Membrane-Active Antibiotics Affect Domains in Bacterial Membranes as the First Step of Their Activity

The need to combat antimicrobial resistance is becoming more and more pressing. Here we investigate the working mechanism of a small cationic agent, <i>N</i>-alkylamide 3d, by conventional and high-speed atomic force microscopy. We show that <i>N</i>-alkylamide 3d interacts with the membrane of <i>Staphylococcus aureus</i>, where it changes the organization and dynamics of lipid domains. After this initial step, supramolecular structures of the antimicrobial agent attach on top of the affected membrane gradually, covering it entirely. These results demonstrate that lateral domains in the bacterial membranes might be affected by small antimicrobial agents more often than anticipated. At the same time, we show a new dual-step activity of <i>N</i>-alkylamide 3d that not only destroys the lateral membrane organization but also effectively covers the whole membrane with aggregates. This final step could render the membrane inaccessible from the outside and possibly prevent signaling and waste disposal of living bacteria.