Signatures of tRNA-specificity in proteobacterial glutamyl-tRNA synthetases.

The canonical function of glutamyl-tRNA synthetase (GluRS) is to glutamylate tRNA. Yet not all bacterial GluRSs glutamylate tRNA; many glutamylate both tRNA and tRNA, while some glutamylate only tRNA and not the cognate substrate tRNA. Understanding the basis of the unique specificity of tRNA is important. Mutational studies have hinted at hotspot residues, both on tRNA and GluRS, which play crucial roles in tRNA-specificity. However, its underlying structural basis remains unexplored. The majority of biochemical studies related to tRNA-specificity have been performed on GluRS from Escherichia coli and other proteobacterial species. However, since the early crystal structures of GluRS and tRNA-bound GluRS were from non-proteobacterial species (Thermus thermophilus), proteobacterial biochemical data have often been interpreted in the context of non-proteobacterial GluRS structures. Marked differences between proteobacterial and non-proteobacterial GluRSs have been demonstrated; therefore, it is important to understand tRNA-specificity vis-a-vis proteobacterial GluRS structures. To this end, we solved the crystal structure of a double mutant GluRS from E. coli. Using the solved structure and several other currently available proteo- and non-proteobacterial GluRS crystal structures, we probed the structural basis of the tRNA-specificity of bacterial GluRSs. Specifically, our analyses suggest a unique role played by the tRNA D-helix contacting loop of GluRS in the modulation of tRNA-specificity. While earlier studies have identified functional hotspots on tRNA that control the tRNA-specificity of GluRS, this is the first report of complementary signatures of tRNA-specificity in GluRS.