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Updated: May 5, 2026
Establishment of a High-throughput Setup for Screening Small Molecules That Modulate c-di-GMP Signaling in Pseudomonas aeruginosa
Published on: June 30, 2016
Discovery of a Pseudomonas aeruginosa-specific small molecule targeting outer membrane protein OprH-LPS interaction
Bradley E Poulsen1, Thulasi Warrier1, Sulyman Barkho2
1Department of Molecular Biology and Center for Computational and Integrative Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Abstract:
The surge of antimicrobial resistance threatens efficacy of current antibiotics, particularly against Pseudomonas aeruginosa, a highly resistant gram-negative pathogen. The asymmetric outer membrane (OM) of P. aeruginosa combined with its array of efflux pumps provide a barrier to xenobiotic accumulation, thus making antibiotic discovery challenging. We adapted PROSPECT, a target-based, whole-cell screening strategy, to discover small molecule probes that kill P. aeruginosa mutants depleted for essential proteins localized at the OM. We identified BRD1401, a small molecule that has specific activity against a P. aeruginosa mutant depleted for the essential lipoprotein, OprL. Genetic and chemical biological studies identified that BRD1401 acts by targeting the OM β-barrel protein OprH to disrupt its interaction with LPS and increase membrane fluidity. Studies with BRD1401 also revealed an interaction between OprL and OprH, directly linking the OM with peptidoglycan. Thus, a whole-cell, multiplexed screen can identify species-specific chemical probes to reveal pathogen biology.
Insights
Researchers discovered a new compound, BRD1401, that effectively targets the outer membrane of Pseudomonas aeruginosa. This finding offers a novel approach to combatting antimicrobial resistance in this challenging pathogen.
Area of Science:
- Microbiology
- Drug Discovery
- Biochemistry
Background:
- Antimicrobial resistance is a growing global health threat, particularly against Gram-negative bacteria like Pseudomonas aeruginosa.
- The unique outer membrane and efflux pumps of P. aeruginosa present significant challenges for antibiotic penetration and efficacy.
- Discovering new antibiotics against P. aeruginosa requires innovative strategies that overcome its inherent resistance mechanisms.
Purpose of the Study:
- To adapt a target-based, whole-cell screening strategy (PROSPECT) for identifying novel antimicrobial compounds against P. aeruginosa.
- To discover small molecule probes that specifically target essential proteins in the P. aeruginosa outer membrane.
- To elucidate the mechanism of action of newly identified compounds and their impact on bacterial cell biology.
Main Methods:
- Utilized the PROSPECT screening strategy to identify compounds targeting P. aeruginosa mutants lacking essential outer membrane proteins.
- Conducted genetic and chemical biological studies to determine the molecular targets and mechanisms of action of identified compounds.
- Investigated the interaction between outer membrane proteins and lipopolysaccharide (LPS) to understand membrane dynamics.
Main Results:
- Identified BRD1401, a small molecule with specific activity against P. aeruginosa mutants deficient in the essential lipoprotein OprL.
- Determined that BRD1401 targets the outer membrane β-barrel protein OprH, disrupting its interaction with LPS and increasing membrane fluidity.
- Revealed a novel interaction between OprL and OprH, establishing a link between the outer membrane and the peptidoglycan layer.
Conclusions:
- Whole-cell, multiplexed screening is an effective approach for discovering species-specific chemical probes against challenging pathogens.
- BRD1401 represents a promising lead compound for developing new therapeutics against P. aeruginosa by targeting outer membrane integrity.
- The study provides new insights into the complex biology of the P. aeruginosa outer membrane and its essential protein interactions.
