Effect of flanking residues on the conformational sampling of the internal fusion peptide from Ebola virus

Abstract

Fusion peptides mediate viral and host-cell membrane fusion during viral entry. The monomeric form of the internal fusion peptide from Ebola virus was studied in membrane bilayer and water environments with computer simulations using replica exchange sampling and an implicit solvent description of the environment. Wild-type Ebola fusion peptide (EFP), the W8A mutant form, and an extended construct with flanking residues were examined. It was found that the monomeric form of wild-type EFP adopts coil-helix-coil structure with a short helix from residues 8 to 11 mostly sampling orientations parallel to the membrane surface. W8A mutation disrupts the helicity in the N-terminal region of the peptide and leads to a preference for slightly oblique orientation relative to the membrane surface. The addition of flanking residues also alters the fusion peptide conformation with either a helix-break-helix structure or extended N and C-termini and reduced membrane insertion. In water, the fusion peptide is found to adopt structures with low helicity.

Figure 1

Schematic representation of Ebola fusion protein in fusiogenic state. The globular domain GP1 is initially responsible for binding to the host cell receptor. The GP2 domain contains a helical bundle with the fusion peptide near the N-terminus.

Figure 2

Left panel: Potentials of mean force (PMFs) for EFP (A), W8A (B), and EFP-X (C) from weighted-histogram analysis (WHAM), projected onto insertion angle (in degrees) and distance of the peptide center of mass from the membrane center (in Å). Different colors reflect relative free energies as indicated by the color bar (in kcal/mol). Right panel: Dominant structures for simulations with EFP (A1–A4), W8A (B1–B4) and EFP-X(C1–C4) from clustering. The backbone is shown as a ribbon and side chains are shown as stick representation. Charged residues are colored in red, glycines in green, prolines pale cyan, aromatic residues in white and hydrophobic residues in gray. The N-terminus is marked by a green sphere. Residue 8 is shown in ball and stick representation and colored magenta (W8) and orange (A8 in mutant). The z-axis is pointing up in all figures. All figures were generated with pyMol.

Figure 3

Left panel: Potential of mean force (PMF) of EFP-AQ from weighted-histogram analysis (WHAM), projected onto average helicity based on hydrogen bond criterion and radius of gyration (in Å). Different colors reflect relative free energies as indicated by the color bar (in kcal/mol). Right panel: Dominant structures for simulations with EFP-AQ from clustering as in Fig. 2.

Figure 4

Average fraction of helical conformations as a function of residue for EFP (circle), W8A (triangle), EFP-X (square) and EFP-AQ (cross). A:Helicity is defined by O(i) – H-N(i+4) hydrogen bonding (< 2.6 Å distance); B: Helicity is defined by −70≤ψ≤−50 and −35≤φ≤−55; C: 3₁₀ helicity is defined by O(i) – H-N(i+3) hydrogen bonding (< 2.6 Å distance)