Crystal structure of Staphylococcus aureus tyrosyl-tRNA synthetase in complex with a class of potent and specific inhibitors

Abstract

SB-219383 and its analogues are a class of potent and specific inhibitors of bacterial tyrosyl-tRNA synthetases. Crystal structures of these inhibitors have been solved in complex with the tyrosyl-tRNA synthetase from Staphylococcus aureus, the bacterium that is largely responsible for hospital-acquired infections. The full-length enzyme yielded crystals that diffracted to 2.8 A resolution, but a truncated version of the enzyme allowed the resolution to be extended to 2.2 A. These inhibitors not only occupy the known substrate binding sites in unique ways, but also reveal a butyl binding pocket. It was reported that the Bacillus stearothermophilus TyrRS T51P mutant has much increased catalytic activity. The S. aureus enzyme happens to have a proline at position 51. Therefore, our structures may contribute to the understanding of the catalytic mechanism and provide the structural basis for designing novel antimicrobial agents.

Fig. 1.

Chemical structures of the S. aureus tyrosyl-tRNA synthetase (YRS) inhibitors. The IC₅₀ values shown in this figure are cited from published reports, which were resolved by a full aminoacylation assay (Brown et al. 1999).

Fig. 2.

Structure of the YRS monomer. (A) Alignment of the YRS sequence (Sa, BAB42818) with that of bsTyrRS (Bs, P00952, 61% identity). C-terminal domains are omitted. The bsTyrRS numbering system is adopted for YRS. Conserved residues are in bold letters. Residues interacting with tyrosyl-adenylate are shown in red, and those of the class defining ms in blue. Helices are shaded in yellow; strands are shaded in light grey. Secondary structure elements of YRS are derived from the YRS^tr485 structure and named according to those of bsTyrRS. (B) Ribbon diagram of the YRS^tr485 structure. The N-terminal domain is in yellow (helices) and red (strands), the linker domain in blue and the α-helical domain in green. The inhibitor SB-284485 is shown in a black ball-and-stick model. All helices are labeled and the strands are in the order of A-F-E-B-C-D from the front to the back of the view.

Fig. 2.

Structure of the YRS monomer. (A) Alignment of the YRS sequence (Sa, BAB42818) with that of bsTyrRS (Bs, P00952, 61% identity). C-terminal domains are omitted. The bsTyrRS numbering system is adopted for YRS. Conserved residues are in bold letters. Residues interacting with tyrosyl-adenylate are shown in red, and those of the class defining ms in blue. Helices are shaded in yellow; strands are shaded in light grey. Secondary structure elements of YRS are derived from the YRS^tr485 structure and named according to those of bsTyrRS. (B) Ribbon diagram of the YRS^tr485 structure. The N-terminal domain is in yellow (helices) and red (strands), the linker domain in blue and the α-helical domain in green. The inhibitor SB-284485 is shown in a black ball-and-stick model. All helices are labeled and the strands are in the order of A-F-E-B-C-D from the front to the back of the view.

Fig. 3.

Stereoview of the differences in the vicinity of the T51P mutation. Carbon atoms of YRS are in yellow, those of bsTyrRS in grey, and those of tyrosyl-adenylate (as observed in its complex with bsTyrRS) in purple. Colors of the other atoms are: oxygen–red, nitrogen–cyan, sulfur/phosphorous–green. The tyrosyl group of the tyrosyl-adenylate is omitted in this figure for clarity.

Fig. 4.

Stereoview of the interactions involving the bicyclic ring of SB-219383. The inhibitor carbon atoms are drawn in purple and the YRS carbon atoms are shown in yellow. Colors of the other atoms are: oxygen–red, nitrogen–cyan, sulfur/phosphorous–green. Hydrogen bonds are illustrated by dashed lines.

Fig. 5.

The superposition of SB-239629 and SB-219383 in their complexes with YRS. The former inhibitor is shown in yellow, while the latter is in purple. This view is similar to that of Figure 4 ▶.

Fig. 6.

Stereoview of the SB-243545 butyl binding pocket in YRS. Protein carbon atoms in the YRS545 structure are shown in yellow, while those of SB-243545 are shown in purple. The carbon atoms in the YRS629 structure are drawn in thinner grey lines. Oxygen is red and nitrogen is cyan.

Fig. 7.

The binding of SB-284485 to YRS. (A) Schematic diagram showing all the hydrogen bonding interactions (dashed lines) between the inhibitor and YRS. The hydrogen bonding distances are labeled. (B) Stereoview of the fucose binding mode. Hydrogen bonds are shown in dashed lines. SB-243545 is included for comparison. Protein carbons are in yellow, SB-284485 carbons are in purple, and SB-243545 carbons are in grey. Oxygen is red and nitrogen is cyan. (C) Stereoview of the superposition of SB-284485 (purple) and tyrosyl adenylate (green). The latter is shown with ribose hydrogen bonds and is modeled based on bsTyrRS structures.

Fig. 7.

The binding of SB-284485 to YRS. (A) Schematic diagram showing all the hydrogen bonding interactions (dashed lines) between the inhibitor and YRS. The hydrogen bonding distances are labeled. (B) Stereoview of the fucose binding mode. Hydrogen bonds are shown in dashed lines. SB-243545 is included for comparison. Protein carbons are in yellow, SB-284485 carbons are in purple, and SB-243545 carbons are in grey. Oxygen is red and nitrogen is cyan. (C) Stereoview of the superposition of SB-284485 (purple) and tyrosyl adenylate (green). The latter is shown with ribose hydrogen bonds and is modeled based on bsTyrRS structures.

Fig. 7.

The binding of SB-284485 to YRS. (A) Schematic diagram showing all the hydrogen bonding interactions (dashed lines) between the inhibitor and YRS. The hydrogen bonding distances are labeled. (B) Stereoview of the fucose binding mode. Hydrogen bonds are shown in dashed lines. SB-243545 is included for comparison. Protein carbons are in yellow, SB-284485 carbons are in purple, and SB-243545 carbons are in grey. Oxygen is red and nitrogen is cyan. (C) Stereoview of the superposition of SB-284485 (purple) and tyrosyl adenylate (green). The latter is shown with ribose hydrogen bonds and is modeled based on bsTyrRS structures.

Fig. 8.

Stereoviews of the Electron Density (2Fo–Fc) Maps for the YRS Inhibitors. (A) The density for SB-219383 contoured at 1 σ. (B) The density of SB239629 contoured at 1 σ. (C) The density for SB-243545 contoured at 1 σ. (D) The density for SB-284485 contoured at 1.5 σ.

Fig. 8.

Stereoviews of the Electron Density (2Fo–Fc) Maps for the YRS Inhibitors. (A) The density for SB-219383 contoured at 1 σ. (B) The density of SB239629 contoured at 1 σ. (C) The density for SB-243545 contoured at 1 σ. (D) The density for SB-284485 contoured at 1.5 σ.

Fig. 8.

Stereoviews of the Electron Density (2Fo–Fc) Maps for the YRS Inhibitors. (A) The density for SB-219383 contoured at 1 σ. (B) The density of SB239629 contoured at 1 σ. (C) The density for SB-243545 contoured at 1 σ. (D) The density for SB-284485 contoured at 1.5 σ.

Fig. 8.

Stereoviews of the Electron Density (2Fo–Fc) Maps for the YRS Inhibitors. (A) The density for SB-219383 contoured at 1 σ. (B) The density of SB239629 contoured at 1 σ. (C) The density for SB-243545 contoured at 1 σ. (D) The density for SB-284485 contoured at 1.5 σ.