Young Investigator Highlight

Chloe Mitchell from The Hospital for Sick Children is one of the winners of the 2022 Schram Young Investigators Oral Presentations Award, presented at this year's American Peptide Society Symposium in Whistler, B.C., Canada. The title of her talk was Peptide-Based Disruption of Membrane-Embedded Protein-Protein Interactions in Bacterial Efflux Pumps.

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Antibiotic resistance is a global crisis that first appeared in 1940 shortly after the introduction of penicillin and has been a steady battle ever since. With the advent of each new antibiotic, bacteria compensate with mechanisms to avoid their lethal effects and continue to proliferate in our health care systems. Dr. Charles Deber’s lab studies one of these bacterial defense mechanisms – drug efflux pumps. These pumps lie in the bacterial membrane and are capable of recognizing the antibiotics we administer and removing them from the bacterial cell.

The Deber lab studies principally the small multidrug resistance (SMR) efflux protein. SMR’s homodimerize through their 4th transmembrane helix (TM4) to function and are primarily responsible for expelling detergents and disinfectants from the cells. We have employed a rational drug design to target the TM4-TM4 interaction by designing peptide inhibitors that contain the TM4 Gly-Gly heptad motif that mediates dimerization. The peptide inhibitors also contain two tags: a C-terminal positively-charged Lys tag to direct the peptide to the negatively-charged bacterial membrane, and a non-charged N-terminal peptoid tag to promote membrane insertion. These peptides should therefore insert into the membrane, align with - and competitively disrupt - the membrane-embedded TM4-TM4 interface, and thereby disable substrate efflux.

To date, studies have been performed on the SMR from Pseudomonas aeruginosa, the bacterium that infects patients with cystic fibrosis. Cystic fibrosis is a genetic disease that leads to a heavy mucus layer in the lungs, providing a moist, rich environment for bacteria to flourish. This particular species has become extremely resistant and difficult to treat. We have earlier shown that these peptides reduce SMR-mediated efflux activity, ostensibly through the specifically designed mechanism. However, the architecture of these peptides resembles that of cationic antimicrobial peptides, as both categories of peptides have a hydrophobic and charged domain. Therefore, we’ve explored in structural detail the effects inhibitor peptides have on the bacterial membrane and found the peptides to be stably inserting and causing minimal non-specific membrane disruption. Our results show promise for targeting these bacterial efflux pumps, as well as representing a new approach to disrupting membrane-embedded protein-protein interactions.

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