Overcoming the Transport Limitation of the Lasso Peptide Citrocin Using Antimicrobial Synergy

Abstract

Previous work in the discovery of the lasso peptide citrocin demonstrated the potent RNA polymerase inhibition of the peptide in vitro. However, corresponding antimicrobial susceptibility tests to determine the minimum inhibitory concentration against E. coli strains revealed only moderate antimicrobial activity. This work suggested that citrocin may be limited in its uptake by target cells. One goal of this work was to demonstrate that the phenomenon of antimicrobial synergy could be applied to overcome the transport limitation of citrocin. Synergy occurs when the combination of two antimicrobial agents provides a more effective killing response than if either were used alone. Checkerboard assays for synergy between citrocin and several other antibiotic compounds were completed against two different Gram-negative bacterial strains, E. coli MG1655 and A. baumannii ATCC BAA 1605. Synergy was observed between citrocin and members of the membrane disrupting polymyxin antimicrobial class of compounds in both strains. Exclusive to E. coli MG1655, however, synergy was also observed between the lasso peptide and trimethoprim, an antibiotic that inhibits the folate biosynthesis pathway, and chloramphenicol, a drug that inhibits ribosomal translation. Another goal of this work was to determine whether resistance to citrocin could be overcome by potentiating its activity with other antibiotics. E. coli ∆sbmA does not express a crucial inner membrane transport protein, SbmA, needed for citrocin uptake, making it resistant to the lasso peptide. The polymyxins, which exhibited the strongest degree of synergy against E. coli MG1655 did not enable citrocin to overcome resistance.