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Query: UMLS:C1175175 (SARS)
 19,188 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The SARS-CoV spike protein, a glycoprotein essential for viral entry, is a primary target for vaccine and drug development. Two peptides denoted HR-N(SN50) and HR-C(SC40), corresponding to the Leu/Ile/Val-rich heptad-repeat regions from the N-terminal and C-terminal segments of the SARS-CoV spike S2 sequence, respectively, were synthesized and predicted to form trimeric assembly of hairpin-like structures. The polyclonal antibodies produced by recombinant S2 protein were tested for antigenicity of the two heptad repeats. We report here the first crystallographic study of the SARS spike HR-N/HR-C complex. The crystal belongs to the triclinic space group P1 and the data-set collected to 2.98 A resolution showed noncrystallographic pseudo-222 and 3-fold symmetries. Based on these data, comparative modeling of the SARS-CoV fusion core was performed. The immunological and structural information presented herein may provide a more detailed understanding of the viral fusion mechanism as well as the development of effective therapy against SARS-CoV infection.
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PMID:Immunological, structural, and preliminary X-ray diffraction characterizations of the fusion core of the SARS-coronavirus spike protein. 1547 92

The lectins DC-SIGN and DC-SIGNR can augment viral infection; however, the range of pathogens interacting with these attachment factors is incompletely defined. Here we show that DC-SIGN and DC-SIGNR enhance infection mediated by the glycoprotein (GP) of Marburg virus (MARV) and the S protein of severe acute respiratory syndrome coronavirus and might promote viral dissemination. SIGNR1, a murine DC-SIGN homologue, also enhanced infection driven by MARV and Ebola virus GP and could be targeted to assess the role of attachment factors in filovirus infection in vivo.
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PMID:DC-SIGN and DC-SIGNR interact with the glycoprotein of Marburg virus and the S protein of severe acute respiratory syndrome coronavirus. 1547 53

Angiotensin-converting enzyme 2 (ACE2) is a receptor for SARS-CoV, the novel coronavirus that causes severe acute respiratory syndrome [Li, W. Moore, M. J., Vasilieva, N., Sui, J., Wong, S. K., Berne, M. A., Somasundaran, M., Sullivan, J. L., Luzuriaga, K., Greenough, T. C., et al. (2003) Nature 426, 450-454]. We have identified a different human cellular glycoprotein that can serve as an alternative receptor for SARS-CoV. A human lung cDNA library in vesicular stomatitis virus G pseudotyped retrovirus was transduced into Chinese hamster ovary cells, and the cells were sorted for binding of soluble SARS-CoV spike (S) glycoproteins, S(590) and S(1180). Clones of transduced cells that bound SARS-CoV S glycoprotein were inoculated with SARS-CoV, and increases in subgenomic viral RNA from 1-16 h or more were detected by multiplex RT-PCR in four cloned cell lines. Sequencing of the human lung cDNA inserts showed that each of the cloned cell lines contained cDNA that encoded human CD209L, a C-type lectin (also called L-SIGN). When the cDNA encoding CD209L from clone 2.27 was cloned and transfected into Chinese hamster ovary cells, the cells expressed human CD209L glycoprotein and became susceptible to infection with SARS-CoV. Immunohistochemistry showed that CD209L is expressed in human lung in type II alveolar cells and endothelial cells, both potential targets for SARS-CoV. Several other enveloped viruses including Ebola and Sindbis also use CD209L as a portal of entry, and HIV and hepatitis C virus can bind to CD209L on cell membranes but do not use it to mediate virus entry. Our data suggest that the large S glycoprotein of SARS-CoV may use both ACE2 and CD209L in virus infection and pathogenesis.
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PMID:CD209L (L-SIGN) is a receptor for severe acute respiratory syndrome coronavirus. 1549 74

Two candidate small interfering RNAs (siRNAs) corresponding to severe acute respiratory syndrome-associated coronavirus (SARS-CoV) spike gene were designed and in vitro transcribed to explore the possibility of silencing SARS-CoV S gene. The plasmid pEGFP-optS, which contains the codon-optimized SARS-CoV S gene and expresses spike-EGFP fusion protein (S-EGFP) as silencing target and expressing reporter, was transfected with siRNAs into HEK 293T cells. At various time points of posttransfection, the levels of S-EGFP expression and amounts of spike mRNA transcript were detected by fluorescence microscopy, flow cytometry, Western blot, and real-time quantitative PCR, respectively. The results showed that the cells transfected with pEGFP-optS expressed S-EGFP fusion protein at a higher level compared with those transfected with pEGFP-S, which contains wildtype SARS-CoV spike gene sequence. The green fluorescence, mean fluorescence intensity, and SARS-CoV S RNA transcripts were found significantly reduced, and the expression of SARS-CoV S glycoprotein was strongly inhibited in those cells co-transfected with either EGFP- or S-specific siRNAs. Our findings demonstrated that the S-specific siRNAs used in this study were able to specifically and effectively inhibit SARS-CoV S glycoprotein expression in cultured cells through blocking the accumulation of S mRNA, which may provide an approach for studies on the functions of SARS-CoV S gene and development of novel prophylactic or therapeutic agents for SARS-CoV.
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PMID:Silencing of SARS-CoV spike gene by small interfering RNA in HEK 293T cells. 1550 39

The recently emerged severe acute respiratory syndrome coronavirus (SARS-CoV) contains four structural genes, two replicase-transcriptase open reading frames, and more than five potential genes of unknown function. Despite this relative simplicity, the molecular regulation of SARS-CoV replication and assembly is not understood. Here, we report that two viral genes, encoding the SARS-CoV membrane (M) and nucleocapsid (N) proteins, are necessary and sufficient for formation of virus-like particles. Expression vectors encoding these two proteins were synthesized by using preferred human codons. When M and N expression plasmids were cotransfected into human 293 renal epithelial cells, pseudoparticles formed readily. The addition of a third gene, encoding the spike (S) glycoprotein, facilitated budding of particles that contained a corona-like halo resembling SARS-CoV when examined by transmission electron microscopy, with a buoyant density characteristic of coronaviruses. Specific biochemical interactions of these proteins were also shown in vitro. The S, M, and N proteins of the SARS-CoV are, therefore, necessary and sufficient for pseudovirus assembly. These findings advance the understanding of the morphogenesis of SARS-CoV and enable the generation of safe, conformational mimetics of the SARS virus that may facilitate the development of vaccines and antiviral drugs.
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PMID:Generation of synthetic severe acute respiratory syndrome coronavirus pseudoparticles: implications for assembly and vaccine production. 1550 43

The severe acute respiratory syndrome-coronavirus (SARS-CoV) spike (S) glycoprotein alone can mediate the membrane fusion required for virus entry and cell fusion. It is also a major immunogen and a target for entry inhibitors. Recent rapid advances in our knowledge of the structure and function of this protein have lead to the development of a number of candidate vaccine immunogens and SARS-CoV entry inhibitors.
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PMID:The SARS-CoV S glycoprotein. 1552 50

Severe acute respiratory syndrome (SARS), a life-threatening disease, is caused by the newly identified virus SARS coronavirus (SARS-CoV). In order to study the spike (S) protein of this highly contagious virus, we established a clonal cell-line, CHO-SG, from the Chinese hamster ovary cells that stably expresses C-terminally EGFP-tagged SARS-CoV S protein (S-EGFP). The ectodomain of the S glycoprotein is localized on the surface of CHO-SG cells with N-acetyl-glucosamine-terminated carbohydrate structure. CHO-SG cells associated tightly with Vero E6 cells, a SARS-CoV receptor (ACE2) expressing cell-line, and the interaction remained stable under highly stringent condition (1M NaCl). This interaction could be blocked by either the serum from a SARS convalescent patient or a goat anti-ACE2 antibody, indicating that the interaction is specific. A binding epitope with lesser degree of glycosylation and native conformation was localized by using rabbit anti-sera raised against five denatured recombinant S protein fragments expressed in Escherichia coli. One of the sera obtained from the fragment encompassing amino acids 48-358 significantly blocked the interaction between CHO-SG and Vero E6 cells. The region is useful for studying neutralizing antibodies in future vaccine development. This paper describes an easy and safe cell-based assay suitable for studying the binding between SARS-CoV S protein and its receptor.
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PMID:A novel cell-based binding assay system reconstituting interaction between SARS-CoV S protein and its cellular receptor. 1558 97

Severe acute respiratory syndrome coronavirus (SARS-CoV) is the etiological agent of SARS. Analysis of SARS-CoV spike glycoprotein (S) using recombinant plasmid and virus infections demonstrated that the S-precursor (proS) exists as a approximately 190 kDa endoplasmic reticulum form and a approximately 210 kDa Golgi-modified form. ProS is subsequently processed into two C-terminal proteins of approximately 110 and approximately 80 kDa. The membrane-bound proprotein convertases (PCs) furin, PC7 or PC5B enhanced the production of the approximately 80 kDa protein. In agreement, proS processing, cytopathic effects, and viral titers were enhanced in recombinant Vero E6 cells overexpressing furin, PC7 or PC5B. The convertase inhibitor dec-RVKR-cmk significantly reduced proS cleavage and viral titers of SARS-CoV infected cells. In addition, inhibition of processing by dec-RVKR-cmk completely abrogated the virus-induced cellular cytopathicity. A fluorogenically quenched synthetic peptide encompassing Arg(761) of the spike glycoprotein was efficiently cleaved by furin and the cleavage was inhibited by EDTA and dec-RVKR-cmk. Taken together, our data indicate that furin or PC-mediated processing plays a critical role in SARS-CoV spread and cytopathicity, and inhibitors of the PCs represent potential therapeutic anti-SARS-CoV agents.
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PMID:Implication of proprotein convertases in the processing and spread of severe acute respiratory syndrome coronavirus. 1559 35

The spike (S) glycoprotein is one of the major structure proteins of SARS-associated coronavirus (CoV). Fragment 450-650 (S450-650) of the S protein contains receptor-binding domain and neutralizing epitopes. In this study, S450-650 was expressed with a histidine tag in Escherichia coli BL21. Bacterial inclusion bodies containing the recombinant S450-650 were solubilized with 8 M urea and then applied onto a Ni-nitrilotriacetic acid column. On-column refolding and purification was performed. Reduced glutathione and oxidized glutathione were included in the refolding buffer. In the wash and elution buffers, glycerol and glucose were necessary additives to prevent protein aggregation during purification. This refolding and purification procedure allowed production of S450-650 at up to 500 microg/ml in soluble form, which maintained appropriate antigenicity and immunogenicity. It was able to induce strong IgG responses in BALB/c mice. In Western blot assays, the recombinant S450-650 was recognized by monoclonal Ab against the His-tag and also sera from a convalescent SARS patient. S450-650-based ELISA system was able to detect anti-SARS-CoV IgG Abs in patient sera.
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PMID:Prokaryotic expression, refolding, and purification of fragment 450-650 of the spike protein of SARS-coronavirus. 1564 67

Molecular characterization of the severe acute respiratory syndrome coronavirus has revealed genetic diversity among isolates. The spike (S) glycoprotein, the major target for vaccine and immune therapy, shows up to 17 substitutions in its 1,255-aa sequence; however, the biologic significance of these changes is unknown. Here, the functional effects of S mutations have been determined by analyzing their affinity for a viral receptor, human angiotensin-converting enzyme 2 (hACE-2), and their sensitivity to Ab neutralization with viral pseudotypes. Although minor differences among eight strains transmitted during human outbreaks in early 2003 were found, substantial functional changes were detected in S derived from a case in late 2003 from Guangdong province [S(GD03T0013)] and from two palm civets, S(SZ3) and S(SZ16). S(GD03T0013) depended less on the hACE-2 receptor and was markedly resistant to Ab inhibition. Unexpectedly, Abs that neutralized most human S glycoproteins enhanced entry mediated by the civet virus S glycoproteins. The mechanism of enhancement involved the interaction of Abs with conformational epitopes in the hACE-2-binding domain. Finally, improved immunogens and mAbs that minimize this complication have been defined. These data show that the entry of severe acute respiratory syndrome coronaviruses can be enhanced by Abs, and they underscore the need to address the evolving diversity of this newly emerged virus for vaccines and immune therapies.
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PMID:Evasion of antibody neutralization in emerging severe acute respiratory syndrome coronaviruses. 1564 42


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