the misacylated tRNA is released from the synthetase, captured by elongation factor, and used at the ribosome for peptide synthesis. To prevent mistranslation, some AARSs have separate editing activities that hydrolyze the misacylated amino acid from the tRNA (1–3). Because an editing-defective tRNA synthetase is toxic to bacterial and mammalian cells (4, 5), and is causally linked to disease in animals (6), strong selective pressure retains these editing activities throughout evolution.

Lipid II-independent trans Editing of Mischarged tRNAs by the Penicillin Resistance Factor MurM

“…This is higher than the overall error rate for translation, which is typically from 1/3000 to 1/10,000 (10). Amino acid activation errors can be corrected both by the synthetase itself at a distinct editing site, as is the case for AlaRS, and also by free-standing editing domain homologues, as exemplified by the widely conserved AlaXPs proteins that edit Ser-tRNA Ala (17,18). S. pneumoniae encodes no known AlaXPs, suggesting that the genes encoding these AlaRS editing domain homologues may have been lost from the pneumococcal genome during gene shuffling, which occurs rapidly within the organism as a result of exposure to antibiotics (19).…”

Ancestral AlaX Editing Enzymes for Control of Genetic Code Fidelity Are Not tRNA-specific

“…AlaX-M contains an additional Gly-rich motif in its N-terminal region (31,32). Finally, AlaX-L closely resembles the entire editing domain of AlaRS and contains an additional C-terminal domain (C-Ala) (27,33).…”

“…1) (24). They are homologous to the editing domains of alanyl-and threonyl-tRNA synthetases (AlaRS and ThrRS) (10,22,(25)(26)(27)(28). There are three types of freestanding AlaXs: AlaX-S, AlaX-M, and AlaX-L, which stand for small, medium, and large, respectively ( Fig.…”

“…Although AlaX-M, AlaX-L, and AlaRS hydrolyze both Ser-tRNA Ala and Gly-tRNA Ala (22,25,27,32), AlaX-S only hydrolyzes mischarged Ser-tRNA Ala (29), possibly because of the lack of the Gly-rich domain ( Fig. 2A), which results in an enlarged amino acid binding pocket and a much lower binding affinity toward Gly (29).…”

p23 H implicated as cis/trans regulator of AlaXp-directed editing for mammalian cell homeostasis

“…1A). This extra domain brings together aminoacylation and editing functions to act on bound tRNA Ala (21).…”