Peptide-induced hydration of lipid bilayers modulates packing pattern and conformations of hydrocarbon chains - a potential pathway for peptide translocation?

Cell-penetrating peptides (CPPs) with a cationic-hydrophobic character are recognized as carriers for delivering various therapeutics and diagnostic agents across cell membranes and into the cells. Among the most studied CPPs, nona-arginine (R9) exhibits superior penetration compared to nona-lysine (K9), suggesting that the penetration ability depends not only on charge, distribution and concentration of peptides but also on the lipid membrane composition. However, for heptapeptides composed of arginine (R), lysine (K) and phenylalanine (F) residues, which show some CPPs properties, these interactions remain unexplored. This study sheds light on the adsorption of R5F2/K5F2 on model prokaryotic (PRO) and eukaryotic (EU) lipid membranes containing a zwitterionic lipid (phosphatidylcholine; PC) and an anionic lipid (either phosphatidylglycerol, PG, in the PRO model, or phosphatidylserine, PS in EU) at the 90:10 molar ratio. Using differential scanning calorimetry (DSC) and temperature-dependent UV-Vis spectroscopy, we observed peptide-induced changes in the interfacial water layer that affect the fluidity and rigidity of lipid bilayers. The distinct adsorption behavior of R5F2/K5F2 on PRO and EU lipid bilayers revealed the changes in lipid packing and hydrocarbon chain conformations as exclusively peptide-dependent features. The peptide-induced formation of vacancies in the non-polar bilayer part is consistent with partial leakage observed in giant unilamellar vesicles. The synchronized arrangement could represent a mechanism for the concerted translocation of CPPs, along with their potential cargo across the lipid membrane. This study provides new insights into the peptide-lipid interactions underlying CPPs functionality.