Adaptation to tRNA acceptor stem structure by flexible adjustment in the catalytic domain of class I tRNA synthetases

Acceptor stem recognition by EcMetRS and EcCysRS. (A) The tRNA-bound structure of A. aeolicus MetRS, showing the CP1 domain of MetRS in green, the Rossmann fold in light green, and tRNA in orange (Protein Data Bank [PDB] identification 2CT8). (B) The tRNA-bound structure of EcCysRS, showing the CP1 domain in magenta, the Rossmann fold in light pink, and tRNA in orange (PDB identification 1U0B). (C) Superimposition of the tRNA-bound EcCysRS and EcMetRS by aligning the 5′ half of tRNA backbones (nucleotides 1–34), revealing the presence of the inserted α-helical motif in EcCysRS (circled in black). (D) Interaction between the α-helical motif in EcCysRS and the G1-C72 nucleotides in EctRNA^Cys. The helical motif is shown in light green, while the tRNA acceptor end is shown in light orange in the background. The residues Q158, Q162, and G166 are highlighted in sticks with carbons in green, while G1 and C72 are shown in sticks with carbons in cyan. (E) Effect of the triple mutations (Q158A, Q162A, and G166A) in EcCysRS on aminoacylation of tRNA. The activity is shown in logarithmic scale, while the 1-72 base pairs of the tRNA are indicated at the bottom of the bar graph. (WT) Wild-type EcCysRS; (TM) triple mutant harboring the Q158A, Q162A, and G166A mutations. Graphs showing the structures are generated by PyMol (Delano Scientific).