Mohammad Alwarawrah

A Molecular View of the Cholesterol Condensing Effect in DOPC Lipid Bilayers

The condensing effect of cholesterol in dioleoylphosphatidylcholine (DOPC) lipid bilayers was systematically investigated via atomistic molecular dynamics (MD) simulation. Fourteen independent 200 ns simulations, spanning the entire range of cholesterol mole fraction (x(c)) in DOPC bilayers (i.e., from x(c) = 0 to 0.66), were performed at 323 K. The molecular areas occupied by DOPC and cholesterol at different distances from the bilayer center were analyzed using a slicing method based on the VDW radii of atoms. Curiously, while the average area per lipid and the cholesterol tilt angle, with respect to the bilayer normal, both show monotonic decreases as x(c) increases, the average bilayer height shows a significant decrease for x(c) > 0.35, following an initial increase. The calculated partial-specific areas of lipids clearly show the condensing effect of cholesterol. The VDW areal analysis showed that the condensing effect is limited only to the cholesterol sterol ring region, where the acyl chains of DOPC are severely compressed by cholesterol. As x(c) increases, the headgroups of DOPC gradually expand along the bilayer-aqueous interface to occupy more lateral area. Thus, it confirmed a key prediction of the umbrella model. At high cholesterol mole fractions, the calculated area per DOPC and area per cholesterol using some existing methods showed an inconsistent result: both increase while the overall area per lipid decreases. The inconsistency stems from the problematic assumption that cholesterol and DOPC have cylindrical shape and the same height. Our results showed that the total area of a PC/cholesterol bilayer is primarily determined by the molecular packing in the cholesterol sterol ring region. An alternative analysis of area per molecule within this region is proposed, which takes into account the cholesterol tilt angle and the practical incompressibility of cholesterol sterol rings. The new calculation shows that the majority of the area lost due to the cholesterol condensing effect is taken from PC molecules.

Investigation of the Effect of Bilayer Composition on PKCα-C2 Domain Docking Using Molecular Dynamics Simulations

The protein kinase Cα (PKCα) enzyme is a member of a broad family of serine/threonine kinases, which are involved in varied cellular signaling pathways. The initial step of PKCα activation involves the C2 subunit docking with the cell membrane, which is followed by interactions of the C1 domains with diacylglycerol (DAG) in the membrane. Notably, the molecular mechanisms of these interactions remain poorly understood, especially what effects, if any, DAG may have on the initial C2 docking. To further understand this process, we have performed a series of conventional molecular dynamics simulations to systematically investigate the interaction between PKCα-C2 domains and lipid bilayers with different compositions to examine the effects of POPS, PIP2, and 1-palmitoyl-2-oleoyl-sn-glycerol (POG) on domain docking. Our results show that the PKCα-C2 domain does not interact with the bilayer surface in the absence of POPS and PIP2. In contrast, the inclusion of POPS and PIP2 to the bilayer resulted in strong domain docking in both perpendicular and parallel orientations, whereas the further inclusion of POG resulted in only parallel domain docking. In addition, lysine residues in the C2 domain formed hydrogen bonds with PIP2 molecule bilayers containing POG. These effects were further explored with umbrella sampling calculations to estimate the free energy of domain docking to the lipid bilayer in the presence of one or two PIP2 molecules. The results show that the binding of one or two PIP2 molecules is thermodynamically favorable, although stronger in bilayers lacking POG. However, in POG-containing bilayers, the binding mode of the C2 domain appears to be more flexible, which may have implications for activation of full-length PKCα. Together, our results shed new insights into the process of C2 bilayer binding and suggest new mechanisms for the roles of different phospholipids in the activation process of PKCα.

Molecular Dynamics Simulation of Diacylglycerols in Phosphatidycholine Lipid Bilayers

In this study, atomistic MD simulations were performed to investigate the interactions between diacylglycerols (i.e. DPG, POG, or DOG) and phosphatidylcholine (i.e. POPC or DOPC) bilayers. Our results show that diacylglycerols (DAG) increase acyl chain order, headgroup spacing and bilayer thickness, and reduce area-per-lipid. In a lipid bilayer, in order to avoid the unfavorable exposure of DAG hydrophobic parts to water, neighboring phospholipid (PC) headgroups move toward DAG to provide cover. This interaction between DAG and phospholipid is explained by the Umbrella Model. Comparing the three types of DAG in POPC and DOPC bilayers, DOG is located closer to the bilayer/aqueous interfaces than DPG and POG and it requires more coverage according to our umbrella index calculation, likely due to its longer and unsaturated chains. The potentials of mean force were calculated for POPC in its bilayers containing DPG, POG, or DOG. Our results show that POPC flip-flop induces pores in pure PC bilayer, while adding diacylglycerol prevents the pore formation during flip-flop. System that contains POG has a higher free energy of desorption and lower free energy barriers for flip-flop, compared with systems that contain DOG or DPG. This indicates that POG is more favorable in POPC bilayer than the other DAGs, since its acyl chains are similar to POPC.

Inhibition of Melittin Activity by Cholesterol, Unsaturated Lipid, and Negatively Charged Lipids Studied by Molecular Dynamics Simulation

Melittin is shown to cause membrane lyses. However, its lytic activity depends on the lipid composition of the membrane. Experiments have shown that the presence of some lipid components in the membrane can inhibit melittins lytic power against the membrane. For the sake of atomistic details, molecular dynamics simulations was used to investigate the inhibition of melittin activity by cholesterol, unsaturated phospholipid (POPE), and negatively charged phospholipid (POPS). A pure DPPC lipid bilayer with melittin was simulated as a control, and significant disturbance of the bilayer by melittin was found. The order parameter of DPPC changed dramatically and a large curvature was observed in one of the membrane leaflets, probably leading to a future membrane rupture. The DPPC bilayer with cholesterol showed strong resistance to melittin: Melittin can hardly bind to the membrane surface. In the simulation with unsaturated lipid, melittin can only bind either its N or C terminal region to the bilayer, but the interaction between the body of melittin and the lipids is weak. Melittin binds strongly to negatively charged lipids; however, it cannot induce membrane curvature nor disruption. In above three simulations, C termini of melittin were observed to be able to bind to the membrane more easily than N termini. A deep and stable adsorption of melittin to the membrane requires the binding of both N and C-terminal regions to the membrabe. The tendency of melittin to aggregate was observed in all simulations, especially in the simulation containing cholesterol. This study provides insight into the possible mechanisms of the inhibition of melittins lytic activity by different membrane components.

Study of Cholesterol-Diacylglycerol-Pc Interactions in POPC Bilayer using Molecular Dynamics Simulation

Diacylglycerol(DAG)-cholesterol-phospholipid interactions have received increasing attention for their involvement in lipid rafts. The interaction of DAG with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol were systematically investigated via atomistic molecular dynamics (MD) simulation. Four independent bilayer simulations (i.e., pure POPC, POPC/Cholesterol, POPC/DAG, and POPC/DAG/Cholesterol) were performed at 310K. Cholesterol and DAG both have small hydrophilic headgroups and large hydrophobic bodies. The 2D radial distribution functions of molecules show an Umbrella effect between cholesterol and POPC, and also between DAG and POPC. In the ternary mixture of POPC/DAG/cholesterol, cholesterol increases the order parameters of POPC acyl chains as well as DAG acyl chains. The average distances between acyl chains of POPC and DAG decrease with the addition of cholesterol. The cholesterol tilt angle with the bilayer normal in POPC/DAG/cholesterol bilayer is smaller than that in POPC/cholesterol bilayer.

Molecular Interactions in Phase Separation of DOPC/DSPC/cholesterol Ternary Mixtures

With the aim of investigating molecular interactions between lipids involved in lipid-raft formation, the experimental phase diagram of a DOPC/DSPC/cholesterol ternary system was simulated using Monte Carlo simulation. Both pairwise (Ising-like) and multibody interactions were used to simulate the phase boundary of liquid-ordered phase and liquid-disordered (Lo-Ld) phase coexistence regions. The “Composition Histogram Method” (CHM) was specifically developed to quickly determine the compositions of coexisting phases as well as the thermodynamic tie-lines. The simulation demonstrated that the phase boundaries produced by pairwise (Ising-like) interactions alone generally do not agree with the experimental phase boundary. A much better fit for the experimental phase boundary was obtained by including a “domain edge energy” term, which is expressed in a form ofssmultibody interaction. Our result shows that the “domain edge energy” is essential for creating phase separation in lipid raft mixtures. The magnitude of this interaction energy determines the location of the critical point, the shape the phase boundary, and the size distribution of lipid domains in lipid raft mixtures. Any experimental condition that alters the domain edge energy, could significantly change the shape and location of the Lo-Ld phase boundary.