Structure of Francisella tularensis peptidyl-tRNA hydrolase

Abstract

The rational design of novel antibiotics for bacteria involves the identification of inhibitors for enzymes involved in essential biochemical pathways in cells. In this study, the cloning, expression, purification, crystallization and structure of the enzyme peptidyl-tRNA hydrolase from Francisella tularensis, the causative agent of tularemia, was performed. The structure of F. tularensis peptidyl-tRNA hydrolase is comparable to those of other bacterial peptidyl-tRNA hydrolases, with most residues in the active site conserved amongst the family. The resultant reagents, structural data and analyses provide essential information for the structure-based design of novel inhibitors for this class of proteins.

Figure 1

(a) Ribbon diagram of the secondary structure of F. tularensis peptidyl-tRNA hydrolase (FtPth), with N- and C-termini labeled. The structure comprises a monomer with a single α/β globular domain built around a twisted β-sheet. The coloring scheme for the secondary-structure elements is as follows: α-helices are colored red, β-strands yellow and loops green. Chloride ions are represented by gray spheres, while PEG and ethylene glycol molecules are shown as sticks colored by atom type. (b) Superposition with the published E. coli structure (PDB entry 2pth). This figure was generated using PyMOL (DeLano, 2002 ▶). (c) Crystal-packing diagram for the F. tularensis Pth apo structure. The C-terminus of each molecule (red ribbon) resides in the active site of a neighbouring symmetry mate (gray surface). (d) Superposition of the F. tularensis (colored red), E. coli (colored green; PDB entry 3ofv; R. Lam, T. E. McGrath, V. Romanov, S. A. Gothe, S. R. Peddi, E. Razumova, R. S. Lipman, A. A. Branstrom & N. Y. Chirgadze, unpublished work) and M. tuberculosis (colored blue; PDB entry 2z2i; Selvaraj et al., 2007 ▶) Pth crystal structures. The core secondary-structure elements and protein fold are conserved in all species. As shown by the superposition, the highly mobile C-terminal helix and adjacent hinge region represent opportunities for designing new constructs (for the identification of feasible crystal forms).