A Screening Platform for Identification of Compounds That Inhibit Protein Synthesis in Pseudomonas Aeruginosa

Pseudomonas aeruginosa is a Gram-negative pathogenic bacteria and a common cause of nosocomial infections. This pathogen has recently garnered attention due to its pan-resistance. We have developed a poly(U) mRNA-directed aminoacylation/translation (A/T) protein synthesis system from P. aeruginosa. The concentration of each component of the system has been arrived at through multiple rounds of optimization. Poly-phenylalanine synthesis in the system was monitored using scintillation proximity assays (SPA). This system has been used to screen two natural compound libraries (\u3e1100) and a number of the compounds that inhibit greater than 50% of the activity of the system have been identified. The molecular targets for the hit compounds were determined. Minimum inhibitory concentrations (MIC) were determined per NCCLS guidelines. Three of the compounds were tested in time-kill assays to determine their mode of inhibition and tested for their ability to inhibit eukaryotic cytosolic and mitochondrial protein synthesis and none was determined