In Silico Studies of Asymmetric Membranes Perturbations Caused by Dynamic Aggregation of a Cell-Penetrating Peptide

Abstract

Membrane active peptides are therapeutically relevant for a variety of purposes. However a better understanding of their mechanisms of interaction with lipid bilayers is needed in order to maximise both efficiency and selectivity. In the case of cell-penetrating peptides (CPP) it is particularly important to avoid membrane toxicity while maintaining translocation across the plasma membrane. Experimental in vitro studies based on light microscopy and dye release have shown CPP can be internalised via both endocytic and energy-independent pathways. However, uncertainties remain concerning the mechanisms involved in membrane translocation and perturbation. In silico studies using molecular dynamics (MD) simulations have hitherto mainly focused on the interactions of peptides with relatively simple lipid bilayer models.Here we present coarse-grained simulations of the interactions between transportan, a CPP known to perturb cell membranes, and large bilayers with biologically relevant lipid composition. We observe that transportan forms dynamic, unstructured and transient clusters that catalyse the formation of local defects such as bilayer thinning, lipid redistribution and decrease of the lipid tail order. We present a novel analytic approach which shows that the extent of the membrane perturbations induced by CPP clusters depends on their size and varies in time. In particular, anionic lipid flip-flops are consistently observed above a certain cluster size. The importance of asymmetric lipid composition in the bilayer is also investigated and found to impact the stability of the peptide aggregates. We also apply our approach to extended bilayer systems that contain approximately 50,000 lipids and hundreds of transportan peptides and thus allow comparison with high-resolution light microscopy results.