Molecular Mechanism of Polycation-Induced Pore Formation in Biomembranes

Abstract

Polycations are an attractive class of macromolecules with promising applications as drug/gene carriers and biocides. The chemical structure and concentration of a polycation determine its interaction with cellular membranes and, hence, are crucial parameters for designing efficient nontoxic polycations. However, the interaction of polycations with biomembranes at the molecular level and the corresponding free-energy landscape is not well understood. In this work, we investigate the molecular mechanism of interaction between a strong polycation substituted with alkyl moieties and zwitterionic membranes via long-time-scale allatom molecular dynamics simulations and free-energy calculations combined with Langmuir monolayer, atomic force microscopy, and calcein-release experimental measurements. We found that the membrane activity of the polycation and its ability to induce pores in the membranes can be attributed to the polycation-induced changes in the bilayer organization, such as reduced membrane thickness, increased disorder of the acyl chains, reduced packing, and electrostatic field gradients between membrane leaflets. These changes facilitate the penetration of water into the membrane and the formation of aqueous defects/pores. The calculated free-energy profiles indicate that the polycation lowers the nucleation barrier for pore opening and the free energy for pore formation in a concentration-dependent manner. Above the critical coverage of the membrane, the polycation nucleates spontaneous pores in zwitterionic membranes. Our work demonstrates the potential of combining enhanced sampling methods in MD simulations with experiments for a quantitative description of various events in the polycation-membrane interaction cycle, such as strong adsorption on the membrane due to hydrophobic and electrostatic interactions, and pore formation.

Polycation–Anionic Lipid Membrane Interactions

“…A local gradient of the electrostatic potential appears between the leaflets, which creates the potential difference between the two leaflets, referred to as interleaflet voltage. 19 However, as M20-1, M20-2, and M40-1 systems contain very few mobile counterions needed to neutralize the overall charge of the system, we suspect that the calculated potential in a periodic box might not be representative of the actual potential, which can then lead to unexpected effects. (See water ordering above and the Discussion section.)…”

“…Therefore, PMAPTAC is preferentially located in the polar region, close to the headgroups, and does not penetrate into the center of the bilayer ( Figure S4 ), in contrast to hydrophobically modified polycations. 18 , 19 , 48 …”

“…The experiments were performed in triplicate as previously described. 19 Fluorescence spectra of the released calcein were recorded at 25 °C using a PerkinElmer LSD 50B spectrofluorometer. The amount of calcein released after time t , RF( t ), was calculated according to the equation where I 0 , I t , and I max are fluorescence intensities measured for the calcein-loaded liposomes before the polymer addition, at time t after the polymer introduction, and after the Triton X-100 addition (corresponding to complete calcein release), respectively.…”

A joint reaction coordinate for computing the free energy landscape of pore nucleation and pore expansion in lipid membranes

“…Previous simulations investigated pore formation in membrane of various size, spanning systems between 64 and 512 lipids. 17,20,[24][25][26][28][29][30][47][48][49][50] However, we previously found that the system size may strongly influence the free energy landscape of pore formation. In small membrane systems, the open pore may be destabilized because there is no flat membrane between the pore and its periodic image.…”

Biosynthetic Polymalic Acid as a Delivery Nanoplatform for Translational Cancer Medicine

“…In a scenario where anionic PMLA segments are preferentially situated on the surface of polyelectrolyte complexes, they are internalized via non-specific endocytosis. Conversely, cationized PMLA complexes translocate into cells in a similar manner to polycations, which first adsorb onto the hydrophilic outer surface of the cell membrane and then induce the formation of aqueous pores or defects in the membrane hydrophobic core, and subsequently integrate into or permeabilize the cell membrane [ 62 ]. Once transported into the cytoplasm, pH reduction or, to a lesser extent an increase in ionic strength, would accelerate hydrolytic degradation of the PMLA backbone and dissociation of ionic bonds, liberating the complexed moiety for intracellular utility [ 61 ].…”