The vertical location of 30 mol % cholesterol in a hydrated dimyristoylphosphatidylcholine (DMPC) membrane was determined by neutron diffraction on annealed samples containing deuterated or protonated cholesterol at 10, 20, 25, 30, and 50 °C. The sterol was deuterium-labeled in positions 2, 2, 3, 4, 4, and 6, and proton-deuterium contrast techniques were used to locate the position of the labeled part of the steroid in the membrane. Cholesterol is found well embedded in the membrane, with ring A at 16.3 ( 0.5 Å from the bilayer center at 10 °C. This location linearly decreases to 15.1 ( 0.5 Å at 50 °C, demonstrating that the sterol is not expelled from the membrane on crossing the former gel-to-fluid phase transition of pure DMPC (24 °C). Molecular dynamics were also performed on well-hydrated membranes in the presence and absence of cholesterol. Neutron scattering 1D profiles were then calculated for comparison with experimental neutron scattering data. The profile obtained from pure fluid-phase lipids is in nice agreement both in shape and in bilayer hydrophobic thickness with the experiment. The pure gel-phase calculation leads to the correct line shape but with an overestimated bilayer thickness. In the presence of cholesterol, only the calculation performed with initial gel-phase conditions leads to a hydrophobic thickness in agreement with neutron data. Ring A of cholesterol is found at 15.2 ( 0.5 Å at 10 °C, underestimating the experimental value by only 1 Å. Molecular dynamics show that the hydroxyl group of cholesterol is hydrated and in such a proximity to the carboxyl oxygens of the phospholipids that it can make hydrogen bonds. The ability for molecular dynamics calculations on membranes to determine structural data in membranes is finally discussed.
We present here the results of 1-ns molecular dynamics (MD) simulations of two ideally amphipathic lytic peptides, namely LK(15) and LK(9), in a 1,2-dimyristoylphosphatidylcholine monolayer with two different cross-sectional areas per lipid of 80 A(2) (loose film) and 63 A(2) (tight standard film). These peptides are lytic, ideally amphipathic with a minimalist composition L(i)K(j) and the following sequences: H(2)N-KLLKLLLKLLLKLLK-CO-Ph for LK(15) and H(2)N-KLKLKLKLK-CO-Ph for LK(9). From experimental data, LK(15) exhibits an alpha-helical secondary structure, whereas LK(9) was found to form antiparallel beta-sheets at the interface of a DMPC monolayer. Whatever the specific lipid surface is, the two peptides exhibit very different behavior: the alpha-helix inserts deeply into the monolayer whereas the beta-sheeted peptide stays at the surface within the upper polar part of the monolayer. In all cases, a loose monolayer (80 A(2)) results in noticeable artifacts whereas a monolayer with standard specific surface leads to very reliable behavior well in accordance with experimental data. Despite their different insertion depth, the two peptides exhibit identical lytic efficiency. This is very likely a direct consequence of the very strong Van der Waals interactions between the fatty alkyl chains of the lipids and the highly lipophilic lower part of the peptide, resulting in an identical thinning of the two monolayers.
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