Membrane and lipopolysaccharide interactions of C-terminal peptides from S1 peptidases

The mechanisms underlying antimicrobial and anti-endotoxic effects were investigated for a series of structurally related peptides derived from the C-terminal region of S1 peptidases. For this purpose, results on bacterial killing were compared to those on peptide-induced liposome leakage, and to ellipsometry and dual polarization interferometry results on peptide binding to, and disordering of, supported lipid bilayers. Furthermore, the ability of these peptides to block endotoxic effects caused by bacterial lipopolysaccharide (LPS), monitored through NO production in macrophages, was compared to the binding of these peptides to LPS, and to secondary structure formation in the peptide/LPS complex. Bacteria killing, occurring through peptide-induced membrane lysis, was found to correlate with liposome rupture, and with the extent of peptide binding to the lipid membrane, no adsorption threshold for peptide insertion being observed. Membrane and LPS binding was found to depend on peptide net charge, illustrated by LPS binding increasing with increasing peptide charge, and peptides with net negative charge being unable to lyse membranes, kill bacteria, and block LPS-induced endotoxic effect. These effects were, however, also influenced by peptide hydrophobicity. LPS binding was furthermore demonstrated to be necessary, but not sufficient, for anti-endotoxic effect of these peptides. Circular dichroism spectroscopy showed that pronounced helix formation occurs in peptide/LPS complexes for all peptides displaying anti-endotoxic effect, hence potentially linked to this functionality. Similarly, ordered secondary structure formation was correlated to membrane binding, lysis, and antimicrobial activity of these peptides. Finally, preferential binding of these peptides to LPS over the lipid membrane was demonstrated.

Graphical abstractBacterial membranes present multiple binding sites for C-terminal peptides derived from S1 peptidases. Apart from interacting with phospholipid membranes, thereby causing membrane rupture and resulting in antimicrobial effects, they bind to bacterial lipopolysaccharides (LPS), a necessary but not sufficient criterium for their anti-endotoxic effect. LPS and membrane binding is promoted by electrostatic and hydrophobic interactions. Membrane destabilization and antimicrobial effect, as well as anti-endotoxic effect, is correlated to helix formation in the presence of lipid membranes and LPS, respectively.Figure optionsDownload full-size imageDownload high-quality image (170 K)Download as PowerPoint slideHighlights► S1 peptide interaction with lipid membrane and bacterial lipopolysaccharide (LPS) investigated. ► Membrane rupture increases with peptide adsorption and insertion. ► LPS/membrane binding depends on peptide charge, hydrophobicity, and helix content. ► Preferential binding of S1 peptides to LPS over lipid membranes. ► Helix formation important for both membrane rupture and anti-endotoxic effect.