Monoclonal Antibodies That Bind to Domain III of Dengue Virus E Glycoprotein Are the Most Efficient Blockers of Virus Adsorption to Vero Cells

Abstract

The specific mechanisms by which antibodies neutralize flavivirus infectivity are not completely understood. To study these mechanisms in more detail, we analyzed the ability of a well-defined set of anti-dengue (DEN) virus E-glycoprotein-specific monoclonal antibodies (MAbs) to block virus adsorption to Vero cells. In contrast to previous studies, the binding sites of these MAbs were localized to one of three structural domains (I, II, and III) in the E glycoprotein. The results indicate that most MAbs that neutralize virus infectivity do so, at least in part, by the blocking of virus adsorption. However, MAbs specific for domain III were the strongest blockers of virus adsorption. These results extend our understanding of the structure-function relationships in the E glycoprotein of DEN virus and provide the first direct evidence that domain III encodes the primary flavivirus receptor-binding motif.The flavivirus E glycoprotein is the primary antigen inducing protective immunity, is essential for membrane fusion, and mediates binding to cellular receptors. Therefore, this protein directly affects host range, cellular tropism, and, in part, the virulence of these viruses (17, 18). The crystal structure of the ectodomain of the tick-borne encephalitis (TBE) virus E-glycoprotein homodimer was recently solved at high resolution (16). Multiple lines of evidence indicate that this E-glycoprotein structure is strongly conserved across the Flaviviridae (16). This protein contains three structural domains. The central domain, domain I (DI), contains predominately type-specific nonneutralizing epitopes and is theorized to be the molecular hinge region involved in low-pH-triggered conformational changes (19). The dimerization domain, domain II (DII), makes important contacts with itself in the homodimer, is involved in virus-mediated membrane fusion, and contains many cross-reactive epitopes eliciting neutralizing and nonneutralizing monoclonal antibodies (MAbs) (16,19). Domain III (DIII) is characterized by an immunoglobulin-like structure containing the most distal projecting loops from the virion surface. It contains multiple type-and subtype-specific epitopes eliciting only virus-neutralizing MAbs and has been hypothesized to contain the host cell-binding antireceptor (16,18,19). As part of our ongoing research to elucidate the structure-function relationships of the dengue (DEN) virus E glycoprotein, we have assessed the ability of a well-characterized panel of E-glycoprotein-specific MAbs to block virus adsorption to Vero cells. These results provide the first direct evidence that E glycoprotein DIII encodes a receptor-binding motif.

Interaction of West Nile Virus with αvβ3 Integrin Mediates Virus Entry into Cells

“…Ϫ, infectious titre of 10 2 PFU/ml or less; ϩ, infectious titer between 10 3 and 10 5 PFU/ml; ϩϩ, infectious titer of 10 6 PFU/ml and above. domain III of the envelope protein are noted to be the strongest blockers of virus entry (9). Numerous neutralization epitopes for WNV have also been mapped within domain III of the envelope protein (25).…”

Preparation and immunogenic properties of a recombinant West Nile subunit vaccine

“…Antibodies directed against particular epitopes of the envelope protein are capable of virus neutralization, i.e., the inhibition of virus infection of susceptible cells in vitro. The dominant neutralizing epitopes have been mapped to one of three domains of the envelope protein, domain III, using monoclonal antibodies for dengue virus [9], as well as JEV [10] and WNV [11]. Additional neutralizing epitopes have recently been identified on domains I and II of the envelope protein as well [12].…”

Mapping to completeness and transplantation of a group-specific, discontinuous, neutralizing epitope in the envelope protein of dengue virus

“…Numerous data indicate that ED3 plays an important role in the infectious process (Hung et al, 2004;Se-Thoe et al, 2000;Thullier et al, 2001). In particular, many antibodies that are specific for ED3 are neutralizing (Crill & Roehrig, 2001;Roehrig, 2003). Of note, their epitopes are generally sensitive to elimination of the disulfide bond (Roehrig et al, 2004).…”