Recombinant production, crystallization and X-ray crystallographic structure determination of peptidyl-tRNA hydrolase from Salmonella typhimurium

Peptidyl-tRNA hydrolase (Pth; EC 3.1.1.29) from the pathogenic bacterium Salmonella typhimurium has been cloned, expressed in Escherichia coli and crystallized for X-ray analysis. Crystals were grown using hanging-drop vapor diffusion against a reservoir solution consisting of 0.03 M citric acid, 0.05 M bis-tris propane, 1% glycerol, 3% sucrose, 25% PEG 6000 pH 7.6. Crystals were used to obtain the three-dimensional structure of the native protein at 1.6 angstrom resolution. The structure was determined by molecular replacement of the crystallographic data processed in space group P2(1)2(1)2(1) with unit-cell parameters a = 62.1, b = 64.9, c = 110.5 angstrom, alpha = beta = gamma = 90 degrees. The asymmetric unit of the crystallographic lattice was composed of two copies of the enzyme molecule with a 51% solvent fraction, corresponding to a Matthews coefficient of 2.02 angstrom(3) Da(-1). The structural coordinates reported serve as a foundation for computational and structure-guided efforts towards novel small-molecule Pth1 inhibitors and potential antibacterial development.