Receptor mimicry by antibody F045-092 facilitates universal binding to the H3 subtype of influenza virus

Abstract

Influenza viruses present a significant health challenge each year, as in the H3N2 epidemic of 2012-2013. Here we describe an antibody, F045-092, that possesses broadly neutralizing activity against the entire H3 subtype and accommodates the natural variation and additional glycosylation in all strains tested from 1963 to 2011. Crystal structures of F045-092 in complex with HAs from 1975 and 2011 H3N2 viruses reveal the structural basis for its neutralization breadth through insertion of its 23-residue HCDR3 into the receptor-binding site that involves striking receptor mimicry. F045-092 extends its recognition to divergent subtypes, including H1, H2 and H13, using the enhanced avidity of its IgG to overcome lower-affinity Fab binding, as observed with other antibodies that target the receptor-binding site. This unprecedented level of antibody cross-reactivity against the H3 subtype can potentially inform on development of a pan-H3 vaccine or small-molecule therapeutics.

Figure 1. Antibody F045-092 binds the influenza HA receptor-binding site

Crystal structures of F045-092 Fab in complex with (a) Vic1975/H3 HA and (b) Vic2011/H3 HA. One Fab-HA protomer is colored with HA1 in light blue, HA2 in green, the Fab heavy chain in purple, and the Fab light chain in pink. Glycans are shown as spheres with carbon in yellow, oxygen in red, and nitrogen in blue. (c) F045-092 binds HA using only its heavy chain and inserts its HCDR3 into the HA receptor-binding site. The HCDR3 contains a disulfide bridge, which is shown in yellow sticks.

Figure 2. Receptor-binding site recognition by F045-092 to Vic1975/H3 in comparison to the α2,6 sialoglycan receptor

(a) F045-092 inserts its HCDR3 into the HA receptor-binding site. The carboxylate side chain of F045-092 Asp100e overlaps with the (b) carboxylate moiety of the sialic acid of the α2,6 sialoglycan receptor (PDB 2YP4) and uses identical hydrogen bonding interactions, which are shown as black dashed lines. An additional hydrogen bond between the main chains of F045-092 and HA is circled in red dashed lines. The Leu100d side chain of F045-092 overlaps with the acetamide moiety of sialic acid.

Figure 3. Sequence conservation of the F045-092 epitope across human H3 viruses

(a) The F045-092 contact residues on HA are depicted as sticks. The percent conservation for the most common residue found in the H3 subtype is labeled, which is not always identical to the residue at that position in the Vic1975/H3 sequence. (b) Interaction of F045-092 with Vic1975/H3 illustrating the sequence conservation of neutralizing epitope on the HA surface. For identification of the HA residues and positions that are color coded on the surface, see (a). The F045-092 contact residues are labeled, colored by HCDR, and shown as sticks. (c) Percent identity of the most common residue among human H3 isolates plotted against the linear sequence of HA. Residues contacted by F045-092 or CR9114 are indicated by red or green diamonds, respectively. Although F045-092 contacts more hypervariable residues than CR9114, it can still broadly neutralize the H3 subtype.

Figure 4. Emergence of predicted N-linked glycans (PNGs) across human H3 viruses and evasion by F045-092

(a) Stacked graph showing the normalized percentage of strains that contain an additional PNG on HA1 at each indicated position. Seven PNGs have been incorporated in the HA since 1968 in addition to the five absolutely conserved PNGs in H3 strains (HA1 Asn22, Asn38, Asn165, Asn285; HA2 Asn154), except for the PNG at Asn81 (1968-1977). (b) F045-092 evades the glycan at Asn133 proximal to the receptor-binding site in the Vic2011/H3 complex. The glycan is shown as spheres, with carbon in yellow, oxygen in red, and nitrogen in blue. No density corresponding to the PNG at Asn144 (shown as sticks) was observed.

Figure 5. Common trends in recognition of the HA by receptor-binding site targeted antibodies

(a) Antibodies HC19 (red, PDB 2VIR), HC63 (light blue, PDB 1KEN), CH65 (green, PDB 3SM5), and 5J8 (orange, PDB 4M5Z) use an Asp residue to insert a carboxylate in the receptor-binding site that would be occupied by the carboxylate of sialic acid of the α2,6 sialoglycan receptor (dashed red circle). (b) Antibodies HC19 (red, PDB 2VIR), C05 (yellow, PDB 4FP8), 8F8 (blue, PDB 4HF5), 8M2 (pink, PDB 4HFU), and 2G1 (green, PDB 4HG4) use hydrophobic residues to target the receptor-binding site that would be occupied by the acetamide moiety of sialic acid (dashed blue circle). The disulfide loop of the F045-092 HCDR3 is shown and colored purple.