Structural basis for broad neutralization of ebolaviruses\xa0by an antibody targeting the glycoprotein fusion loop

Benjamin M Janus,Nydia Van Dyk,Yuxing Li,M Javad Aman,Xuelian Zhao,Cinque Soto,Gilad Ofek,Thomas R Fuerst,Katie A Howell

doi:10.1038/s41467-018-06113-4

Abstract

The severity of the 2014–2016 ebolavirus outbreak in West Africa expedited clinical development of therapeutics and vaccines though the countermeasures on hand were largely monospecific and lacked efficacy against other ebolavirus species that previously emerged. Recent studies indicate that ebolavirus glycoprotein (GP) fusion loops are targets for cross-protective antibodies. Here we report the 3.72 Å resolution crystal structure of one such cross-protective antibody, CA45, bound to the ectodomain of Ebola virus (EBOV) GP. The CA45 epitope spans multiple faces of the fusion loop stem, across both GP1 and GP2 subunits, with ~68% of residues identical across > 99.5% of known ebolavirus isolates. Extensive antibody interactions within a pan-ebolavirus small-molecule inhibitor binding cavity on GP define this cavity as a novel site of immune vulnerability. The structure elucidates broad ebolavirus neutralization through a highly conserved epitope on GP and further enables rational design and development of broadly protective vaccines and therapeutics.

Highlights

The severity of the 2014–2016 ebolavirus outbreak in West Africa expedited clinical development of therapeutics and vaccines though the countermeasures on hand were largely monospecific and lacked efficacy against other ebolavirus species that previously emerged
Depending on the viral assay and species, CA45 neutralized with IC50 values ranging from 0.9 to 78 nM, while it protected against Ebola virus (EBOV), SUDV, and BDBV virus challenge in mice, guinea pigs, or ferrets, when administered alone or in conjunction with a previously characterized receptor-binding region antibody FVM045,7,19
The purified glycosylated GPΔMuc protein was incubated with the antigen-binding fragment (Fab) of antibody CA45, and the resulting complex separated and further purified by size exclusion chromatography (SEC)

Summary

Results and Discussion

The CA45 epitope on GP observed in the crystal structure was predominantly located in and around the fusion loop stem, made up of strand β19–β20 positioned along the equatorial GP1–GP2 interface of GP. Four residues on GP were previously shown to ablate CA45 binding in alanine scan saturation mutagenesis analyses, one on GP1—R64—and three on GP2—Y517, G546, and N5507 All four of these residues were observed to lie within the CA45-GP interface in the present structure, with Y517 present within the DFF inhibitor-binding pocket, R64 on the lower outer edge of the pocket, and G546 present on the outer face of the β20 fusion loop strand (Fig. 2b–d and below). To further establish the structural basis for breadth of CA45 recognition, we examined sequence variation at each of the 34 CA45 epitope residue positions across the ebolavirus genus (Fig. 3c, d and Supplementary Table 2). Alanine mutation at three additional epitope residues, E103, L515, and W518, knocked out CA45

F98 LCDR1 N40

F98 V100c I100e

Methods