Computational Insights into Membrane Disruption by Cell-Penetrating Peptides

Cell-penetrating peptides (CPPs) can translocate into cells without inducing cytotoxicity. The internalization process implies several steps at different time scales ranging from microseconds to minutes. We combine adaptive Steered Molecular Dynamics (aSMD) with conventional Molecular Dynamics (cMD) to observe nonequilibrium and equilibrium states to study the early mechanisms of peptide–bilayer interaction leading to CPPs internalization. We define three membrane compositions representing bilayer sections, neutral lipids (i.e., upper leaflet), neutral lipids with cholesterol (i.e., hydrophobic core), and neutral/negatively charged lipids with cholesterol (i.e., lower leaflet) to study the energy barriers and disruption mechanisms of Arg9, MAP, and TP2, representing cationic, amphiphilic, and hydrophobic CPPs, respectively. Cholesterol and negatively charged lipids increase the energetic barriers for the peptide–bilayer crossing. TP2 interacts with the bilayer by hydrophobic insertion, while Arg9 disrupts the bilayer by forming transient or stable pores. MAP has shown both behaviors. Collectively, these findings underscore the significance of innovative computational approaches in studying membrane-disruptive peptides and, more specifically, in harnessing their potential for cell penetration.