Unravelling the plasticity of substrate recognition by Staphylococcus aureus lysyl-tRNA synthetase and its implications for misacylation

Abstract

Transfer RNA (tRNA) misacylation is a widespread phenomenon that affects translational fidelity due to the incorporation of non-cognate amino acids into proteins. We investigated the structural basis for the misacylation of tRNALys by Staphylococcus aureus lysyl-tRNA synthetase (SaLysRS). Activity studies showed that SaLysRS misacylated tRNALys with methionine and arginine. In vivo studies and MALDI-TOF analysis revealed the utilisation of these mischarged tRNAs in protein translation, as deciphered from the incorporation of non-cognate methionine and arginine into proteins. The misincorporation was also detrimental to cell growth. The three-dimensional structure of SaLysRS with its cognate substrate lysine was resolved at 2.3 Å resolution, which revealed key residues and conserved motifs needed for substrate recognition. Structural and mutational analysis and molecular dynamics simulations identified Glu233, Tyr273 and Glu420 as crucial residues for both cognate and non-cognate ligand binding. These insights, well-supported by structural, biochemical and computational data, enhance our knowledge of the mechanisms underlying misacylation in tRNA synthetases and its implications for cell growth.