Conserved Neutralizing Epitope at Globular Head of Hemagglutinin in H3N2 Influenza Viruses

Abstract

Neutralizing antibodies that target the hemagglutinin of influenza virus either inhibit binding of hemagglutinin to cellular receptors or prevent the low-pH-induced conformational change in hemagglutinin required for membrane fusion. In general, the former type of antibody binds to the globular head formed by HA1 and has narrow strain specificity, while the latter type binds to the stem mainly formed by HA2 and has broad strain specificity. In the present study, we analyzed the epitope and function of a broadly neutralizing human antibody against H3N2 viruses, F005-126. The crystal structure of F005-126 Fab in complex with hemagglutinin revealed that the antibody binds to the globular head, spans a cleft formed by two hemagglutinin monomers in a hemagglutinin trimer, and cross-links them. It recognizes two peptide portions (sites L and R) and a glycan linked to asparagine at residue 285 using three complementarity-determining regions and framework 3 in the heavy chain. Binding of the antibody to sites L (residues 171 to 173, 239, and 240) and R (residues 91, 92, 270 to 273, 284, and 285) is mediated mainly by van der Waals contacts with the main chains of the peptides in these sites and secondarily by hydrogen bonds with a few side chains of conserved sequences in HA1. Furthermore, the glycan recognized by F005-126 is conserved among H3N2 viruses. F005-126 has the ability to prevent low-pH-induced conformational changes in hemagglutinin. The newly identified conserved epitope, including the glycan, should be immunogenic in humans and may induce production of broadly neutralizing antibodies against H3 viruses. IMPORTANCEAntibodies play an important role in protection against influenza virus, and hemagglutinin is the major target for virus neutralizing antibodies. It has long been believed that all effective neutralizing antibodies bind to the surrounding regions of the sialic acid-binding pocket and inhibit the binding of hemagglutinin to the cellular receptor. Since mutations are readily introduced into such epitopes, this type of antibody shows narrow strain specificity. Recently, however, broadly neutralizing antibodies have been isolated. Most of these bind either to conserved sites in the stem region or to the sialic acid-binding pocket itself. In the present study, we identified a new neutralizing epitope in the head region recognized by a broadly neutralizing human antibody against H3N2. This epitope may be useful for design of vaccines.

A human monoclonal antibody derived from a vaccinated volunteer recognizes heterosubtypically a novel epitope on the hemagglutinin globular head of H1 and H9 influenza A viruses

“…Most heterosubtypic crossneutralizing HuMAb have been reported to recognize epitopes located mainly in the most conserved HA stem region [7,10,14,23]. Also, a few HuMAbs that target relatively conserved epitopes adjacent to or overlapping the RBS in the head region were shown to be subtype-specific [17,25,26]. To our knowledge, 5D7 is the first heterosubtypic neutralizing HuMAb that targets a conserved epitope distinct from the RBS in the HA globular head region.…”

Epitope-focused vaccine design against influenza A and B viruses

“…Third, the head contains epitopes that are conserved in diverse strains within a HA subtype. These epitopes are located in the RBS [22][23][24][25][26][27] or outside the RBS [27][28][29][30][31]32 ]. The RBS binding antibodies are common in vaccine-induced antibody responses.…”

“…Fourth, an antibody recognizes the epitopes located at the HA head/stem interface [34], which are similar to anti-HIV-1 antibody 35O22 [35] or anti-EBOV GP antibody 2G4 [36]. Two antibodies recognize the quaternary epitopes [31,37], which are similar to anti-HIV-1 antibody PGT151 [38] or anti-RSV profusion F antibody AM14 [39]. Fifth, the antibody repertoire against the epitopes in the head is much more diverse than the antibody repertoire against the epitopes in the stem [33 ].…”

Phage and Yeast Display

“…While these antibodies are capable of binding HA proteins across multiple viral subtypes, globular head-directed mAbs tend to be less heteroreactive, given the lack of strict conservation among these epitopes. Studies involving phage library selections yielded headdirected mAbs capable of neutralizing single or multiple subtypes, including H1N1, H2N2, and H3N2 viruses (53)(54). Despite this limited scope of reactivity, mAbs that are neutralizing to the globular head are typically more potent, a trend likely due to epitope availability on the viral surface (41).…”