Molecular Dynamics Simulations Unveil the Aggregation Patterns and Salting out of Polyarginines at Zwitterionic POPC Bilayers in Solutions of Various Ionic Strengths

This study employs molecular dynamics (MD) simulations to investigate the adsorption and aggregation behavior of simple polyarginine cell-penetrating peptides (CPPs), specifically modeled as R ₉ peptides, at zwitterionic phosphocholine POPC membranes under varying ionic strengths of two peptide concentrations and two concentrations of NaCl and CaCl ₂ . The results reveal an intriguing phenomenon of R ₉ aggregation at the membrane, which is dependent on the ionic strength indicating a salting-out effect. As the peptide concentration and ionic strength increase, peptide aggregation also increases, with aggregate lifetimes and sizes showing a corresponding rise, accompanied by the total decrease of adsorbed peptides at the membrane surface. Notably, in high ionic strength environments, large R ₉ aggregates, such as octamers, are also observed occasionally. The salting-out, typically uncommon for short positively charged peptides, is attributed to the unique properties of arginine amino acid, specifically by its side chain containing amphiphilic guanidinium (Gdm ⁺ ) ion which makes both intermolecular hydrophobic like-charge Gdm ⁺ – Gdm ⁺ and salt-bridge Gdm+ – C-terminus interactions, where the former are increased with the ionic strength, and the latter decreased due to electrostatic screening. The aggregation behavior of R ₉ peptides at membranes can also linked to their CPP translocation properties, suggesting that aggregation may aid in translocation across cellular membranes.