Characterization of the water defect at the HIV-1 gp41 membrane spanning domain in bilayers with and without cholesterol using molecular simulations

Abstract

The membrane spanning domain (MSD) of human immunodeficiency virus 1 (HIV-1) envelope glycoprotein gp41 is important for fusion and infection. We used molecular dynamics (MD) simulations (3.4 μs total) to relate membrane and peptide properties that lead to water solvation of the α-helical gp41 MSD's midspan arginine in pure dipalmitoylphosphatidylcholine (DPPC) and in 50/50 DPPC/cholesterol membranes. We find that the midspan arginine is solvated by water that penetrates the inner leaflet, leading to a so-called water defect. The water defect is surprisingly robust across initial conditions and membrane compositions, but the presence of cholesterol modulates its behavior in several key ways. In the cholesterol-containing membranes, fluctuations in membrane thickness and water penetration depth are localized near the midspan arginine, and the MSD helices display a tightly regulated tilt angle. In the cholesterol-free membranes, thickness fluctuations are not as strongly correlated to the peptide position and tilt angles vary significantly depending on protein position relative to boundaries between domains of differing thickness. Cholesterol in an HIV-1 viral membrane is required for infection. Therefore, this work suggests that the colocalized water defect and membrane thickness fluctuations in cholesterol-containing viral membranes play an important role in fusion by bringing the membrane closer to a stability limit that must be crossed for fusion to occur.

Keywords: Membrane simulation; Metadynamics; Midspan arginine; Molecular dynamics; Snorkeling; Transmembrane.

Fig. 1

Final cumulative, conformational potential of mean force (PMF) in kcal/mol vs. collective variable (RMSD of backbone compared to perfect helix) in Å from the R694L peptide metadynamics, in black. The PMF vs. CV from the WT1 peptide metadynamics from our previous study is also shown here, in red diamonds [18]. Error bars represent standard deviations of the last 20 ns.

Fig. 2

Collective variable (RMSD of backbone compared to perfect helix) in Å vs. simulation time in ns and histogram of RMSD vs. frequency for equilibrium MD of (A) WT1, (B) R694L, and (C) WT1ΔChol systems.

Fig. 3

(A, C, and E) Mass density in 10² amu/ų along membrane normal in Å during equilibrium MD for various components of the system: protein, local water (defined as within 4 Å of protein), and midspan residue (arginine or leucine). (B, D, and F) Mass density in amu/ų along membrane normal in Å during equilibrium MD for various components of the system: lipid tails, lipid headgroups, water, and cholesterol. A and B are averages for WT1,WT2, and WT3 systems. E and F are averages for WT1ΔChol, WT2ΔChol, and WT3ΔChol systems. All statistics are from the last 100 ns of each trajectory and error bars in A, B, E and F represent standard error.

Fig. 4

Representative system configurations from equilibrium MD at 300 ns rendered in VMD [42] for the WT1, R694L, WT1ΔChol, and WT3ΔChol systems. Lipid headgroups are in red vdW, lipid tails in white vdW, cholesterol in yellow vdW, water in cyan isosurface, peptide in orange new cartoon, and R694 or R694L in orange vdW. Water with oxygens within 4 Å of the protein and located on the inner leaflet of the membrane are shown in cyan vdW. For clarity, lipids, cholesterol, and water in the foreground of all four configurations are not shown.

Fig. 5

Number of water molecules within 4 Å of protein vs. amino acid that each water molecule is uniquely attributed to during equilibrium MD. All statistics are from the last 100 ns of each trajectory and error bars represent standard deviation.

Fig. 6

Tilt angle in degrees vs. simulation time in ns.

Fig. 7

Maps of membrane thickness, L, (top) in Å and standard deviation, σ_L, (bottom) in Å for the WT1, R694L, WT1ΔChol, and WT3ΔChol systems during 100 ns intervals of equilibrium MD. Overlaid on the maps are the x and y positions of the non-hydrogen atoms of the peptide (black circles) and the 694 residue (white circles) from the last frame of the trajectories. The N- and C-termini are labeled.

Fig. 8

Maps of average minimum distance along membrane normal in the inner leaflet of water molecules from global center of mass of lipid bilayer, Min (top), in Å and standard deviation, σ_Min, (bottom) in Å for WT1, WT1ΔChol, and WT3ΔChol systems.