Structural Insights into the Interaction of Filovirus Glycoproteins with the Endosomal Receptor Niemann-Pick C1: A Computational Study

Abstract

Filoviruses, including marburgviruses and ebolaviruses, have a single transmembrane glycoprotein (GP) that facilitates their entry into cells. During entry, GP needs to be cleaved by host proteases to expose the receptor-binding site that binds to the endosomal receptor Niemann-Pick C1 (NPC1) protein. The crystal structure analysis of the cleaved GP (GPcl) of Ebola virus (EBOV) in complex with human NPC1 has demonstrated that NPC1 has two protruding loops (loops 1 and 2), which engage a hydrophobic pocket on the head of EBOV GPcl. However, the molecular interactions between NPC1 and the GPcl of other filoviruses remain unexplored. In the present study, we performed molecular modeling and molecular dynamics simulations of NPC1 complexed with GPcls of two ebolaviruses, EBOV and Sudan virus (SUDV), and one marburgvirus, Ravn virus (RAVV). Similar binding structures were observed in the GPcl-NPC1 complexes of EBOV and SUDV, which differed from that of RAVV. Specifically, in the RAVV GPcl-NPC1 complex, the tip of loop 2 was closer to the pocket edge comprising residues at positions 79-88 of GPcl; the root of loop 1 was predicted to interact with P116 and Q144 of GPcl. Furthermore, in the SUDV GPcl-NPC1 complex, the tip of loop 2 was slightly closer to the residue at position 141 than those in the EBOV and RAVV GPcl-NPC1 complexes. These structural differences may affect the size and/or shape of the receptor-binding pocket of GPcl. Our structural models could provide useful information for improving our understanding the differences in host preference among filoviruses as well as contributing to structure-based drug design.

Keywords: Niemann-Pick C1; ebolavirus; filovirus; glycoprotein; marburgvirus; molecular dynamics; molecular modeling; structure.

Figure 1

Three-dimensional structure of the EBOV GPcl–NPC1 complex and amino acid sequences of the receptor-binding domain of EBOV, SUDV, and RAVV GPs. (A) The three-dimensional structures of EBOV GPcl trimer and human NPC1-C (PDB ID: 5F1B) are represented as a surface and a ribbon model, respectively. On the GPcl trimer, one monomer (center) is colored white and the others are colored black and dark gray. The GPcl-binding interface, including NPC1 loop 1, and loop 2 (indicated in violet and sky blue, respectively), is shown in the boxed areas. Nitrogen and oxygen atoms are shown in blue and red, respectively. The amino acid residues of loop 1 and loop 2 in NPC1 are displayed. (B) Three receptor-binding domain sequences of filovirus GPcl were aligned using EBOV numbering. Conserved amino acid residues among EBOV, SUDV, and RAVV GPs are shown in red. Solid triangles indicate the positions of contact residues of EBOV GPcl with NPC1 observed in the crystal structure.

Figure 2

Per-residue contribution of NPC1 loops in the GPcl–NPC1 complexes to the binding free energy. Per-residue contribution to the binding free energies of loop 1 and loop 2 in the EBOV, SUDV and RAVV GPcl–NPC1 complexes were calculated using the MM/GBSA free energy decomposition method. Each MD simulation was conducted three times, and average and standard errors (SE) are shown.

Figure 3

Per-residue contribution to GPcl–NPC1 binding. (A) Per-residue contributions to the binding free energy of GPcl in EBOV, SUDV, and RAVV GPcl–NPC1 complexes were calculated using the MM/GBSA free energy decomposition method. Amino acid residues that are distinct from those of EBOV GPcl are shown in red. The numbering scheme for GPcl was adapted from EBOV. Each MD simulation was conducted three times: data are represented as average ± SE. (B) The edge of the hydrophobic pocket is formed by three regions (i.e., amino acid positions 79–88, 111–116, and 141–148). Three-dimensional structures of GPcl in the EBOV GPcl–NPC1 complex are represented as surface models. Dashed arrows indicate the direction of amino acid residues, where the numbers indicate the amino acid position of the cleft of the receptor binding pocket. (C) Differences of binding free energies were calculated by subtraction of the value of EBOV GPcl from those of SUDV and RAVV GPcl molecules. (D) The three-dimensional structures of GPcl in the complexes are represented as surface models. Blue and red denote favorable and unfavorable contributions, respectively, to the binding free energy; the energies less than −3.0 kcal/mol, equal to 0 kcal/mol, and more than +3.0 kcal/mol are colored blue, white, red, respectively.

Figure 4

Interface contact residues in the GPcl–NPC1complexes. (A) Residue contact pairs of the GPcl–NPC1 interfaces for EBOV, SUDV, and RAVV were analyzed using the MOE software (version 2018; Chemical Computing Group, Montreal, Canada). Residue–residue contact pairs that appeared in at least 50% of the 1500 MD simulation frames are shown for EBOV (blue), SUDV (red), and RAVV (yellow) GPcl–NPC1 complexes. In each panel, the light, intermediate dark, and darkest colors represent the contact pairs containing van der Waals interactions (vdW), vdW + hydrogen bonds (hb), and vdW + hb + salt bridges (sb), respectively. Amino acid residues that are distinct from those of EBOV GPcl are shown in red. The numbering scheme for GPcl was adapted from EBOV. (B) Venn diagram for the number of residue contact pairs observed for EBOV (blue), SUDV (red), and RAVV (yellow) GPcl–NPC1 complexes.

Figure 5

Differences in the GPcl–NPC1 binding structures between EBOV, SUDV, and RAVV. (A) Residue contact pairs of the GPcl–NPC1 interfaces for EBOV, SUDV, and RAVV were analyzed using the MOE software (version 2018; Chemical Computing Group, Montreal, Canada). Residue–residue contact pairs that appeared in at least 50% of the 1500 MD simulation frames, are shown for EBOV (blue), SUDV (red), and RAVV (yellow) GPcl–NPC1 complexes. In each panel, the light, intermediate dark, and darkest colors represent the contact pairs containing van der Waals interactions (vdW), vdW + hydrogen bonds (hb), and vdW + hb + salt bridges (sb), respectively. Amino acid residues that are distinct from those of EBOV GPcl are shown in red. The numbering scheme for GPcl was adapted from EBOV. (B) Venn diagram for the number of residue contact pairs observed for EBOV (blue), SUDV (red), and RAVV (yellow) GPcl–NPC1 complexes.

Figure 6

MD simulations for free GPcl molecules of EBOV, SUDV, and RAVV. (A) The volumes of the receptor-binding pocket in the free GPcl molecules of EBOV (blue), SUDV (red), and RAVV (yellow) were calculated with the POVME 3.0 program. The values in the plot area represent the mean and standard deviation (SD) of the pocket volumes from each monomer in the GPcl trimer. In each box plot, the mean and median values are indicated by “x” and a solid horizontal bar, respectively. The top and bottom edges of the box mark the first and third quartile, respectively. Outliers, shown as open circles, are cases with values more than 1.5 times the interquartile range away from the upper or lower quartile. The whiskers extending from the box indicate the highest and lowest values, excluding the outliers. (B) RMSF values were plotted for free EBOV (blue), SUDV (red), and RAVV (orange) GPcl structures. Gray regions between the numbers above the plot area represent the constituent residues of the receptor binding pocket. (C) The Cα−Cα distances (Å) of V79–V141, I79–A141, and P79–I141 were measured for each monomer in the GPcl trimers of EBOV, SUDV, and RAVV, respectively. Distributions of the Cα distances during MD simulations are shown for EBOV (blue), SUDV (red), and RAVV (orange) GPcl molecules. The average distance and SD are indicated in each panel. A total of 1500 snapshots from each trajectory of the last 50 ns were used for all the analyses (A–C).