scholarly journals Supramolecular Mechanism of Viral Envelope Disruption by Molecular Tweezers

2020 ◽  
Vol 142 (40) ◽  
pp. 17024-17038 ◽  
Author(s):  
Tatjana Weil ◽  
Rüdiger Groß ◽  
Annika Röcker ◽  
Kenny Bravo-Rodriguez ◽  
Christian Heid ◽  
...  
Keyword(s):  
Viral Envelope ◽  
Molecular Tweezers

Biological and Surface Properties of C-Type Virus Particles Produced by Novikoff Ascites Hepatoma Cells

1977 ◽  
Vol 35 ◽  
pp. 388-389
Author(s):  
D.C. Hixson ◽  
J.C. Chan ◽  
J.M. Bowen ◽  
E.F. Walborg
Keyword(s):  
Surface Properties ◽  
Ricinus Communis ◽  
Rat Kidney ◽  
Leukemia Virus ◽  
Viral Envelope ◽  
Virus Particles ◽  
Chemically Induced ◽  
Sarcoma Virus ◽  
Hepatocarcinoma Cells ◽  
Type C

Several years ago Karasaki (1) reported the production of type C virus particles by Novikoff ascites hepatocarcinoma cells. More recently, Weinstein (2) has reported the presence of type C virus particles in cell cultures derived from transplantable and primary hepatocellular carcinomas. To date, the biological function of these virus and their significance in chemically induced hepatocarcinogenesis are unknown. The present studies were initiated to determine a possible role for type C virus particles in chemically induced hepatocarcinogenesis. This communication describes results of studies on the biological and surface properties of type C virus associated with Novikoff hepatocarcinoma cells.Ecotropic and xenotropic murine leukemia virus (MuLV) activity in ascitic fluid of Novikoff tumor-bearing rats was assayed in murine sarcoma virus transformed S+L- mouse cells and S+L- mink cells, respectively. The presence of sarcoma virus activity was assayed in non-virus-producing normal rat kidney (NRK) cells. Ferritin conjugates of concanavalin A (Fer-Con wheat germ agglutinin (Fer-WGA), and Ricinus communis agglutinins I and II (Fer-RCAI and Fer-RCAII) were used to probe the structure and topography of saccharide determinants present on the viral envelope.

scholarly journals Digital Droplet PCR for SARS-CoV-2 Resolves Borderline Cases

2021 ◽  
Author(s):  
Jing Xu ◽  
Timothy Kirtek ◽  
Yan Xu ◽  
Hui Zheng ◽  
Huiyu Yao ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Limit Of Detection ◽  
False Negative ◽  
Accurate Method ◽  
Viral Envelope ◽  
Chain Reaction ◽  
Borderline Cases ◽  
Droplet Pcr ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Abstract Objectives The Bio-Rad SARS-CoV-2 ddPCR Kit (Bio-Rad Laboratories) was the first droplet digital polymerase chain reaction (ddPCR) assay to receive Food and Drug Administration (FDA) Emergency Use Authorization approval, but it has not been evaluated clinically. We describe the performance of ddPCR—in particular, its ability to confirm weak-positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) results. Methods We clinically validated the Bio-Rad Triplex Probe ddPCR Assay. The limit of detection was determined by using serial dilutions of SARS-CoV-2 RNA in an artificial viral envelope. The ddPCR assay was performed according to the manufacturer’s specifications on specimens confirmed to be positive (n = 48) or negative (n = 30) by an FDA-validated reverse transcription–polymerase chain reaction assay on the m2000 RealTime system (Abbott). Ten borderline positive cases were also evaluated. Results The limit of detection was 50 copies/mL (19 of 20 positive). Forty-seven specimens spanning a range of quantification cycles (2.9-25.9 cycle numbers) were positive by this assay (47 of 48; 97.9% positive precent agreement), and 30 negative samples were confirmed as negative (30 of 30; 100% negative percent agreement). Nine of 10 borderline cases were positive when tested in triplicate. Conclusions The ddPCR of SARS-CoV-2 is an accurate method, with superior sensitivity for viral RNA detection. It could provide definitive evaluation of borderline positive cases or suspected false-negative cases.

scholarly journals Prefusion structure of human cytomegalovirus glycoprotein B and structural basis for membrane fusion

2021 ◽  
Vol 7 (10) ◽  
pp. eabf3178
Author(s):  
Yuhang Liu ◽  
Kyle P. Heim ◽  
Ye Che ◽  
Xiaoyuan Chi ◽  
Xiayang Qiu ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Human Cytomegalovirus ◽  
Transmembrane Domain ◽  
Proximal Region ◽  
Viral Envelope ◽  
Vaccine Antigen ◽  
Glycoprotein B ◽  
Structural Basis ◽  
Fusion Inhibitor ◽  
Host Cell Membrane

Human cytomegalovirus (HCMV) causes congenital disease with long-term morbidity. HCMV glycoprotein B (gB) transitions irreversibly from a metastable prefusion to a stable postfusion conformation to fuse the viral envelope with a host cell membrane during entry. We stabilized prefusion gB on the virion with a fusion inhibitor and a chemical cross-linker, extracted and purified it, and then determined its structure to 3.6-Å resolution by electron cryomicroscopy. Our results revealed the structural rearrangements that mediate membrane fusion and details of the interactions among the fusion loops, the membrane-proximal region, transmembrane domain, and bound fusion inhibitor that stabilized gB in the prefusion state. The structure rationalizes known gB antigenic sites. By analogy to successful vaccine antigen engineering approaches for other viral pathogens, the high-resolution prefusion gB structure provides a basis to develop stabilized prefusion gB HCMV vaccine antigens.

scholarly journals Viral envelope fails to deliver?

Nature ◽  
10.1038/35504 ◽  
1998 ◽  
Vol 391 (6668) ◽  
pp. 638-639 ◽  
Author(s):  
Dani P. Bolognesi ◽  
Thomas J. Matthews
Keyword(s):  
Viral Envelope

scholarly journals Site-specific N-glycosylation analysis of animal cell culture-derived Zika virus proteins

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alexander Pralow ◽  
Alexander Nikolay ◽  
Arnaud Leon ◽  
Yvonne Genzel ◽  
Erdmann Rapp ◽  
...  
Keyword(s):  
Zika Virus ◽  
Animal Cell ◽  
Gradient Centrifugation ◽  
Viral Envelope ◽  
Protein Glycosylation ◽  
High Yield ◽  
Structural Protein ◽  
Site Specific ◽  
E Protein ◽  
The Impact

AbstractHere, we present for the first time, a site-specific N-glycosylation analysis of proteins from a Brazilian Zika virus (ZIKV) strain. The virus was propagated with high yield in an embryo-derived stem cell line (EB66, Valneva SE), and concentrated by g-force step-gradient centrifugation. Subsequently, the sample was proteolytically digested with different enzymes, measured via a LC–MS/MS-based workflow, and analyzed in a semi-automated way using the in-house developed glyXtoolMS software. The viral non-structural protein 1 (NS1) was glycosylated exclusively with high-mannose structures on both potential N-glycosylation sites. In case of the viral envelope (E) protein, no specific N-glycans could be identified with this method. Nevertheless, N-glycosylation could be proved by enzymatic de-N-glycosylation with PNGase F, resulting in a strong MS-signal of the former glycopeptide with deamidated asparagine at the potential N-glycosylation site N444. This confirmed that this site of the ZIKV E protein is highly N-glycosylated but with very high micro-heterogeneity. Our study clearly demonstrates the progress made towards site-specific N-glycosylation analysis of viral proteins, i.e. for Brazilian ZIKV. It allows to better characterize viral isolates, and to monitor glycosylation of major antigens. The method established can be applied for detailed studies regarding the impact of protein glycosylation on antigenicity and human pathogenicity of many viruses including influenza virus, HIV and corona virus.

scholarly journals Specific inhibition of the Survivin–CRM1 interaction by peptide-modified molecular tweezers

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Annika Meiners ◽  
Sandra Bäcker ◽  
Inesa Hadrović ◽  
Christian Heid ◽  
Christine Beuck ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Rational Design ◽  
Nuclear Export Signal ◽  
Signal Interference ◽  
Receptor Interaction ◽  
Dual Function ◽  
Experimental Proof ◽  
Molecular Tweezers ◽  
Recognition Element ◽  
Dynamics Simulations

AbstractSurvivin’s dual function as apoptosis inhibitor and regulator of cell proliferation is mediated via its interaction with the export receptor CRM1. This protein–protein interaction represents an attractive target in cancer research and therapy. Here, we report a sophisticated strategy addressing Survivin’s nuclear export signal (NES), the binding site of CRM1, with advanced supramolecular tweezers for lysine and arginine. These were covalently connected to small peptides resembling the natural, self-complementary dimer interface which largely overlaps with the NES. Several biochemical methods demonstrated sequence-selective NES recognition and interference with the critical receptor interaction. These data were strongly supported by molecular dynamics simulations and multiscale computational studies. Rational design of lysine tweezers equipped with a peptidic recognition element thus allowed to address a previously unapproachable protein surface area. As an experimental proof-of-principle for specific transport signal interference, this concept should be transferable to any protein epitope with a flanking well-accessible lysine.

scholarly journals Role of Envelope Glycoprotein Complexes in Cell-Associated Spread of Human Cytomegalovirus

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 614
Author(s):  
Nina Weiler ◽  
Caroline Paal ◽  
Kerstin Adams ◽  
Christopher Calcaterra ◽  
Dina Fischer ◽  
...  
Keyword(s):  
Human Cytomegalovirus ◽  
Envelope Glycoprotein ◽  
Infectious Virus ◽  
Viral Envelope ◽  
Accessory Proteins ◽  
Viral Growth ◽  
Cell Association ◽  
Quantitative Immunoblotting ◽  
Model Virus

The role of viral envelope glycoproteins, particularly the accessory proteins of trimeric and pentameric gH/gL-complexes, in cell-associated spread of human cytomegalovirus (HCMV) is unclear. We aimed to investigate their contribution in the context of HCMV variants that grow in a strictly cell-associated manner. In the genome of Merlin pAL1502, the glycoproteins gB, gH, gL, gM, and gN were deleted by introducing stop codons, and the mutants were analyzed for viral growth. Merlin and recent HCMV isolates were compared by quantitative immunoblotting for expression of accessory proteins of the trimeric and pentameric gH/gL-complexes, gO and pUL128. Isolates were treated with siRNAs against gO and pUL128 and analyzed regarding focal growth and release of infectious virus. All five tested glycoproteins were essential for growth of Merlin pAL1502. Compared with this model virus, higher gO levels were measured in recent isolates of HCMV, and its knockdown decreased viral growth. Knockdown of pUL128 abrogated the strict cell-association and led to release of infectivity, which allowed cell-free transfer to epithelial cells where the virus grew again strictly cell-associated. We conclude that both trimer and pentamer contribute to cell-associated spread of recent clinical HCMV isolates and downregulation of pentamer can release infectious virus into the supernatant.

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