Molecular Mechanism of Resveratrol's Lipid Membrane Protection

Abstract

Resveratrol, a natural compound found in red wine and various vegetables, has drawn increasing interest due to its reported benefit in cardiovascular protection, neurodegenerative disorders, and cancer therapy. The mechanism by which resveratrol exerts such pleiotropic effects remains unclear. It remains as one of the most discussed polyphenol compounds in the debating "French Paradox". In this study, using molecular dynamics simulations of dipalmitoyl phosphatidylcholine (DPPC) bilayer with resveratrol, we generated a free energy map of resveratrol's location and orientation of inside the lipid bilayer. We found that resveratrol increases the surface area per lipid and decreases membrane thickness, which is the opposite effect of the well-studied cholesterol on liquid phase DPPC. Most importantly, based on the simulation observation that resveratrol has a high probability of forming hydrogen bonds with sn-1 and sn-2 ester groups, we discovered a new mechanism using experimental approach, in which resveratrol protects both sn-1 and sn-2 ester bonds of DPPC and distearoyl phosphatidylcholine (DSPC) from phospholipase A1 (PLA1) and phospholipase A2 (PLA2) cleavage. Our study elucidates the new molecular mechanism of potential health benefits of resveratrol and possibly other similar polyphenols and provides a new paradigm for drug design based on resveratrol and its analogs.

Figure 1

The two dimensional (a), and three-dimensional structure (b) of a trans-resveratrol molecule.

Figure 2

DSC Scans of phospholipid membranes with various amounts of resveratrol (mole %): (a) DPPC, (b) DSPC.

Figure 3

Distribution of resveratrol molecules in DPPC lipid bilayer. First snapshot and last snapshot of the simulated system at 310 K and 323 K. 18 resveratrol molecules are shown in VDW mode with atom color and DPPC lipids are shown in line with atom color: green for carbon, blue for nitrogen, red for oxygen, tan for phosphorus. All hydrogen atoms and water molecules are omitted for clarity.

Figure 4

Depth of resveratrol molecules in DPPC lipid bilayer. The distance between the center of mass of 18 resveratrol molecules and bilayer center along z-axis during the 300 ns simulations at two temperatures, 310 K (top) and 323 K (bottom). Each resveratrol is indicated with unique color code. Bilayer center is located at z = 0 Å and the average center of mass of polar head are located at z = 20.5, illustrated on a lipid molecule.

Figure 5

The effect of resveratrol molecules on DPPC lipid surface area and bilayer thickness over last 100 ns simulations at 310 K and 323 K. Left panel shows the time evolution of surface area per lipid and the corresponding distribution for the DPPC simulation with resveratrol. Right panel shows the time evolution of bilayer thickness and the corresponding distribution for the DPPC simulation with resveratrol. The average values are compared with pure DPPC simulations in the Table 2.

Figure 6

Calculated NMR deuterium order parameters (S_CD) for DPPC bilayer with (dashed line) and without (solid line) resveratrol at two temperatures from simulations. The experimental values at 323 K are taken from refs^– (black).

Figure 7

Free energy potential map of resveratrol distribution in the lipid membrane. The x-axis indicates the free energy of the angle distribution (illustrated on a resveratrol molecule) and y-axis are the free energy along membrane normal (illustrated on a DPPC molecule). The 2D-free energy maps are calculated from the last 100 ns simulation at 310 K and 325 K temperatures. The integrated 1D-free energy profiles of resveratrol’s angle along z-axis are shown at the top and bottom of the maps. The integrated 1D-free energy profiles of resveratrol’s center of mass z-position along membrane normal are shown on the left of the maps.

Figure 8

Probability of various types of hydrogen bonds (H-bonds) between resveratrol (Resv) and DPPC. Those that have higher than 10% probability are illustrated with insets. NA: no H-bond; H-bonds between two groups are indicated by the following four-letter codes.To: resv 3,5-OH; B: resv 4′-OH; H: DPPC phosphate group; Ta: DPPC ester group. For example, ToH: H-bonds between resv 3,5-OH and lipid phosphate oxygen; ToTa: H-bonds between Resv 3,5-OH and lipid ester group; ToTaH: H-bonds between resv 3,5-OH and both lipid phosphate group and ester group; BH: Hbonds between resv 4′-OH and lipid phosphate group.

Figure 9

Resveratrol’s protective effect on DPPC and DSPC membranes from the hydrolytic activity of PLA2 or PLA1 at room temperature: Protection of resveratrol from PLA 2 on DPPC (a); or DSPC membranes (b). Protection of resveratrol from PLA 1 on DPPC (c); or DSPC membranes (d). Data are presented as free fatty acid (FFA) vs. DPPC or DSPC lipid membranes with different percentages of resveratrol. Results were analyzed using one-way ANOVA followed by Tuckey’s multiple comparison tests. The values are mean values ± the standard error of the mean (n = 3). Bars with asterisks denote significant difference from control (lipid membranes without resveratrol) (****p < 0.0001, ***p < 0.0006, *p < 0.02).

Figure 10

Mechanism of resveratrol protects lipid from PLA1 and PLA2. (a) Hydrolytic attack by PLA1 or PLA2 (red scissors) on the sn-1 or sn-2 ester bond of a phospholipid molecule, respectively. (b) Phospholipid bilayer membranes attacked by PLAs (1 or 2) release free fatty acid (yellow bars) and lysolipid molecules (yellow bars with red dots). Lysolipids then form micelles. In the presence of resveratrol, PLAs (1 or 2) phospholipid membranes are protected at most concentrations tested. Figure (b) is modified from Phospholipid Bilayer by OpenStax (used under CC BY 4.0).