How arginine derivatives alter the stability of lipid membranes: dissecting the roles of side chains, backbone and termini

Abstract

Arginine (R)-rich peptides constitute the most relevant class of cell-penetrating peptides and other membrane-active peptides that can translocate across the cell membrane or generate defects in lipid bilayers such as water-filled pores. The mode of action of R-rich peptides remains a topic of controversy, mainly because a quantitative and energetic understanding of arginine effects on membrane stability is lacking. Here, we explore the ability of several oligo-arginines R_n and of an arginine side chain mimic R_mathrm {Side} to induce pore formation in lipid bilayers employing MD simulations, free-energy calculations, breakthrough force spectroscopy and leakage assays. Our experiments reveal that R_mathrm {Side} but not R_n reduces the line tension of a membrane with anionic lipids. While R_n peptides form a layer on top of a partly negatively charged lipid bilayer, R_mathrm {Side} leads to its disintegration. Complementary, our simulations show R_mathrm {Side} causes membrane thinning and area per lipid increase beside lowering the pore nucleation free energy. Model polyarginine R_8 similarly promoted pore formation in simulations, but without overall bilayer destabilization. We conclude that while the guanidine moiety is intrinsically membrane-disruptive, poly-arginines favor pore formation in negatively charged membranes via a different mechanism. Pore formation by R-rich peptides seems to be counteracted by lipids with PC headgroups. We found that long R_n and R_mathrm {Side} but not short R_n reduce the free energy of nucleating a pore. In short R_n, the substantial effect of the charged termini prevent their membrane activity, rationalizing why only longer mathrm {R}_{n} are membrane-active.