Structural and Functional Studies of Herpes Simplex Virus Glycoprotein D

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). The vaccine consisted of a soluble form of herpes simplex virus 2 (HSV-2) glycoprotein D (gD2) with adjuvant. The goal of the current study was to examine the composition of the humoral response to gD2 within a selected subset of vaccinated individuals. Serum samples from 30 vaccine recipients were selected based upon relative enzyme-linked immunosorbent assay (ELISA) titers against gD2; 10 samples had high titers, 10 had medium titers, and the remaining 10 had low ELISA titers. We employed a novel, biosensor-based monoclonal antibody (MAb)-blocking assay to determine whether gD2 vaccination elicited IgG responses against epitopes overlapping those of well-characterized MAbs. Importantly, IgGs from the majority of gD2-immunized subjects competed for gD binding with four antigenically distinct virus-neutralizing MAbs (MC2, MC5, MC23, and DL11). Screening of patient IgGs against overlapping peptides spanning the gD2 ectodomain revealed that about half of the samples contained antibodies against linear epitopes within the N and C termini of gD2. We found that the virus-neutralizing abilities of the 10 most potent samples correlated with overall gD-binding activity and to an even greater extent with the combined content of IgGs against the epitopes of MAbs MC2, MC5, MC23, and DL11. This suggests that optimal virus-neutralizing activity is achieved by strong and balanced responses to the four major discontinuous neutralizing epitopes of gD2. IMPORTANCESeveral herpes simplex virus 2 (HSV-2) subunit vaccine studies have been conducted in human subjects using a recombinant form of HSV-2 glycoprotein D (gD2). Although several distinct, well-characterized virus-neutralizing epitopes on gD2 are targeted by murine monoclonal antibodies, it is not known whether the same epitopes are targeted by the humoral response to gD2 in humans. We have developed a novel, biosensor-based competition assay to directly address this important question. Using this approach, we identified epitopes that elicit strong humoral responses in humans, as well as other epitopes that elicit much weaker responses. These data provide new insight into the human response to known neutralizing gD2 epitopes and reveal characteristics of this response that may guide future vaccine development.

Section: History Of Samplesmentioning

confidence: 99%

“…1B) and in complex with both HVEM and nectin-1 has been solved (15)(16)(17). In addition, because of its ability to induce potent virus-neutralizing antibodies, the antigenic structure of gD has been extensively studied (18,19) using a large panel of anti-gD MAbs (Fig. 1A).…”

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confidence: 99%

Repertoire of Epitopes Recognized by Serum IgG from Humans Vaccinated with Herpes Simplex Virus 2 Glycoprotein D

Whitbeck

Huang

Cairns

et al. 2014

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“…Also, gD-1(⌬290-299t) and wild-type gDt competed for binding to HVEMt. It is well established that gD can induce potent virus-neutralizing antibodies (Abs) (7,9,25,34,40,43,49). Monoclonal Abs (MAbs) against gD have been demonstrated to block a postadsorption step in virus entry prior to virus-cell fusion (20,21,25,52).…”

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confidence: 99%

Monoclonal Antibodies to Distinct Sites on Herpes Simplex Virus (HSV) Glycoprotein D Block HSV Binding to HVEM

Nicola

Leon

et al. 1998

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124

HVEM (for herpesvirus entry mediator) is a member of the tumor necrosis factor receptor superfamily and mediates entry of many strains of herpes simplex virus (HSV) into normally nonpermissive Chinese hamster ovary (CHO) cells. We used sucrose density centrifugation to demonstrate that purified HSV-1 KOS virions bind directly to a soluble, truncated form of HVEM (HVEMt) in the absence of any other cell-associated components. Therefore, HVEM mediates HSV entry by serving as a receptor for the virus. We previously showed that soluble, truncated forms of HSV glycoprotein D (gDt) bind to HVEMt in vitro. Here we show that antibodies specific for gD, but not the other entry glycoproteins gB, gC, or the gH/gL complex, completely block HSV binding to HVEM. Thus, virion gD is the principal mediator of HSV binding to HVEM. To map sites on virion gD which are necessary for its interaction with HVEM, we preincubated virions with gD-specific monoclonal antibodies (MAbs). MAbs that recognize antigenic sites Ib and VII of gD were the only MAbs which blocked the HSV-HVEM interaction. MAbs from these two groups failed to coprecipitate HVEMt in the presence of soluble gDt, whereas the other anti-gD MAbs coprecipitated HVEMt and gDt. Previous mapping data indicated that site VII includes amino acids 11 to 19 and site Ib includes 222 to 252. The current experiments indicate that these sites contain residues important for HSV binding to HVEM. Group Ib and VII MAbs also blocked HSV entry into HVEM-expressing CHO cells. These results suggest that the mechanism of neutralization by these MAbs is via interference with the interaction between gD in the virus and HVEM on the cell. Group Ia and II MAbs failed to block HSV binding to HVEM yet still neutralized HVEM-mediated entry, suggesting that these MAbs block entry at a step other than HVEM binding.

“…Using a complementation assay, it was shown that gD-2 can replace gD-1 in the infection process (36), also suggesting that the two glycoproteins interact with the same receptor on Vero cells. Both proteins induce production of neutralizing antibody and stimulate a cross-protective immune response in animal models (reviewed in references 3,4,10,30,35).…”

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confidence: 99%

High-level expression and purification of secreted forms of herpes simplex virus type 1 glycoprotein gD synthesized by baculovirus-infected insect cells

Sisk

Bradley

Leipold

et al. 1994

107

Two forms of herpes simplex virus glycoprotein gD were recombined into Autographa californica nuclear polyhedrosis virus (baculovirus) and expressed in infected Spodoptera Jfugiperda (Sf9) cells. Each protein was truncated at residue 306 of mature gD. One form, gD-1(306t), contains the coding sequence of Patton strain herpes simplex virus type 1 gD; the other, gD-1(QAAt), contains three mutations which eliminate all signals for addition of N-linked oligosaccharides. Prior to recombination, each gene was cloned into the baculovirus transfer vector pVT-Bac, which permits insertion of the gene minus its natural signal peptide in frame with the signal peptide of honeybee melittin. As is the case with many other baculovirus transfer vectors, pVT-Bac also