2003 ME Graduate Student Conference

Cellular processes depend on controlled transport of material across the cell membrane, typically by the formation of ion specific channels and pores. The major difficulty in studying trans-membrane pores experimentally is that they are usually below the theoretical resolution of light microscope. Thus, pores have been produced and investigated extensively in membranes using indirect techniques by the application of mechanical stresses or electric fields. During the last decade, realistic atomic-level computer simulations have evolved as complementary techniques in the study of bilayer of lipid molecules. In this study we report molecular dynamic simulation studies that for the first time delineate with atomistic details, the entropy-driven nucleation and growth of hydrophilic pores in lipid bilayers in the presence of edge-active agent. Dimyristoylphosphatidylcholine (DMPC) lipid bilayer was chosen as the model membrane, since it is quite prevalent in biological membranes and is well studied both computationally and experimentally. As edge-active agent we used dimethylsulfoxide (DMSO), one of the most widely used solvents in cell biology and cryopreservation. Our simulations show that when the line tension of the DMPC bilayer is lowered below a certain threshold *, pores can be thermally nucleated and grow over atomistically accessible timescales (a few tens of nanoseconds). The lipid bilayer systems investigated in this study by molecular dynamics (MD) consists of 96 molecules (48 in each leaflet) of DMPC immersed in DMSO-water solutions at various concentrations. In all MD simulations reported in this study, the bilayer systems were followed over 100 ns. The simulation results corroborate with and provide atomistic support and understanding for the phenomenological theory and lattice model predictions of entropy driven pore growth studies of Shillcock and Seifert [1]. SIMULATION METHODOLOGY The MD simulations were performed with the GROMACS molecular dynamics package. Several sets of MD simulations were carried out on DMPC lipid bilayers immersed in pure water, and DMSO-water solution at four concentrations. Systems at four DMSO concentrations (3 mol %, 6 mol %, 9 mol%, and 11.3 mol %) were thoroughly investigated with the lipid bilayers in fluid phase and in equilibrium at 323K. Details of the simulation methodology can be found in Moldovan et al [2] and in Pinisetty et al [3]. The line tensions, , of a linear DMPC bilayer edge, in different DMSO-water solutions, were calculated from separate simulations using the method developed by Tolpekina et al [4]. Ribbon-like (strip) bilayers, consisting of 96 lipids surrounded by water (or by water and DMSO) molecules in the normal (x) and one of the lateral (y) directions, were simulated using periodic boundary conditions in all three directions. The length of the simulation box along the z axis (parallel to the ribbon edge) was fixed, whereas the box dimensions in the x and y directions were scaled jointly. Through this semi-isotropic pressure coupling scheme, a positive line tension is supported along the ribbon edges while system’s volume is allowed to change and equilibrate at 1 atm pressure. The positive line tension, , was calculated from the diagonal elements of the pressure tensor according to the relation , where <...> stands for time averaging. Further simulation details are available elsewhere [1,4]. RESULTS AND DISSCUSSION The structural integrity of the lipid bilayers is investigated by visual inspection of both top and cross view snapshots. Figure 1 shows the formation of hydrophobic pore at 10ns and its gradual transformation to hydrophilic pore in less than 5ns. The hydrophobic to hydrophilic pore metamorphosis is mediated by intricate atomistic rearrangements of the molecular species around the rim of the ij P ] 2 / ) )[( 2 / ( zz yy xx y x P P P L L    