Lipopolysaccharide interaction is decisive for the activity of the antimicrobial peptide NK-2 against Escherichia coli and Proteus mirabilis
Résumé
Phosphatidylglycerol is a widely used mimetic to study the effects of antimicrobial peptides (AMPs) on the bacterial cytoplasmic membrane. It turned out, however, that the antibacterial activities of novel NK-2-derived AMPs could not sufficiently explained by using this simple model system. Since the lipopolysaccharide (LPS) containing outer membrane is the first barrier of Gram-negative bacteria, here we investigated interactions of NK-2 and a shortened variant thereof with viable Escherichia coli WBB01 and Proteus mirabilis R45, and with model membranes composed of LPS isolated from these two strains. Differences in net charge and charge distribution of the two LPS have been made responsible for the differential sensitivity of the respective bacteria to other AMPs. As imaged by TEM and AFM, NK-2-mediated killing of these bacteria was corroborated by structural alterations of the outer and inner membranes, the release of E. coli cytoplasma, and the formation of unique fibrous structures inside P. mirabilis, suggesting distinct and novel intracellular targets. NK-2 bound to and intercalated into LPS bilayers, and eventually induced the formation of transient, heterogeneous lesions in planar lipid bilayers. However, the discriminative activity of NK-2 against the two bacterial strains was independent from membrane intercalation and lesion formation, which both were indistinguishable for the two LPS. Instead, differences in activity originated from the LPS binding step, which could be demonstrated by NK 2 attachment to intact bacteria, and to solid-supported LPS bilayers on a surface acoustic wave biosensor.
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