title: Recombinant production, crystallization and X-ray crystallographic structure determination of the peptidyl-tRNA hydrolase of Pseudomonas aeruginosa.
abstract: The peptidyl-tRNA hydrolase enzyme from the pathogenic bacterium Pseudomonas aeruginosa (Pth; EC 3.1.1.29) has been cloned, expressed in Escherichia coli and crystallized for X-ray structural analysis. Suitable crystals were grown using the sitting-drop vapour-diffusion method after one week of incubation against a reservoir solution consisting of 20% polyethylene glycol 4000, 100 mM Tris pH 7.5, 10%(v/v) isopropyl alcohol. The crystals were used to obtain the three-dimensional structure of the native protein at 1.77 Å resolution. The structure was determined by molecular replacement of the crystallographic data processed in space group P6(1)22 with unit-cell parameters a=b=63.62, c=155.20 Å, α=β=90, γ=120°. The asymmetric unit of the crystallographic lattice was composed of a single copy of the enzyme molecule with a 43% solvent fraction, corresponding to a Matthews coefficient of 2.43 Å3 Da(-1). The crystallographic structure reported here will serve as the foundation for future structure-guided efforts towards the development of novel small-molecule inhibitors specific to bacterial Pths.
authors.affiliation: Foresight Biosciences Inc., HudsonAlpha Institute for Biotechnology, 601 Genome Way, Huntsville, AL 35806, USA.
authors.forename: Ronny C
authors.initials: RC
authors.lastname: Hughes
authors.forename: Hana
authors.initials: H
authors.lastname: McFeeters
authors.forename: Leighton
authors.initials: L
authors.lastname: Coates
authors.forename: Robert L
authors.initials: RL
authors.lastname: McFeeters
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000096:biosynthesis* / Q000737:chemistry*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000201:enzymology*
chemical_list: D002265:Carboxylic Ester Hydrolases; C019674:aminoacyl-tRNA hydrolase