Structure of Respiratory Syncytial Virus Fusion Glycoprotein in the Postfusion Conformation Reveals Preservation of Neutralizing Epitopes

McLellan, Jason S.; Yang, Yongping; Graham, Barney S.; Kwong, Peter D.

doi:10.1128/jvi.00555-11

Cited by 354 publications

(448 citation statements)

References 46 publications

Supporting

Mentioning

427

Contrasting

Unclassified

Order By: Relevance

“…In addition, HRSV contains a small hydrophobic protein, SH, of unclear function and proteins of cellular origin, such as actin and chaperones (12). The RNP and F have been analyzed using 3D-EM and X-ray crystallography (13,14).HRSV particles are pleomorphic with both spherical and filamentous particles of different sizes (15). Early electron microscopy studies showed the presence of a helical protein layer under the membrane of filamentous HRSV (15).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Architecture of respiratory syncytial virus revealed by electron cryotomography

Liljeroos

Krzyzaniak²,

Helenius³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

177

173

View full text Add to dashboard Cite

Human respiratory syncytial virus is a human pathogen that causes severe infection of the respiratory tract. Current information about the structure of the virus and its interaction with host cells is limited. We carried out an electron cryotomographic characterization of cell culture-grown human respiratory syncytial virus to determine the architecture of the virion. The particles ranged from 100 nm to 1,000 nm in diameter and were spherical, filamentous, or a combination of the two. The filamentous morphology correlated with the presence of a cylindrical matrix protein layer linked to the inner leaflet of the viral envelope and with local ordering of the glycoprotein spikes. Recombinant viruses with only the fusion protein in their envelope showed that these glycoproteins were predominantly in the postfusion conformation, but some were also in the prefusion form. The ribonucleocapsids were left-handed, randomly oriented, and curved inside the virions. In filamentous particles, they were often adjacent to an intermediate layer of protein assigned to M2-1 (an envelopeassociated protein known to mediate association of ribonucleocapsids with the matrix protein). Our results indicate important differences in structure between the Paramyxovirinae and Pneumovirinae subfamilies within the Paramyxoviridae, and provide fresh insights into host cell exit of a serious pathogen.cryo-ET | paramyxovirus | virus structure H uman respiratory syncytial virus (HRSV) causes severe disease, especially in children and the elderly (1, 2). In a study covering data from 2005, it was found that HRSV was responsible for 22% of acute lower respiratory infections worldwide and 66,000-199,000 deaths in children under the age of 5 y (2). Treatment is currently limited to purine analogs (Ribavirin) and passive immune-prophylaxis with humanized monoclonal antibodies (Palivizumab) (3).RSV belongs to the Paramyxoviridae, a large family of enveloped, negative-sense RNA viruses with many members from humans and animals. Together with human metapneumovirus, it constitutes the Pneumovirinae subfamily. Like other paramyxoviruses, it has a plasma-membrane-derived lipid envelope and a helical ribonucleocapsid (RNP) that contains a single viral RNA molecule associated with nucleoproteins (N), and an RNA-dependent RNA polymerase. The virion contains, in addition, a matrix protein (M) (4-7) and a transcription antiterminator, M2-1 (an envelopeassociated protein known to mediate association of RNPs with the M protein) (8-11). The lipid envelope contains two transmembrane glycoproteins: the fusion protein, F, and a glycoprotein, G, used for attachment to host cells. In addition, HRSV contains a small hydrophobic protein, SH, of unclear function and proteins of cellular origin, such as actin and chaperones (12). The RNP and F have been analyzed using 3D-EM and X-ray crystallography (13,14).HRSV particles are pleomorphic with both spherical and filamentous particles of different sizes (15). Early electron microscopy studies showed the presence of a helical p...

show abstract

mentioning

confidence: 99%

mentioning

confidence: 99%

Architecture of respiratory syncytial virus revealed by electron cryotomography

Liljeroos

Krzyzaniak²,

Helenius³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

177

173

View full text Add to dashboard Cite

show abstract

“… Partial amino acid sequences of the hMPV F (blue) and hRSV F (red) antigenic site II (Toiron et al , 1996; Lopez et al , 1998; McLellan et al , 2011b). Partial sequences of the F‐415 and F‐416 chimeras are color coded to indicate the origin of their antigenic site II residues.…”

Section: Resultsmentioning

confidence: 99%

“…pRB21/postfusion hRSV F (Post‐hRSV F): This plasmid encodes residues 1–524 of the hRSV F ectodomain (Long strain) with a deletion of the fusion peptide (residues 137–146) to avoid aggregation (McLellan et al , 2011b; Palomo et al , 2016). …”

Section: Methodsmentioning

confidence: 99%

“…The search for new hRSV vaccines has been reinvigorated by recent advances in structure‐based design of soluble hRSV F proteins folded in either the prefusion (McLellan et al , 2013a; Krarup et al , 2015) or postfusion conformations (McLellan et al , 2011b; Swanson et al , 2011). Stabilized soluble forms of prefusion hRSV F were shown to induce higher levels of neutralizing antibodies than soluble postfusion hRSV F in mice, cotton rats, and non‐human primates (McLellan et al , 2013a; Krarup et al , 2015; Palomo et al , 2016).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Chimeric Pneumoviridae fusion proteins as immunogens to induce cross‐neutralizing antibody responses

et al. 2017

View full text Add to dashboard Cite

Human respiratory syncytial virus (hRSV) and human metapneumovirus (hMPV), two members of the Pneumoviridae family, account for the majority of severe lower respiratory tract infections worldwide in very young children. They are also a frequent cause of morbidity and mortality in the elderly and immunocompromised adults. High levels of neutralizing antibodies, mostly directed against the viral fusion (F) glycoprotein, correlate with protection against either hRSV or hMPV. However, no cross‐neutralization is observed in polyclonal antibody responses raised after virus infection or immunization with purified F proteins. Based on crystal structures of hRSV F and hMPV F, we designed chimeric F proteins in which certain residues of well‐characterized antigenic sites were swapped between the two antigens. The antigenic changes were monitored by ELISA with virus‐specific monoclonal antibodies. Inoculation of mice with these chimeras induced polyclonal cross‐neutralizing antibody responses, and mice were protected against challenge with the virus used for grafting of the heterologous antigenic site. These results provide a proof of principle for chimeric fusion proteins as single immunogens that can induce cross‐neutralizing antibody and protective responses against more than one human pneumovirus.

show abstract

Characterization of neutralizing affinity‐matured human respiratory syncytial virus F binding antibodies in the sub‐picomolar affinity range

Canziani

Melero

Lacy

2012

J of Molecular Recognition

View full text Add to dashboard Cite

In the human adaptation and optimization of a mouse anti-human respiratory syncytial virus neutralizing antibody, affinity assessment was crucial to distinguish among potential candidates and to evaluate whether this correlated with function in vitro and in vivo. This affinity assessment was complicated by the trimeric nature of the antigen target, respiratory syncytial virus F (RSV-F) glycoprotein. In the initial affinity screen, surface plasmon resonance was used to determine the intrinsic binding affinities of anti-RSV-F Fab and immunoglobulin G (IgG) to the extracellular domain of RSV-F. This assessment required minimal biotinylation of the RSV-F protein and design of a capture strategy to minimize avidity effects. Approximately 30 Fabs were selected from three optimization phage display libraries on the basis of an initial ELISA screen. Surface plasmon resonance analysis demonstrated the success of optimization with some candidates from the screened libraries having low picomolar dissociation constants, more than 700-fold tighter than the parental monoclonal antibody (B21M). The affinities of these antibodies were further evaluated by a kinetic exclusion assay, a solution binding technology. One IgG (monoclonal antibody 029) displayed a low picomolar K(D) comparable with that of motavizumab, an RSV antibody in clinical study. Kinetic exclusion assay showed that two other of the matured IgGs (011 and 019) had sub-picomolar dissociation constants that could not be resolved further. We discuss the relevance of these interaction analysis results in the light of recently published data on the mechanism of F-driven viral fusion during paramyxoviral infection and 101F epitope conservation revealed from the recent crystal structure of RSV-F in the post-fusion state.

show abstract

Structure of Respiratory Syncytial Virus Fusion Glycoprotein in the Postfusion Conformation Reveals Preservation of Neutralizing Epitopes

Cited by 354 publications

References 46 publications

Architecture of respiratory syncytial virus revealed by electron cryotomography

Architecture of respiratory syncytial virus revealed by electron cryotomography

Chimeric Pneumoviridae fusion proteins as immunogens to induce cross‐neutralizing antibody responses

Characterization of neutralizing affinity‐matured human respiratory syncytial virus F binding antibodies in the sub‐picomolar affinity range

Contact Info

Product

Resources

About