Where Electrostatics Matter: Bacterial Surface Neutralization and Membrane Disruption by Antimicrobial Peptides SAAP-148 and OP-145

The need for alternative treatment of multi-drug-resistant bacteria led to the increased design of antimicrobial peptides (AMPs). AMPs exhibit a broad antimicrobial spectrum without a distinct preference for a specific species. Thus, their mechanism, disruption of fundamental barrier function by permeabilization of the bacterial cytoplasmic membrane is considered to be rather general and less likely related to antimicrobial resistance. Of all physico-chemical properties of AMPs, their positive charge seems to be crucial for their interaction with negatively charged bacterial membranes. Therefore, we elucidate the role of electrostatic interaction on bacterial surface neutralization and on membrane disruption potential of two potent antimicrobial peptides, namely, OP-145 and SAAP-148. Experiments were performed on <i>Escherichia coli</i>, a Gram-negative bacterium, and <i>Enterococcus hirae</i>, a Gram-positive bacterium, as well as on their model membranes. Zeta potential measurements demonstrated that both peptides neutralized the surface charge of <i>E. coli</i> immediately after their exposure, but not of <i>E. hirae</i>. Second, peptides neutralized all model membranes, but failed to efficiently disrupt model membranes mimicking Gram-negative bacteria. This was further confirmed by flow cytometry showing reduced membrane permeability for SAAP-148 and the lack of OP-145 to permeabilize the <i>E. coli</i> membrane. As neutralization of <i>E. coli</i> surface charges was achieved before the cells were killed, we conclude that electrostatic forces are more important for actions on the surface of Gram-negative bacteria than on their cytoplasmic membranes.