Five of Five VHHs Neutralizing Poliovirus Bind the Receptor-Binding Site

ABSTRACTNanobodies, or VHHs, that recognize poliovirus type 1 have previously been selected and characterized as candidates for antiviral agents or reagents for standardization of vaccine quality control. In this study, we present high-resolution cryo-electron microscopy reconstructions of poliovirus with five neutralizing VHHs. All VHHs bind the capsid in the canyon at sites that extensively overlap the poliovirus receptor-binding site. In contrast, the interaction involves a unique (and surprisingly extensive) surface for each of the five VHHs. Five regions of the capsid were found to participate in binding with all five VHHs. Four of these five regions are known to alter during the expansion of the capsid associated with viral entry. Interestingly, binding of one of the VHHs, PVSS21E, resulted in significant changes of the capsid structure and thus seems to trap the virus in an early stage of expansion.IMPORTANCEWe describe the cryo-electron microscopy structures of complexes of five neutralizing VHHs with the Mahoney strain of type 1 poliovirus at resolutions ranging from 3.8 to 6.3Å. All five VHHs bind deep in the virus canyon at similar sites that overlap extensively with the binding site for the receptor (CD155). The binding surfaces on the VHHs are surprisingly extensive, but despite the use of similar binding surfaces on the virus, the binding surface on the VHHs is unique for each VHH. In four of the five complexes, the virus remains essentially unchanged, but for the fifth there are significant changes reminiscent of but smaller in magnitude than the changes associated with cell entry, suggesting that this VHH traps the virus in a previously undescribed early intermediate state. The neutralizing mechanisms of the VHHs and their potential use as quality control agents for the end game of poliovirus eradication are discussed.

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N-Linked Glycan at Residue 523 of Human Parainfluenza Virus Type 3 Hemagglutinin-Neuraminidase Masks a Second Receptor-Binding Site

Journal of Virology ◽

10.1128/jvi.02331-09 ◽

2010 ◽

Vol 84 (6) ◽

pp. 3094-3100 ◽

Cited By ~ 16

Author(s):

Vasiliy P. Mishin ◽

Makiko Watanabe ◽

Garry Taylor ◽

John DeVincenzo ◽

Michael Bose ◽

...

Keyword(s):

Binding Site ◽

Virus Type ◽

Receptor Binding ◽

Critical Role ◽

Glycosylation Site ◽

Parainfluenza Virus ◽

Receptor Binding Site ◽

Human Parainfluenza Virus ◽

Type 3

ABSTRACT The hemagglutinin-neuraminidase (HN) glycoprotein plays a critical role in parainfluenza virus replication. We recently found that in addition to the catalytic binding site, HN of human parainfluenza virus type 1 (hPIV-1) may have a second receptor-binding site covered by an N-linked glycan at residue 173, which is near the region of the second receptor-binding site identified in Newcastle disease virus (NDV) HN (I. A. Alymova, G. Taylor, V. P. Mishin, M. Watanabe, K. G. Murti, K. Boyd, P. Chand, Y. S. Babu, and A. Portner, J. Virol. 82:8400-8410, 2008). Sequence analysis and superposition of the NDV and hPIV-3 HN dimer structures revealed that, similar to what was seen in hPIV-1, the N-linked glycan at residue 523 on hPIV-3 HN may cover a second receptor-binding site. Removal of this N-linked glycosylation site by an Asn-to-Asp substitution at residue 523 (N523D) changed the spectrum of the mutant virus's receptor specificity, delayed its elution from both turkey and chicken red blood cells, reduced mutant sensitivity (by about half) to the selective HN inhibitor BCX 2855 in hemagglutination inhibition tests, and slowed its growth in LLC-MK2 cells. The neuraminidase activity of the mutant and its sensitivity to BCX 2855 in neuraminidase inhibition assays did not change, indicating that the mutation did not affect the virus's catalytic-binding site and that all observed effects were caused by the exposure of the purported second receptor-binding site. Our data are consistent with the idea that, similar to the case for hPIV-1, the N-linked glycan shields a second receptor-binding site on hPIV-3 HN.

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Length polymorphism within the second variable region of the human immunodeficiency virus type 1 envelope glycoprotein affects accessibility of the receptor binding site.

Journal of Virology ◽

10.1128/jvi.71.1.759-765.1997 ◽

1997 ◽

Vol 71 (1) ◽

pp. 759-765 ◽

Cited By ~ 29

Author(s):

D G Fox ◽

P Balfe ◽

C P Palmer ◽

J C May ◽

C Arnold ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Binding Site ◽

Human Immunodeficiency Virus Type ◽

Receptor Binding ◽

Length Polymorphism ◽

Envelope Glycoprotein ◽

Variable Region ◽

Receptor Binding Site ◽

Immunodeficiency Virus

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Mutation at Residue 523 Creates a Second Receptor Binding Site on Human Parainfluenza Virus Type 1 Hemagglutinin-Neuraminidase Protein

Journal of Virology ◽

10.1128/jvi.00969-06 ◽

2006 ◽

Vol 80 (18) ◽

pp. 9009-9016 ◽

Cited By ~ 18

Author(s):

Tatiana Bousse ◽

Toru Takimoto

Keyword(s):

Amino Acid ◽

Binding Site ◽

Receptor Binding ◽

Reverse Genetics ◽

Sendai Virus ◽

Parainfluenza Virus ◽

Amino Acid Difference ◽

Receptor Binding Site ◽

Human Parainfluenza Virus

ABSTRACT The paramyxovirus hemagglutinin-neuraminidase (HN) is a multifunctional protein mediating hemagglutination (HA), neuraminidase (NA), and fusion promotion activities. It has been a matter of debate whether HN contains combined or separate sites for HA and NA activities. To clear the issue, we determined the presence of the second binding site on human parainfluenza virus (hPIV) type 1, 2, and 3 and Sendai virus (SeV) HN proteins. Results of virus elution from erythrocytes at an elevated temperature and HA inhibition by NA inhibitor BCX-2798 suggest that all hPIVs bind to the receptor only through the NA catalytic site, while SeV HN has an additional receptor binding site. Comparison of SeV and hPIV1 HN sequences revealed two amino acid differences at residues 521 and 523 in the region close to the second binding site identified in Newcastle disease virus HN. We mutated hPIV1 HN at position 523 from Asn to the residue of SeV HN, Asp, and rescued a recombinant SeV that carries the mutated hPIV1 HN by a reverse genetics system. The hPIV1 HN with Asp at position 523 hemagglutinated in the presence of BCX-2798, suggesting that the amino acid difference at position 523 is critical for the formation of a second binding site. Creation of the second binding site on hPIV1 HN, however, did not significantly affect the growth or fusion activity of the recombinant virus. Our study indicates that the presence and requirement of a second binding site vary among paramyxoviruses.

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Chromatin immunoprecipitation (ChIP) assays identify a functional retinoic acid receptor binding site and -549T>C SNP in the PTGDR-5’ region

10.26226/morressier.5acdc65fd462b8029238deb0 ◽

2018 ◽

Author(s):

Esther Maria Moreno

Keyword(s):

Retinoic Acid ◽

Binding Site ◽

Receptor Binding ◽

Chromatin Immunoprecipitation ◽

Retinoic Acid Receptor ◽

Receptor Binding Site ◽

Acid Receptor

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Analysis of the receptor-binding site of murine coronavirus spike protein.

Journal of Virology ◽

10.1128/jvi.70.4.2632-2636.1996 ◽

1996 ◽

Vol 70 (4) ◽

pp. 2632-2636 ◽

Cited By ~ 44

Author(s):

H Suzuki ◽

F Taguchi

Keyword(s):

Binding Site ◽

Receptor Binding ◽

Spike Protein ◽

Receptor Binding Site ◽

Murine Coronavirus

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Mechanism of SARS-CoV-2 polymerase stalling by remdesivir

Nature Communications ◽

10.1038/s41467-020-20542-0 ◽

2021 ◽

Vol 12 (1) ◽

Cited By ~ 1

Author(s):

Goran Kokic ◽

Hauke S. Hillen ◽

Dimitry Tegunov ◽

Christian Dienemann ◽

Florian Seitz ◽

...

Keyword(s):

Electron Microscopy ◽

Rna Polymerase ◽

Binding Site ◽

Molecular Mechanism ◽

Active Center ◽

Rna Synthesis ◽

Active Form ◽

Nucleoside Triphosphate ◽

Rna Dependent Rna Polymerase ◽

Cryo Electron Microscopy

AbstractRemdesivir is the only FDA-approved drug for the treatment of COVID-19 patients. The active form of remdesivir acts as a nucleoside analog and inhibits the RNA-dependent RNA polymerase (RdRp) of coronaviruses including SARS-CoV-2. Remdesivir is incorporated by the RdRp into the growing RNA product and allows for addition of three more nucleotides before RNA synthesis stalls. Here we use synthetic RNA chemistry, biochemistry and cryo-electron microscopy to establish the molecular mechanism of remdesivir-induced RdRp stalling. We show that addition of the fourth nucleotide following remdesivir incorporation into the RNA product is impaired by a barrier to further RNA translocation. This translocation barrier causes retention of the RNA 3ʹ-nucleotide in the substrate-binding site of the RdRp and interferes with entry of the next nucleoside triphosphate, thereby stalling RdRp. In the structure of the remdesivir-stalled state, the 3ʹ-nucleotide of the RNA product is matched and located with the template base in the active center, and this may impair proofreading by the viral 3ʹ-exonuclease. These mechanistic insights should facilitate the quest for improved antivirals that target coronavirus replication.

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Potent sialic acid inhibitors that target influenza A virus hemagglutinin

Scientific Reports ◽

10.1038/s41598-021-87845-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yu-Jen Chang ◽

Cheng-Yun Yeh ◽

Ju-Chien Cheng ◽

Yu-Qi Huang ◽

Kai-Cheng Hsu ◽

...

Keyword(s):

Sialic Acid ◽

Antiviral Activity ◽

Binding Site ◽

Influenza A Virus ◽

Receptor Binding ◽

Influenza A ◽

The United States ◽

Ligand Complex ◽

Receptor Binding Site ◽

Computational Strategy

AbstractEradicating influenza A virus (IAV) is difficult, due to its genetic drift and reassortment ability. As the infectious cycle is initiated by the influenza glycoprotein, hemagglutinin (HA), which mediates the binding of virions to terminal sialic acids moieties, HA is a tempting target of anti-influenza inhibitors. However, the complexity of the HA structure has prevented delineation of the structural characterization of the HA protein–ligand complex. Our computational strategy efficiently analyzed > 200,000 records of compounds held in the United States National Cancer Institute (NCI) database and identified potential HA inhibitors, by modeling the sialic acid (SA) receptor binding site (RBS) for the HA structure. Our modeling revealed that compound NSC85561 showed significant antiviral activity against the IAV H1N1 strain with EC50 values ranging from 2.31 to 2.53 µM and negligible cytotoxicity (CC50 > 700 µM). Using the NSC85561 compound as the template to generate 12 derivatives, robust bioassay results revealed the strongest antiviral efficacies with NSC47715 and NSC7223. Virtual screening clearly identified three SA receptor binding site inhibitors that were successfully validated in experimental data. Thus, our computational strategy has identified SA receptor binding site inhibitors against HA that show IAV-associated antiviral activity.

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