title: Identification of Chemical Compounds That Inhibit Protein Synthesis in Pseudomonas aeruginosa.
abstract: Four inhibitory compounds were identified using a poly-uridylic acid (polyU) mRNA-directed aminoacylation/translation (A/T) protein synthesis system composed of phenylalanyl-tRNA synthetases (PheRS), ribosomes, and ribosomal factors from Pseudomonas aeruginosa in an in vitro screen of a synthetic compound library. The compounds were specific for inhibition of bacterial protein synthesis. In enzymatic assays, the compounds inhibited protein synthesis with IC50values ranging from 20 to 60 μM. Minimum inhibitory concentrations (MICs) were determined in cultures for a panel of pathogenic organisms, including Enterococcus faecalis, Escherichia coli, Haemophilus influenzae, P. aeruginosa, Staphylococcus aureus, and Streptococcus pneumoniae. All the compounds were observed to have broad-spectrum activity and inhibited an efflux pump mutant strain of P. aeruginosa with MICs of 0.5-16 μg/mL. The molecular target of two compounds was determined to be PheRS. These two compounds were bacteriostatic against both Gram-positive and Gram-negative pathogens. In competition assays, they were not observed to compete with the natural substrates ATP or phenylalanine for active site binding. The other two compounds directly inhibited the ribosome and were bactericidal against both Gram-positive and Gram-negative pathogens. In cytotoxicity MTT testing in human cell lines, the compounds were shown to be from 2500- to 30,000-fold less active than the control staurosporine.
authors.affiliation: 1 Chemistry Department, The University of Texas-RGV, Edinburg, TX, USA.
authors.forename: Stephanie O
authors.initials: SO
authors.lastname: Palmer
authors.affiliation: 1 Chemistry Department, The University of Texas-RGV, Edinburg, TX, USA.
authors.forename: Yanmei
authors.initials: Y
authors.lastname: Hu
authors.affiliation: 2 Biology Department, The University of Texas-RGV, Edinburg, TX, USA.
authors.forename: Megan
authors.initials: M
authors.lastname: Keniry
authors.affiliation: 1 Chemistry Department, The University of Texas-RGV, Edinburg, TX, USA.
authors.forename: James M
authors.initials: JM
authors.lastname: Bullard
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D055808:Drug Discovery / Q000379:methods; D006801:Humans; D020128:Inhibitory Concentration 50; D008826:Microbial Sensitivity Tests; D015394:Molecular Structure; D014176:Protein Biosynthesis / Q000187:drug effects*; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics*; D054852:Small Molecule Libraries
chemical_list: D000900:Anti-Bacterial Agents; D054852:Small Molecule Libraries
keywords: Pseudomonas; antibiotics; drug discovery; in vitro screening; protein synthesis; tRNA aminoacylation