Type- and Subcomplex-Specific Neutralizing Antibodies against Domain III of Dengue Virus Type 2 Envelope Protein Recognize Adjacent Epitopes

Abstract

Neutralization of flaviviruses in vivo correlates with the development of an antibody response against the viral envelope (E) protein. Previous studies demonstrated that monoclonal antibodies (MAbs) against an epitope on the lateral ridge of domain III (DIII) of the West Nile virus (WNV) E protein strongly protect against infection in animals.Based on X-ray crystallography and sequence analysis, an analogous type-specific neutralizing epitope for individual serotypes of the related flavivirus dengue virus (DENV) was hypothesized. Using yeast surface display of DIII variants, we defined contact residues of a panel of type-specific, subcomplex-specific, and cross-reactive MAbs that recognize DIII of DENV type 2 (DENV-2) and have different neutralizing potentials. Type-specific MAbs with neutralizing activity against DENV-2 localized to a sequence-unique epitope on the lateral ridge of DIII, centered at the FG loop near residues E383 and P384, analogous in position to that observed with WNV-specific strongly neutralizing MAbs. Subcomplex-specific MAbs that bound some but not all DENV serotypes and neutralized DENV-2 infection recognized an adjacent epitope centered on the connecting A strand of DIII at residues K305, K307, and K310. In contrast, several MAbs that had poor neutralizing activity against DENV-2 and cross-reacted with all DENV serotypes and other flaviviruses recognized an epitope with residues in the AB loop of DIII, a conserved region that is predicted to have limited accessibility on the mature virion. Overall, our experiments define adjacent and structurally distinct epitopes on DIII of DENV-2 which elicit type-specific, subcomplex-specific, and cross-reactive antibodies with different neutralizing potentials.Dengue fever (DF), the most prevalent arthropod-borne viral illness in humans, is caused by dengue virus (DENV). The four serotypes of DENV are transmitted to humans primarily by the mosquitoes Aedes aegypti and Aedes albopictus. DENV is a member of the Flaviviridae family and is related to the viruses that cause yellow fever and the Japanese, St. Louis, and West Nile encephalitides (8). Infection by DENV causes a spectrum of clinical disease, ranging from an acute, debilitating, selflimited febrile illness (DF) to a life-threatening hemorrhagic and capillary leak syndrome (dengue hemorrhagic fever/dengue shock syndrome). At present, no approved antiviral treatment or vaccine is available, and therapy is supportive in nature. DENV causes an estimated 25 to 100 million cases of DF and 250,000 cases of dengue hemorrhagic fever per year worldwide, with 2.5 billion people at risk for infection (27,48).DENV is an enveloped virus with a single-stranded, positivesense RNA genome (11). The 10.7-kilobase genome is translated as a single polyprotein, which is then cleaved into three structural proteins (C, prM/M, and E) and seven nonstructural (NS) proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) by virus-and host-encoded proteases. The 500-Å DENV mature virion has a well-organized outer protein she...

Plasmablasts Generated during Repeated Dengue Infection Are Virus Glycoprotein–Specific and Bind to Multiple Virus Serotypes

“…3B), and no binding to prM was observed, with the exception of the pooled human serum that was used as a positive control. All mAb clones were also tested for binding to EDIII, the E protein domain that contains the most effective neutralizing epitopes (34,35). However, none of the Abs tested bound to EDIII, at least not to the soluble monomeric form used in these assays (Table II).…”

Functional analysis of dengue virus (DENV) type 2 envelope protein domain 3 type-specific and DENV complex-reactive critical epitope residues

“…Extensive antibody mapping studies using mouse mAbs have identified at least 12 different epitopes on the surface of the E protein, with those located on ED3 eliciting some of the most potent neutralizing antibodies Roehrig et al, 1998). Furthermore, two major overlapping antigenic sites have been identified on ED3 and include the DENV type-specific and DEN complex-reactive antigenic sites Gromowski et al, 2008;Roehrig et al, 1998;Sukupolvi-Petty et al, 2007). Epitope mapping studies have demonstrated that these sites are composed of a small number of critically important residues (i.e.…”

“…Epitope mapping studies have demonstrated that these sites are composed of a small number of critically important residues (i.e. critical residues) that significantly reduce antibody binding to recombinant ED3 when mutated Gromowski et al, 2008;Hiramatsu et al, 1996;Sukupolvi-Petty et al, 2007). Critical residues composing the DENV-2 type-specific antigenic site are lysine at position 305 and proline at position 384 (Figs 1a and S1, available in the online Supplementary Material), whereas, the DEN complexreactive antigenic site is composed of one critical residue, lysine at 310 (Fig.…”

Molecular Docking Studies of Selected Medicinal Drugs as Dengue Virus-2 Protease Inhibitors

“…Dengue is an infection caused by the dengue virus (DENV) of the Flaviviridae family (Sukupolvi-Petty et al 2007) and is transmitted by infected female mosquitoes (Bhatt et al 2013;Kyle & Harris 2006). Dengue virus has four serotypes (DENV-1 to -4) that genetically share ~65% similarities (Mustafa et al 2015).…”