Increasing the permeability of Escherichia coli using MAC13243

Area of Science:

• Microbiology
• Bacterial Physiology
• Drug Discovery

Background:

• Gram-negative bacteria possess an outer membrane that acts as a barrier, limiting the entry of large molecules.
• This barrier renders many antibiotic classes ineffective against these bacteria, posing a significant challenge in treating infections.

Purpose of the Study:

• To identify small molecules capable of enhancing the permeability of the gram-negative bacterial outer membrane.
• To investigate potential therapeutic strategies to overcome antibiotic resistance in gram-negative pathogens.

Main Methods:

• Conducted a high-throughput screen to discover compounds that increase outer membrane permeability in Escherichia coli.
• Utilized a fluorescent probe (1-N-phenylnapthylamine) to assess changes in membrane permeability.
• Employed CRISPR interference (CRISPRi) to genetically validate the observed phenotype by depleting the target protein, LolA.

Main Results:

• Identified MAC13243 as a small molecule inhibitor of the periplasmic chaperone LolA.
• Demonstrated that sub-inhibitory concentrations of MAC13243 increase bacterial outer membrane permeability to fluorescent probes.
• Showed that MAC13243 treatment enhances the susceptibility of Escherichia coli to antibiotics with large scaffolds.
• Confirmed that depleting LolA using CRISPRi recapitulates the MAC13243-induced permeability phenotype, ruling out off-target effects.

Conclusions:

• MAC13243, by inhibiting LolA, effectively increases outer membrane permeability in gram-negative bacteria.
• This mechanism enhances the efficacy of large-scaffold antibiotics, offering a potential strategy to combat antibiotic resistance.
• Targeting LolA represents a promising avenue for developing novel therapeutics against gram-negative infections.