# Selective Interaction of the Antimicrobial Peptide RKW with Bacterial Lipid Bilayers: A Biophysical Approach

## TL;DR

This study explores how an antimicrobial peptide called RKW selectively targets bacterial membranes without harming eukaryotic cells, using biophysical methods to understand its mechanism and safety.

## Contribution

The novel contribution is the detailed biophysical characterization of RKW's selective interaction with bacterial lipid bilayers and its safety in a living model.

## Key findings

- RKW preferentially interacts with negatively charged bacterial membranes, especially Gram-negative ones.
- RKW induces membrane permeabilization and leakage in bacterial model membranes.
- Toxicological tests in C. elegans showed no adverse effects, supporting RKW's safety.

## Abstract

Antimicrobial peptides (AMPs) have emerged as promising
candidates
for next-generation antibiotics due to their broad-spectrum activity,
including efficacy against multidrug-resistant bacteria. However,
their clinical application remains limited, primarily because of cytotoxicity
toward host cells. A deeper understanding of AMP–membrane interactions,
particularly through biophysical studies using model membrane systems,
is essential for developing safe and effective AMP-based therapeutics.
In this study, the interaction of a previously designed AMP, named
RKW, with model lipid vesicles mimicking the lipid composition of
both prokaryotic and eukaryotic cell membranes was investigated. RKW
exhibited a strong preference for negatively charged bacterial membrane
models, especially those representing Gram-negative bacteria, while
showing minimal or no affinity for zwitterionic or eukaryotic-like
membranes. These findings imply that electrostatic interactions are
the primary driving force behind its membrane selectivity. Fluorescence
spectroscopy and quenching experiments with acrylamide and lipophilic
probes revealed that RKW localizes mainly at the membrane interface,
likely adopting a parallel orientation relative to the bilayer surface.
Furthermore, RKW induced substantial leakage of carboxyfluorescein
from bacterial model membranes, indicating potent membrane permeabilisation.
This mechanism was corroborated by dynamic light scattering (DLS)
analyses, which provided additional evidence of peptide-induced membrane
disruption. Collectively, this study elucidates the selective mechanism
of action of RKW and underlines its potential as a targeted antimicrobial
agent with reduced cytotoxicity toward eukaryotic cells. Toxicological
assessments using the Caenorhabditis elegans in vivo model further supported its safety, showing no adverse effects
on survival, reproduction, locomotion, or growth.