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

2007 ◽  
Vol 81 (23) ◽  
pp. 12816-12826 ◽  
Author(s):  
Soila Sukupolvi-Petty ◽  
S. Kyle Austin ◽  
Whitney E. Purtha ◽  
Theodore Oliphant ◽  
Grant E. Nybakken ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Surface Display ◽  
Viral Envelope ◽  
Yeast Surface Display ◽  
Neutralizing Activity ◽  
Denv Serotypes ◽  
E Protein ◽  
Lateral Ridge ◽  
Domain Iii

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.

scholarly journals Genotype-Specific Neutralization and Protection by Antibodies against Dengue Virus Type 3

2010 ◽  
Vol 84 (20) ◽  
pp. 10630-10643 ◽  
Author(s):  
James D. Brien ◽  
S. Kyle Austin ◽  
Soila Sukupolvi-Petty ◽  
Katie M. O'Brien ◽  
Syd Johnson ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Surface Display ◽  
Viral Envelope ◽  
Yeast Surface Display ◽  
E Protein ◽  
Lateral Ridge ◽  
Dengue Virus Type 3 ◽  
Domain Iii ◽  
Positive Strand Rna ◽  
Type 3

ABSTRACT Dengue viruses (DENV) comprise a family of related positive-strand RNA viruses that infect up to 100 million people annually. Currently, there is no approved vaccine or therapy to prevent infection or diminish disease severity. Protection against DENV is associated with the development of neutralizing antibodies that recognize the viral envelope (E) protein. Here, with the goal of identifying monoclonal antibodies (MAbs) that can function as postexposure therapy, we generated a panel of 82 new MAbs against DENV-3, including 24 highly neutralizing MAbs. Using yeast surface display, we localized the epitopes of the most strongly neutralizing MAbs to the lateral ridge of domain III (DIII) of the DENV type 3 (DENV-3) E protein. While several MAbs functioned prophylactically to prevent DENV-3-induced lethality in a stringent intracranial-challenge model of mice, only three MAbs exhibited therapeutic activity against a homologous strain when administered 2 days after infection. Remarkably, no MAb in our panel protected prophylactically against challenge by a strain from a heterologous DENV-3 genotype. Consistent with this, no single MAb neutralized efficiently the nine different DENV-3 strains used in this study, likely because of the sequence variation in DIII within and between genotypes. Our studies suggest that strain diversity may limit the efficacy of MAb therapy or tetravalent vaccines against DENV, as neutralization potency generally correlated with a narrowed genotype specificity.

scholarly journals Structure and Function Analysis of Therapeutic Monoclonal Antibodies against Dengue Virus Type 2

2010 ◽  
Vol 84 (18) ◽  
pp. 9227-9239 ◽  
Author(s):  
Soila Sukupolvi-Petty ◽  
S. Kyle Austin ◽  
Michael Engle ◽  
James D. Brien ◽  
Kimberly A. Dowd ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Dengue Virus ◽  
Function Analysis ◽  
Surface Display ◽  
Viral Envelope ◽  
Yeast Surface Display ◽  
Protective Capacity ◽  
Diverse Range ◽  
Lateral Ridge

ABSTRACT Dengue virus (DENV) is the most prevalent insect-transmitted viral disease in humans globally, and currently no specific therapy or vaccine is available. Protection against DENV and other related flaviviruses is associated with the development of antibodies against the viral envelope (E) protein. Although prior studies have characterized the neutralizing activity of monoclonal antibodies (MAbs) against DENV type 2 (DENV-2), none have compared simultaneously the inhibitory activity against a genetically diverse range of strains in vitro, the protective capacity in animals, and the localization of epitopes. Here, with the goal of identifying MAbs that can serve as postexposure therapy, we investigated in detail the functional activity of a large panel of new anti-DENV-2 mouse MAbs. Binding sites were mapped by yeast surface display and neutralization escape, cell culture inhibition assays were performed with homologous and heterologous strains, and prophylactic and therapeutic activity was evaluated with two mouse models. Protective MAbs localized to epitopes on the lateral ridge of domain I (DI), the dimer interface, lateral ridge, and fusion loop of DII, and the lateral ridge, C-C′ loop, and A strand of DIII. Several MAbs inefficiently inhibited at least one DENV-2 strain of a distinct genotype, suggesting that recognition of neutralizing epitopes varies with strain diversity. Moreover, antibody potency generally correlated with a narrowed genotype and serotype specificity. Five MAbs functioned efficiently as postexposure therapy when administered as a single dose, even 3 days after intracranial infection of BALB/c mice. Overall, these studies define the structural and functional complexity of antibodies against DENV-2 with protective potential.

scholarly journals Dengue virus envelope domain III immunization elicits predominantly cross-reactive, poorly neutralizing antibodies localized to the AB loop: implications for dengue vaccine design

2013 ◽  
Vol 94 (10) ◽  
pp. 2191-2201 ◽  
Author(s):  
Xiao-Quan Li ◽  
Li-Wen Qiu ◽  
Yue Chen ◽  
Kun Wen ◽  
Jian-Piao Cai ◽  
...  
Keyword(s):  
Antibody Response ◽  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Surface Display ◽  
Yeast Surface Display ◽  
Protein Domain ◽  
Dengue Vaccine ◽  
Denv Serotypes ◽  
Domain Iii

Dengue virus (DENV) is a mosquito-borne virus that causes severe health problems. An effective tetravalent dengue vaccine candidate that can provide life-long protection simultaneously against all four DENV serotypes is highly anticipated. A better understanding of the antibody response to DENV envelope protein domain III (EDIII) may offer insights into vaccine development. Here, we identified 25 DENV cross-reactive mAbs from immunization with Pichia pastoris-expressed EDIII of a single or all four serotype(s) using a prime–boost protocol, and through pepscan analysis found that 60 % of them (15/25) specifically recognized the same highly conserved linear epitope aa 309–320 of EDIII. All 15 complex-reactive mAbs exhibited significant cross-reactivity with recombinant EDIII from all DENV serotypes and also with C6/36 cells infected with DENV-1, -2, -3 and -4. However, neutralization assays indicated that the majority of these 15 mAbs were either moderately or weakly neutralizing. Through further epitope mapping by yeast surface display, two residues in the AB loop, Q316 and H317, were discovered to be critical. Three-dimensional modelling analysis suggests that this epitope is surface exposed on EDIII but less accessible on the surface of the E protein dimer and trimer, especially on the surface of the mature virion. It is concluded that EDIII as an immunogen may elicit cross-reactive mAbs toward an epitope that is not exposed on the virion surface, therefore contributing inefficiently to the mAbs neutralization potency. Therefore, the prime–boost strategy of EDIII from a single serotype or four serotypes mainly elicited a poorly neutralizing, cross-reactive antibody response to the conserved AB loop of EDIII.

scholarly journals Antibody Recognition and Neutralization Determinants on Domains I and II of West Nile Virus Envelope Protein

2006 ◽  
Vol 80 (24) ◽  
pp. 12149-12159 ◽  
Author(s):  
Theodore Oliphant ◽  
Grant E. Nybakken ◽  
Michael Engle ◽  
Qing Xu ◽  
Christopher A. Nelson ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Surface Display ◽  
Yeast Surface Display ◽  
Virus Envelope ◽  
West Nile ◽  
Lower Efficiency ◽  
E Protein ◽  
Lateral Ridge ◽  
Fusion Loop ◽  
Neutralizing Epitopes

ABSTRACT Previous studies have demonstrated that monoclonal antibodies (MAbs) against an epitope on the lateral surface of domain III (DIII) of the West Nile virus (WNV) envelope (E) strongly protect against infection in animals. Herein, we observed significantly less efficient neutralization by 89 MAbs that recognized domain I (DI) or II (DII) of WNV E protein. Moreover, in cells expressing Fc γ receptors, many of the DI- and DII-specific MAbs enhanced infection over a broad range of concentrations. Using yeast surface display of E protein variants, we identified 25 E protein residues to be critical for recognition by DI- or DII-specific neutralizing MAbs. These residues cluster into six novel and one previously characterized epitope located on the lateral ridge of DI, the linker region between DI and DIII, the hinge interface between DI and DII, and the lateral ridge, central interface, dimer interface, and fusion loop of DII. Approximately 45% of DI-DII-specific MAbs showed reduced binding with mutations in the highly conserved fusion loop in DII: 85% of these (34 of 40) cross-reacted with the distantly related dengue virus (DENV). In contrast, MAbs that bound the other neutralizing epitopes in DI and DII showed no apparent cross-reactivity with DENV E protein. Surprisingly, several of the neutralizing epitopes were located in solvent-inaccessible positions in the context of the available pseudoatomic model of WNV. Nonetheless, DI and DII MAbs protect against WNV infection in mice, albeit with lower efficiency than DIII-specific neutralizing MAbs.

scholarly journals Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

Virology ◽  
2013 ◽  
Vol 441 (2) ◽  
pp. 114-125 ◽  
Author(s):  
John T. Roehrig ◽  
Siritorn Butrapet ◽  
Nathan M. Liss ◽  
Susan L. Bennett ◽  
Betty E. Luy ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Heparan Sulfate ◽  
Membrane Fusion ◽  
Binding Sites ◽  
Envelope Protein ◽  
Vero Cells ◽  
Lateral Ridge ◽  
Domain Iii ◽  
Dengue Virus Type 2

scholarly journals Cryo-EM structure of an antibody that neutralizes dengue virus type 2 by locking E protein dimers

Science ◽  
2015 ◽  
Vol 349 (6243) ◽  
pp. 88-91 ◽  
Author(s):  
Guntur Fibriansah ◽  
Kristie D. Ibarra ◽  
Thiam-Seng Ng ◽  
Scott A. Smith ◽  
Joanne L. Tan ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Secondary Infection ◽  
Human Monoclonal Antibody ◽  
Viral Envelope ◽  
Severe Dengue ◽  
Virus Surface ◽  
Dengue Disease ◽  
Denv Serotypes ◽  
E Protein ◽  
Protein Dimers

There are four closely-related dengue virus (DENV) serotypes. Infection with one serotype generates antibodies that may cross-react and enhance infection with other serotypes in a secondary infection. We demonstrated that DENV serotype 2 (DENV2)–specific human monoclonal antibody (HMAb) 2D22 is therapeutic in a mouse model of antibody-enhanced severe dengue disease. We determined the cryo–electron microscopy (cryo-EM) structures of HMAb 2D22 complexed with two different DENV2 strains. HMAb 2D22 binds across viral envelope (E) proteins in the dimeric structure, which probably blocks the E protein reorganization required for virus fusion. HMAb 2D22 “locks” two-thirds of or all dimers on the virus surface, depending on the strain, but neutralizes these DENV2 strains with equal potency. The epitope defined by HMAb 2D22 is a potential target for vaccines and therapeutics.

scholarly journals Monoclonal Antibodies That Bind to Common Epitopes on the Dengue Virus Type 2 Nonstructural-1 and Envelope Glycoproteins Display Weak Neutralizing Activity and Differentiated Responses to Virulent Strains: Implications for Pathogenesis and Vaccines

2007 ◽  
Vol 15 (3) ◽  
pp. 549-561 ◽  
Author(s):  
Andrew K. I. Falconar
Keyword(s):  
Monoclonal Antibodies ◽  
Dengue Virus ◽  
Neutralizing Activity ◽  
Denv Serotypes ◽  
Cross Reactions ◽  
Virulent Strains ◽  
Highly Virulent ◽  
Dengue Virus Type 2

ABSTRACT The abilities of monoclonal antibodies (MAbs) that bind to defined sequential epitopes on the dengue virus (DENV) nonstructural-1 (NS1) glycoproteins to cross-react with epitopes on the DENV envelope (E) glycoproteins were investigated. In this study, some of these MAbs cross-reacted with the DENV type 2 (DENV-2) E glycoprotein and with synthetic peptides representing X-ray crystallographically confirmed surface-exposed regions on this glycoprotein. MAb 1G5.3 cross-reacted with the flavivirus-conserved 101-WGNGCGLFG-109 fusion sequence, the 273-SSGNL-277 DENV-2 hinge region sequence, and the 156-GKHGKEIKIT-165 sequence of virulent DENV-2 strains. MAb 1G5.4-A1-C3 cross-reacted with the 67-NTTT ESR CPT-76 and 156-GKHGK EIK IT-165 sequences of virulent DENV-2 strains, the 338-EIMDL DNR HV-347 sequence from a highly virulent DENV-2 (M2) strain, and two epitopes on a virulent DENV-3 strain (288-KMD KLELK G-296 and 323- RVEYRGE DAP-332), which all contained target ELK/KLE-type motifs (underlined). These MAbs showed reduced cross-reactions with the corresponding sequences from weakly pathogenic strains of all four DENV serotypes and had either no (MAb 1G5.4-A1-C3) or weak (MAb 1G5.3) neutralizing activity against them. MAb 1G5.3 more strongly neutralized DENV-2 strains with higher pathogenic capacities, while MAb 1G5.4-A1-C3 showed increasing neutralizing titers against the virulent DENV-3 strain and the moderately virulent and highly virulent (M2) DENV-2 strains. These cross-reactions with the E glycoprotein accord with the observation that MAb 1G5.3 caused dramatic and lethal antibody-enhanced replication (AER) of a DENV-2 strain in vivo. Together with in vivo AER studies of these DENV strains using MAb 1G5.4-A1-C3, these results may account for the increased pathogenic capacities of such strains, which is likely to have important implications for pathogenesis and vaccines.

scholarly journals Residues in domain III of the dengue virus envelope glycoprotein involved in cell-surface glycosaminoglycan binding

2012 ◽  
Vol 93 (1) ◽  
pp. 72-82 ◽  
Author(s):  
Daniel Watterson ◽  
Bostjan Kobe ◽  
Paul R. Young
Keyword(s):  
Cell Surface ◽  
Dengue Virus ◽  
Site Directed Mutagenesis ◽  
Cell Surface Receptor ◽  
Protein Fusion ◽  
Virus Envelope ◽  
Putative Receptor ◽  
E Protein ◽  
Surface Receptor ◽  
Domain Iii

The dengue virus (DENV) envelope (E) protein mediates virus entry into cells via interaction with a range of cell-surface receptor molecules. Cell-surface glycosaminoglycans (GAGs) have been shown to play an early role in this interaction, and charged oligosaccharides such as heparin bind to the E protein. We have examined this interaction using site-directed mutagenesis of a recombinant form of the putative receptor-binding domain III of the DENV-2E protein expressed as an MBP (maltose-binding protein)-fusion protein. Using an ELISA-based GAG-binding assay, cell-based binding analysis and antiviral-activity assays, we have identified two critical residues, K291 and K295, that are involved in GAG interactions. These studies have also demonstrated differential binding between mosquito and human cells.

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