Identification of Antibacterial Cyclic Peptides with a High-Throughput Cell-Based Dropout Screen

Area of Science:

• Microbiology
• Drug Discovery
• Genomics

Background:

• Antimicrobial resistance poses a significant global health challenge, driving the urgent need for novel antibiotics.
• Existing antibiotic discovery strategies face limitations in identifying new targets and effective compounds.

Purpose of the Study:

• To establish a target-agnostic screening platform for identifying antibacterial cyclic peptides.
• To discover novel antibacterial agents and intracellular targets using a negative-selection approach.
• To validate the platform by identifying a new cyclic peptide scaffold against an essential bacterial protein.

Main Methods:

• Developed an intracellular screening platform utilizing a library of 3.2 million cyclic peptides.
• Employed next-generation sequencing to identify sequences depleted by antibacterial activity.
• Clustered depleted sequences by shared pharmacophores for robust hit identification.
• Focused library design based on identified pharmacophores.

Main Results:

• Successfully identified antibacterial cyclic peptides through sequence depletion.
• Discovered a novel cyclic peptide scaffold with antibacterial properties.
• Demonstrated inhibition of the essential iron-sulfur cluster carrier protein ErpA, a previously untargeted protein.
• Validated the platform's ability to identify inhibitors of essential intracellular targets.

Conclusions:

• The developed platform offers a general and scalable framework for discovering intracellular antibacterial targets.
• This approach enables the identification of novel antibacterial cyclic peptides and their corresponding targets directly within living cells.
• The findings provide a new strategy for combating antimicrobial resistance by targeting essential bacterial proteins like ErpA.