Structural and Thermodynamic Insight into Spontaneous Membrane-Translocating Peptides Across Model PC/PG Lipid Bilayers.

We present results of Martini coarse-grained force field simulations to estimate the potentials of mean force for a series of recently screened spontaneous membrane-translocating peptides, SMTPs. We consider model bilayer composed of POPC and POPG, the latter providing the anionic component as used in experimental studies. We observe a significant barrier for translocation in the case of the canonical cationic cell-penetrating peptide nona-arginine, ARG9. In the case of the TP1, TP2, and TP3 peptides, potentials of mean force are systematically lower relative to the ARG9 case. Though the barriers predicted by the simulations, on the order of 20 kcal/mol, are still rather large to recapitulate the experimental kinetics of internalization, we emphasize that the qualitative trend of reduction of barrier heights is a significant result. Decomposition of the PMFs indicates that though there is a substantial entropic stability when the peptides reside at bilayer center, barriers as predicted from these force field-based studies are largely determined by enthalpic (potential energy) interactions. We note that the binding of the SMTPs is critically dependent on the mix of hydrophilic and hydrophobic residues that constitute the amino acid motif/sequence of these peptides. For the cationic ARG9 which only contains hydrophilic residues, there is no tight binding observed. The specific motif [Formula: see text] (where [Formula: see text] is a general residue) is a potential sequence in drug/peptide design. The SMTPs with this motif are able to translocate into membrane at a significantly lower free energy cost, compared to the negative control peptides. Finally, we compare the different membrane perturbations induced by the presence of the different peptides in the bilayer center. In some cases, hydrophilic pores are observed to form, thus conferring stability to the internalized state. In other cases, SMTPs are associated only with membrane defects such as induced membrane curvature. These latter observations suggest some influence of membrane rigidity as embodied in the full range of membrane undulatory modes in defining pore-forming propensities in bilayers.