Structural basis for norovirus neutralization by an HBGA blocking human IgA antibody

Abstract

Human noroviruses (HuNoVs) cause sporadic and epidemic gastroenteritis worldwide. They are classified into two major genogroups (GI and GII), with each genogroup further divided into multiple genotypes. Susceptibility to these viruses is influenced by genetically determined histo-blood group antigen (HBGA) expression. HBGAs function as cell attachment factors by binding to a surface-exposed region in the protruding (P) domain of the capsid protein. Sequence variations in this region that result in differential HBGA binding patterns and antigenicity are suggested to form a basis for strain diversification. Recent studies show that serum antibodies that block HBGA binding correlate with protection against illness. Although genogroup-dependent variation in HBGA binding specificity is structurally well characterized, an understanding of how antibodies block HBGA binding and how genotypic variations affect such blockade is lacking. Our crystallographic studies of the GI.1 P domain in complex with the Fab fragment of a human IgA monoclonal antibody (IgA 5I2) with HBGA blocking activity show that the antibody recognizes a conformational epitope formed by two surface-exposed loop clusters in the P domain. The antibody engulfs the HBGA binding site but does not affect its structural integrity. An unusual feature of the antigen recognition by IgA 5I2 is the predominant involvement of the CDR light chain 1 in contrast to the commonly observed CDR heavy chain 3, providing a unique perspective into antibody diversity in antigen recognition. Identification of the antigenic site in the P domain shows how genotypic variations might allow escape from antibody neutralization and exemplifies the interplay between antigenicity and HBGA specificity in HuNoV evolution.norovirus | HBGA-blockade antibody | crystal structure | antibody neutralization | viral entry H uman noroviruses (NoVs; HuNoVs) are the leading cause of viral gastroenteritis. They are associated with almost one fifth of all cases of acute gastroenteritis worldwide (1). It is estimated that ∼200,000 children under the age of 5 y die annually from HuNoV infections (2). Currently, there are no licensed vaccines or antiviral agents to treat the disease, although vaccine candidates are being investigated (3, 4). Development of efficient vaccines is limited by a lack of understanding of the immune correlates of protection and rapid evolution of NoVs based on antigenic variations and differential glycan binding.NoVs are nonenveloped positive-strand RNA viruses belonging to the family Caliciviridae. They are phylogenetically classified into at least six genogroups (GI-GVI), with each genogroup divided into several genotypes. Genogroups GI, GII, and GIV contain human pathogens (5, 6). The prototype Norwalk virus (NV) is classified as genogroup I genotype 1 (i.e., GI.1). NoVs belonging to genotype GII.4 are the most prevalent and are associated with ∼70% of all HuNoV infections (7). HuNoVs recognize and bind to histo-blood group antigens (HBGAs) as receptors/coreceptors for cel...

Nanobodies Targeting Norovirus Capsid Reveal Functional Epitopes and Potential Mechanisms of Neutralization

“…Four α-GI mAbs isolated from chimpanzees challenged with norovirus blocked VLP binding to carbohydrates and inhibited hemagglutination, although their precise binding sites were not described [20]. Recently, a GI.1 specific mAb was discovered that sterically hindered the HBGA pocket [22]. In our study, we showed that Nano-14 overlapped with the GII.10 HBGA binding sites and inhibited HBGA binding by steric interference and competition for the pocket.…”

“…For example, in the case of HIV, with the aid of an extra long CDR3 loop, the neutralizing Nanobody D7 effectively competed for the CD4 binding site on gp120 protein [42]. Previously described Nanobodies and mAbs with therapeutic potential against human norovirus were also proposed to interfere with the HBGA binding site [20,[22][23][24][25][26][27]. MAb termed NV8812 bound to a conformational epitope on the GI.1 P domain and blocked the binding of norovirus VLPs to human and animal cell lines [24].…”

“…Moreover, in a recent enteroid norovirus replication system inhibition in the blocking assay was correlated with neutralization in cell culture. A recent study suggested that antibodies targeting the HBGA pocket could inhibit norovirus replication by steric interference with the GI.1 HBGA pocket [22]. A number of other studies have identified norovirus-specific monoclonal antibodies (mAbs) and single chain variable domains (VHH or Nanobodies) that could block norovirus VLP binding to HBGAs [20,[23][24][25][26][27].…”

Antigen recognition by single-domain antibodies: structural latitudes and constraints

“…81 Likewise, V H Hs targeting the HBGA-binding pocket of norovirus VP1 neutralized a broad range of genotypes, 82 while larger conventional antibodies also made contact with antigenically variable residues surrounding the HBGA pocket and were thus strain-specific. 83 …”

Structural features of glycan recognition among viral pathogens

“…Recently the structure of one such human antibody in complex with the NV P-domain was determined and it revealed steric hindrance as the mechanism of HBGA blockade [23]. Other structural studies have shown that human milk oligosaccharides, 2′-fucosyllactose (2′FL) and 3′-fucosyllactose (3′FL) and molecules like citrate mimic HBGA binding, thus serving as decoy receptors and potential glycomimetics [24,25].…”