The structural basis for filovirus neutralization by monoclonal antibodies

Abstract

Filoviruses, including ebolaviruses and marburgviruses, are the causative agents of highly lethal disease outbreaks. The 2013-2016 Ebola virus outbreak was responsible for >28000 infections and >11000 deaths. Although there are currently no licensed vaccines or therapeutics for any filovirus-induced disease, monoclonal antibodies (mAbs) are among the most promising options for therapeutic development. Hundreds of mAbs have been isolated from human survivors of filovirus infections that target the viral spike glycoprotein (GP). The binding, neutralization, and cross-reactivity of many of these mAbs has been determined. Several mAbs have been characterized structurally, and this information has been crucial for strategizing therapeutic and vaccine design. Here we present an overview of the structural features of the neutralizing/protective epitopes on filovirus glycoproteins.

Figure 1. Antibody epitopes on filovirus GPs

(A) Ebolavirus GP with antibody binding epitopes shown as patches of color on the GP surface (PDB: 5JQ7) [65] and a corresponding sequence map below. Labels for ebolaviruses: SP = Signal Peptide, I = Base, II = Head, CL = Cathepsin Cleavage Loop, III = Glycan Cap, IV = Mucin-like Domain (MLD), V = N-terminal Loop, VI = Fusion Loop, VII = Heptad Repeat 1 (HR1), VIII and IX are together Heptad Repeat 2 (HR2), of which IX = Stalk, X = Membrane Proximal External Region (MPER), and TM = Transmembrane domain. (B) Marburgvirus GP with antibody binding epitopes shown as patches of color on the GP surface (PDB: 6BP2) [25]. Labels for marburgviruses: SP = Signal Peptide, I = GP₁, * = Receptor binding site, II = Glycan Cap, III = MLD, IV = Wing, V = N-terminal loop, VI = Fusion Loop, VII = HR1, VIII = HR2, IX = MPER, and TM = Transmembrane domain. The RBS is illustrated only on marburgvirus GP for clarity; on uncleaved ebolavirus GP, the glycan cap masks the RBS.

Figure 2. Visualization of the Mucin-Like Domain

(A) The crystal structure of the mucin-deleted EBOV GP (PDB: 5JQ3) [65] is shown docked into a subtomogram averaged map of mucin-deleted EBOV GP [66] and a single-particle generated map of intact, mucin-containing EBOV GP (B) [22]. Although the observation of density for mobile regions is limited by technical factors in single particle reconstruction, the regions of the mucin-like domain that are visible appear to extend upwards and outwards from the glycan cap and base region, thereby shielding much of the GP core.

Figure 3. Structural similarities between sGP and GP

(A) The EBOV sGP dimer (PDB: 5KEM) [55] is shown as a cartoon with each monomer shaded orange. (B) The EBOV GP trimer (PDB: 5JQ3) [65] is shown as a cartoon with GP₁ (dark gray) and GP₂ (white). (C) GP₁ and a single monomer of sGP align with a C_α r.m.s.d. of 3.87 Å over 217 aligned residues. sGP and GP share 100% sequence identity for their first 295 amino acids. (D) They are structurally similar between the core residues 66–184 and 216–259 (blue), although parts of these regions are likely obscured from immune surveillance by the dimerization interface. (E) The isolated cores (same as the blue region in (D)) align with a C_α r.m.s.d. of 1.71 Å over 163 residues.

Figure 4. Neutralizing epitopes identified for marburgvirus

(A) Crystal structures of neutralizing antibodies MR191 [25] and MR78 [49] bound within the RBS of RAVV GP. (B) A phenylalanine at the apex of the CDR-H3s of both antibodies reaches into the hydrophobic pocket of the marburgvirus GP in a manner that structurally mimics interactions of ebolavirus GP both with its glycan cap and its host receptor, NPC1-C [12]. (C) A marburgvirus GP is shown with a single GP monomer colored in blue (GP₁) and gold (GP₂). The remaining two monomers are grey. The anchor of the wing domain (orange) is shown wedged underneath the base of GP. (D) Enlarged view of the wing illustrating the β-strand wing anchor region (connected by an 8 aa linker), and the relative position of the 33 aa wing targeted by antibodies.