The permeability and flip-flop-properties of lipid bilayers containing oxysterols was studied using molecular dynamics simulations. The two chosen oxysterols were 27-hydroxy-cholesterol (HCH) and 7β-hydroxycholesterol (BCH). Bilayers containing these oxysterols were compared with bilayer containing non-oxidised cholesterol (CHOL) and bilayer without any sterols (pure POPC). Water molecules and oxalate-ions (OXA) were pulled through bilayers and free energy profiles were measured using umbrella sampling. Gromacs 4.6 and OPLS-AA force field were used in all simulations.
Three different bilayer systems with 90% of POPC and 10% of CHOL/HCH/BCH prepared. Additionally one system with 100% POPC was prepared as a control system. All systems were equilibrated for at least 50 ns. I didn’t observe anything interesting in equillibration of 10%BCH system, but 10%HCH has some intriguing properties. During the equilibration run the tail hydroxyl groups of HCH molecules seem to attract each other and they form clusters in the membrane. The tail hydroxyl groups also attract water molecules on other side of the membrane, and decrease the membrane thickness locally. These “local monolayers” might affect the permeability of membrane.
To generate starting configurations for umbrella sampling simulations one water molecule or oxalate-ion was pulled along z-axis (which was parallel to the normal of the bilayer) to the center of the bilayer. In total 32 configurations per system were extracted from pull simulation trajectories (COM distance z from 0 nm to 3.1 nm with 0.1 nm spacing). From each configuration I started a 50 ns or 60 ns (water or OXA) long umbrella sampling simulation, in which the pulled water molecule was kept in same plane by an umbrella potential. The pull force was saved every 1 ps. From the pull forces I calculated the potential of mean force (PMF) using weighted histogram analysis method (WHAM) with gromacs tool
g_wham . When water (or OXA) is pulled deeper into the membrane it always pulls other water molecules with it, forming a “water defect”. Same phenomena has been reported in other similar studies . When water is pulled deep enough, the defect disappears. This causes the flat region in the PMF curve of water. By looking at the PMF curves we can say that the bilayer with 10%HCH is the most permeable, for both water and OXA, and the system with 10%CHOL is least permeable. Pure POPC bilayer and 10%BCH bilayer are something between them.
To study the energetics of POPC flip-flop, one POPC in the membrane was pulled through the membrane in similar manner as water and OXA. The umbrella force acted to the phosphate atom of POPC, as this was easier than pulling the center of mass of the whole molecule. A bit longer longer (100 ns) umbrella sampling simulations were needed, because it takes some time for that big molecule to find the preferred configuration. The energy penalty of flip-flop is smallest in pure POPC membrane. Also in HCH membrane the energy penalty is smaller compared to 10%CHOL and 10%BCH membranes.
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