The prefusion structure of herpes simplex virus glycoprotein B.
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ABSTRACT:
SUBMITTER: Vollmer B
PROVIDER: S-EPMC7518877 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Binding of herpes simplex virus (HSV) glycoprotein D (gD) to a cell surface receptor is required to trigger membrane fusion during entry into host cells. Nectin-1 is a cell adhesion molecule and the main HSV receptor in neurons and epithelial cells. We report the structure of gD bound to nectin-1 determined by x-ray crystallography to 4.0 Å resolution. The structure reveals that the nectin-1 binding site on gD differs from the binding site of the HVEM receptor. A surface on the first Ig-domain of nectin-1, which mediates homophilic interactions of Ig-like cell adhesion molecules, buries an area composed by residues from both the gD N- and C-terminal extensions. Phenylalanine 129, at the tip of the loop connecting β-strands F and G of nectin-1, protrudes into a groove on gD, which is otherwise occupied by C-terminal residues in the unliganded gD and by N-terminal residues in the gD/HVEM complex. Notably, mutation of Phe129 to alanine prevents nectin-1 binding to gD and HSV entry. Together these data are consistent with previous studies showing that gD disrupts the normal nectin-1 homophilic interactions. Furthermore, the structure of the complex supports a model in which gD-receptor binding triggers HSV entry through receptor-mediated displacement of the gD C-terminal region.
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Project description:The worldwide prevalence of herpes simplex virus (HSV) and the shortage of efficient vaccines and novel therapeutic strategies against HSV are widely global concerns. The abundance on the virion and the major stimulus for the virus-neutralizing antibodies makes gD a predominant candidate for cure of HSV infection. In this study, we generated a monoclonal antibody (mAb), termed m27f, targeting to glycoprotein D (gD) of HSV-2, which also has cross-reactivity against HSV-1 gD. It has a high level of neutralizing activity against both HSV-1 and HSV-2, and binds to a highly conserved region (residues 292-297) within the pro-fusion domain of gD. It can effectively block HSV cell-to-cell spread in vitro. The pre- or post-attachment neutralization assay and syncytium formation inhibition assay revealed that m27f neutralizes HSV at the post-binding stage. Moreover, therapeutic administration of m27f completely prevented infection-related mortality of mice challenged with a lethal dose of HSV-2. Our newly identified epitope for the neutralizing antibody would facilitate studies of gD-based HSV entry or vaccine design, and m27f itself demonstrated a high potential for adaptation as a protective or therapeutic drug against HSV.
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Project description:Herpes simplex virus (HSV) uses the cell adhesion molecule nectin-1 as a receptor to enter neurons and epithelial cells. The viral glycoprotein D (gD) is used as a non-canonical ligand for nectin-1. The gD binding site on nectin-1 overlaps with a functional adhesive site involved in nectin-nectin homophilic trans-interaction. Consequently, when nectin-1 is engaged with a cellular ligand at cell junctions, the gD binding site is occupied. Here we report that HSV gD is able to disrupt intercellular homophilic trans-interaction of nectin-1 and induce a rapid redistribution of nectin-1 from cell junctions. This movement does not require the receptor's interaction with the actin-binding adaptor afadin. Interaction of nectin-1 with afadin is also dispensable for virion surfing along nectin-1-rich filopodia. Cells seeded on gD-coated surfaces also fail to accumulate nectin-1 at cell contact. These data indicate that HSV gD affects nectin-1 locally through direct interaction and more globally through signaling.
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Project description:Herpes simplex viruses display hundreds of gD glycoproteins, and yet their neutralization requires tens of thousands of antibodies per virion, leading us to ask whether a wild-type virion with just a single free gD is still infective. By quantitative analysis of fluorescently labeled virus particles and virus neutralization assays, we show that entry of a wild-type HSV virion to a cell does indeed require just one or two of the approximately 300 gD glycoproteins to be left unbound by monoclonal antibody. This indicates that HSV entry is an extraordinarily efficient process, functioning at the level of single molecular complexes.
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Project description:Four glycoproteins (gD, gB, gH, and gL) are essential for herpes simplex virus (HSV) entry into cells. An early step of fusion requires gD to bind one of several receptors, such as nectin-1 or herpesvirus entry mediator (HVEM). We hypothesize that a conformational change in gD occurs upon receptor binding that triggers the other glycoproteins to mediate fusion. Comparison of the crystal structures of gD alone and gD bound to HVEM reveals that upon HVEM binding, the gD N terminus transitions from a flexible stretch of residues to a hairpin loop. To address the contribution of this transition to the ability of gD to trigger fusion, we attempted to "lock" the gD N terminus into a looped conformation by engineering a disulfide bond at its N and C termini. The resulting mutant (gD-A3C/Y38C) failed to trigger fusion in the absence of receptor, suggesting that formation of the loop is not the sole fusion trigger. Unexpectedly, although gD-A3C/Y38C bound HVEM, it failed to bind nectin-1. This was due to the key role played by Y38 in interacting with nectin-1. Since tyrosines are often "hot spot" residues at the center of protein-protein interfaces, we mutated residues that surround Y38 on the same face of gD and tested their binding and functional properties. Our results suggest that this region of gD is important for nectin-1 interaction and is distinct from but partially overlaps the site of HVEM binding. Unique gD mutants with altered receptor usage generated in this study may help dissect the roles played by various HSV receptors during infection.
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Project description:Herpesviruses include many important human pathogens such as herpes simplex virus, cytomegalovirus, varicella-zoster virus, and the oncogenic Epstein-Barr virus and Kaposi sarcoma-associated herpesvirus. Herpes virions contain a large icosahedral capsid that has a portal at a unique 5-fold vertex, similar to that seen in the tailed bacteriophages. The portal is a molecular motor through which the viral genome enters the capsid during virion morphogenesis. The genome also exits the capsid through the portal-vertex when it is injected through the nuclear pore into the nucleus of a new host cell to initiate infection. Structural investigations of the herpesvirus portal-vertex have proven challenging, owing to the small size of the tail-like portal-vertex-associated tegument (PVAT) and the presence of the tegument layer that lays between the nucleocapsid and the viral envelope, obscuring the view of the portal-vertex. Here, we show the structure of the herpes simplex virus portal-vertex at subnanometer resolution, solved by electron cryomicroscopy (cryoEM) and single-particle 3D reconstruction. This led to a number of new discoveries, including the presence of two previously unknown portal-associated structures that occupy the sites normally taken by the penton and the Ta triplex. Our data revealed that the PVAT is composed of 10 copies of the C-terminal domain of pUL25, which are uniquely arranged as two tiers of star-shaped density. Our 3D reconstruction of the portal-vertex also shows that one end of the viral genome extends outside the portal in the manner described for some bacteriophages but not previously seen in any eukaryote viruses. Finally, we show that the viral genome is consistently packed in a highly ordered left-handed spool to form concentric shells of DNA. Our data provide new insights into the structure of a molecular machine critical to the biology of an important class of human pathogens.
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Project description:Herpes simplex virus type 1 (HSV-1) encodes 11 envelope glycoproteins, of which glycoprotein G-1 (gG-1) induces a type-specific antibody response. Variability of the gG-1 gene among wild-type strains may be a factor of importance for a reliable serodiagnosis and typing of HSV-1 isolates. Here, we used a gG-1 type-specific monoclonal antibody (MAb) to screen for mutations in the immunodominant region of this protein in 108 clinical HSV-1 isolates. Of these, 42 isolates showed no reactivity to the anti-gG-1 MAb. One hundred five strains were further examined by DNA sequencing of the middle part of the gG-1 gene, encompassing 106 amino acids including the immunodominant region and epitope of the anti-gG-1 MAb. By phylogenetic comparisons based on the sequence data, we observed two (main) genetic variants of the gG-1 gene among the clinical isolates corresponding to reactivity or nonreactivity to the anti-gG-1 MAb. Furthermore, four strains appeared to be recombinants of the two gG-1 variants. In addition, one strain displayed a gG-1-negative phenotype due to a frameshift mutation, in the form of insertion of a cytosine nucleotide. When immunoglobulin G reactivity to HSV-1 in sera from patients infected with either of the two variants was investigated, no significant differences were found between the two groups, either in a type-common enzyme-linked immunosorbent assay (ELISA) or in a type-specific gG-1 antigen-based ELISA. Despite the here-documented existence of two variants of the gG-1 gene affecting the immunodominant region of the protein, other circumstances, such as early phase of infection, might be sought for explaining the seronegativity to gG-1 commonly found in a proportion of the HSV-1-infected patients.
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