title: Mutant Alleles of lptD Increase the Permeability of Pseudomonas aeruginosa and Define Determinants of Intrinsic Resistance to Antibiotics.
abstract: Gram-negative bacteria provide a particular challenge to antibacterial drug discovery due to their cell envelope structure. Compound entry is impeded by the lipopolysaccharide (LPS) of the outer membrane (OM), and those molecules that overcome this barrier are often expelled by multidrug efflux pumps. Understanding how efflux and permeability affect the ability of a compound to reach its target is paramount to translating in vitro biochemical potency to cellular bioactivity. Herein, a suite of Pseudomonas aeruginosa strains were constructed in either a wild-type or efflux-null background in which mutations were engineered in LptD, the final protein involved in LPS transport to the OM. These mutants were demonstrated to be defective in LPS transport, resulting in compromised barrier function. Using isogenic strain sets harboring these newly created alleles, we were able to define the contributions of permeability and efflux to the intrinsic resistance of P. aeruginosa to a variety of antibiotics. These strains will be useful in the design and optimization of future antibiotics against Gram-negative pathogens.
authors.affiliation: Merck Research Laboratories, Kenilworth, New Jersey, USA carl.balibar@merck.com.
authors.forename: Carl J
authors.initials: CJ
authors.lastname: Balibar
authors.affiliation: Department of Molecular Biology, Princeton University, Princeton, New Jersey, USA.
authors.forename: Marcin
authors.initials: M
authors.lastname: Grabowicz
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D001692:Biological Transport / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics*; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000235:genetics*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D008070:Lipopolysaccharides / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics*
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D008070:Lipopolysaccharides; D026901:Membrane Transport Proteins