Combining Cell Envelope Stress Reporter Assays in a Screening Approach to Identify BAM Complex Inhibitors

The development of new antibiotics is particularly problematic in Gram-negative bacteria due to the presence of the outer membrane (OM), which serves as a permeability barrier. Recently, the beta-barrel assembly machine (BAM), located in the OM and responsible for beta-barrel type OM protein (OMP) assembly, has been validated as a novel target for antibiotics. Here, we identified potential BAM complex inhibitors using a screening approach that reports on cell envelope sigma(E) and Rcs stress in Escherichia coli. Screening a library consisting of 316 953 compounds yielded five compounds that induced sigma(E) and Rcs stress responses, while not inducing the intracellular heat-shock response. Two of the five compounds (compounds 2 and 14) showed the characteristics of known BAM complex inhibitors: synergy with OMP biogenesis mutants, decrease in the abundance of various OMPs, and loss of OM integrity. Importantly, compound 2 also inhibited BAM-dependent OMP folding in an in vitro refolding assay using purified BAM complex reconstituted in proteoliposomes.

Automated summary

Researchers identified potential BAM complex inhibitors as novel targets for antibiotics, which induced cell envelope sigma(E) and Rcs stress in bacteria. Two of the compounds showed characteristics of known BAM complex inhibitors, including reducing the abundance of OMPs and disrupting OM integrity.