Structure and antigenicity of divergent Henipavirus fusion glycoproteins

Abstract

In August 2022, a novel henipavirus (HNV) named Langya virus (LayV) was isolated from patients with severe pneumonic disease in China. This virus is closely related to Mòjiāng virus (MojV), and both are divergent from the bat-borne HNV members, Nipah (NiV) and Hendra (HeV) viruses. The spillover of LayV is the first instance of a HNV zoonosis to humans outside of NiV and HeV, highlighting the continuing threat this genus poses to human health. In this work, we determine the prefusion structures of MojV and LayV F proteins via cryogenic electron microscopy to 2.66 and 3.37 Å, respectively. We show that despite sequence divergence from NiV, the F proteins adopt an overall similar structure but are antigenically distinct as they do not react to known antibodies or sera. Glycoproteomic analysis revealed that while LayV F is less glycosylated than NiV F, it contains a glycan that shields a site of vulnerability previously identified for NiV. These findings explain the distinct antigenic profile of LayV and MojV F, despite the extent to which they are otherwise structurally similar to NiV. Our results carry implications for broad-spectrum HNV vaccines and therapeutics, and indicate an antigenic, yet not structural, divergence from prototypical HNVs.

Fig. 1. Cryo-EM structure of LayV F glycoprotein in the prefusion form.

a Gene schematic of LayV F with each domain coloured. Domains coloured in white are not included in the final structure. b SDS-PAGE of purified LayV F proteins under reducing conditions either stabilized with clamp2 or unstabilised. c SEC of LayV F clamp2 ran on Superose 6 Increase 10/300 GL column. Trimer peak is indicated. d Cryo-EM map (grey) resolved to 3.67 Å and generated model coloured as in (a) of prefusion LayV F. Source data are provided as a Source Data file.

Fig. 2. Sequence and structural comparison between LayV and MojV F proteins.

a MojV F protein surface representation coloured in light grey with a single monomer coloured in purple. Conserved mutations between MojV and LayV F are coloured in green and non-conserved mutations are coloured in orange. b Model comparisons between LayV F (grey) and MojV F (purple) within different domains. Conserved mutations are coloured green and non-conserved mutations are coloured orange. Non-conserved MojV F side chains are depicted as sticks with oxygens coloured red and nitrogen coloured blue.

Fig. 3. LayV and MojV F are antigenically distinct from NiV F.

a LayV F (purple), MojV F (pink) and NiV F (white) trimer overlay. Inset 1 shows inter-protomer DIII contacts which are shared for LayV and MojV but distinct from NiV, a shift in a surface loop (arrow) projects into the interprotomer interface resulting in lateral translocation of the opposing DIII helix. Inset 2 shows structural homology between LayV and NiV at the F1/F2 cleavage site compared to MojV, cathepsin cleavage site is indicated. Antibody binding site and footprint are shown for 5B3 (b), mAb66 (c), 12B2 (d) and 1F5 (e). Indirect ELISAs of each mAb are shown to the right against prefusion clamp2 stabilized NiV F, MojV F and LayV F. Two replicates for each datapoint are shown. In each antibody footprint map, NiV F is shown with conserved residues in LayV F depicted as green and non-conserved as grey, with glycans coloured in purple. Source data are provided as a Source Data file.

Fig. 4. Glycan occupancy in LayV F.

a LayV F cryo-EM map (grey) glycan identification fitted with generated model (purple). N-linked sites coloured red and Asn residues shown as sticks. Glycan structures observed at N65 (b) and N459 (c) by mass spectrometry glycoproteomics. N-Acetylglucosamine is presented as blue squares, mannose as green circles and fucose as red triangles. d Schematics highlighting the position of the occupied glycosylation sites in LayV and NiV F and the proportion of each glycan type at the sites in LayV.