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.

FIG. 1

Representative dose-dependent blocking of adsorption by a DIII MAb, 9D12, in the pre- and postadsorption assays.

FIG. 2

Comparison of percent blocking of adsorption (±95% CI) by representative MAbs for the three distinct E-glycoprotein domains. (A) DI MAb 1B4C-2; (B) DII MAb 4E5; (C) DIII MAb 9D12; (D) DII MAb 1A5D-1; (E) DEN-2 virus polyclonal murine HIAF; (F) DEN-1-specific negative-control MAb 1F1. Panel D illustrates enhancement of virus adsorption.

FIG. 3

Summary of biological activities and spatial arrangements of the DEN-2 virus E-glycoprotein epitopes listed in Table 1. The biological activities of MAbs elicited by these epitopes were hemagglutination inhibition (HI) and virus neutralization (N). The maximal values for the blocking of virus adsorption are indicated as follows: black, greater than 40%; dark gray, 30 to 40%; light gray, 20 to 30%; white, less than 20% (for A2) or not tested (for A4, C2, C3 and C4); cross-hatched, binding enhancement. Overlapping circles indicate spatially proximal epitopes. Epitope designations: A, DII; B, DIII; C, DI. The HA and N activities of these epitopes were reported previously (19).