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1.
Biomaterials ; 286: 121585, 2022 Jul.
Article in English | MEDLINE | ID: covidwho-1881707

ABSTRACT

Among all the biological entities involved in the immune response, galectins, a family of glycan-binding proteins, have been described as key in immune cell homeostasis and modulation. More importantly, only some galectin family members are crucial in the resolution of inflammation, while others perpetuate the immune response in a pathological context. As they are expressed in most major diseases, their potential as targets for new therapies seems promising. Most of the galectin family members' ubiquitous expression points to the need for targeted treatments to ensure effectiveness. Engineered biomaterials are emerging as a promising method to improve galectin-targeted strategies' therapeutic performance. In this review, we provide an overview of the role of galectins in health and disease and their potential as therapeutic targets, as well as the state-of-the-art and future directions of galectin-targeted biomaterials.


Subject(s)
Biocompatible Materials , Galectins , Galectins/metabolism , Galectins/therapeutic use , Humans , Inflammation , Polysaccharides/metabolism
2.
Nutrients ; 14(11)2022 May 27.
Article in English | MEDLINE | ID: covidwho-1869720

ABSTRACT

Fucoidan, a sulfated polysaccharide extracted from brown seaweed, has been proposed to effectively treat and prevent various viral infections. However, the mechanisms behind its antiviral activity are not completely understood. We investigate here the global transcriptional changes in bone marrow-derived dendritic cells (BMDCs) using RNA-Seq technology. Through both analysis of differentially expressed genes (DEG) and gene set enrichment analysis (GSEA), we found that fucoidan-treated BMDCs were enriched in virus-specific response pathways, including that of SARS-CoV-2, as well as pathways associated with nucleic acid-sensing receptors (RLR, TLR, NLR, STING), and type I interferon (IFN) production. We show that these transcriptome changes are driven by well-known regulators of the inflammatory response against viruses, including IRF, NF-κB, and STAT family transcription factors. Furthermore, 435 of the 950 upregulated DEGs are classified as type I IFN-stimulated genes (ISGs). Flow cytometric analysis additionally showed that fucoidan increased MHCII, CD80, and CD40 surface markers in BMDCs, indicative of greater antigen presentation and co-stimulation functionality. Our current study suggests that fucoidan transcriptionally activates PRR signaling, type I IFN production and signaling, ISGs production, and DC maturation, highlighting a potential mechanism of fucoidan-induced antiviral activity.


Subject(s)
COVID-19 , Dendritic Cells , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Humans , Polysaccharides/metabolism , Polysaccharides/pharmacology , SARS-CoV-2
3.
Molecules ; 27(9)2022 Apr 26.
Article in English | MEDLINE | ID: covidwho-1810050

ABSTRACT

Plant polysaccharides can increase the number and variety of beneficial bacteria in the gut and produce a variety of active substances, including short-chain fatty acids (SCFAs). Gut microbes and their specific metabolites have the effects of promoting anti-inflammatory activity, enhancing the intestinal barrier, and activating and regulating immune cells, which are beneficial for improving immunity. A strong immune system reduces inflammation caused by external viruses and other pathogens. Coronavirus disease 2019 (COVID-19) is still spreading globally, and patients with COVID-19 often have intestinal disease and weakened immune systems. This article mainly evaluates how polysaccharides in plants can improve the immune system barrier by improving the intestinal microecological balance, which may have potential in the prevention and treatment of COVID-19.


Subject(s)
COVID-19 , Gastrointestinal Microbiome , COVID-19/drug therapy , Fatty Acids, Volatile/metabolism , Humans , Immunity , Polysaccharides/metabolism , Polysaccharides/pharmacology , Polysaccharides/therapeutic use
4.
Int J Mol Sci ; 23(5)2022 Feb 25.
Article in English | MEDLINE | ID: covidwho-1736943

ABSTRACT

Rouleaux (stacked clumps) of red blood cells (RBCs) observed in the blood of COVID-19 patients in three studies call attention to the properties of several enveloped virus strains dating back to seminal findings of the 1940s. For COVID-19, key such properties are: (1) SARS-CoV-2 binds to RBCs in vitro and also in the blood of COVID-19 patients; (2) although ACE2 is its target for viral fusion and replication, SARS-CoV-2 initially attaches to sialic acid (SA) terminal moieties on host cell membranes via glycans on its spike protein; (3) certain enveloped viruses express hemagglutinin esterase (HE), an enzyme that releases these glycan-mediated bindings to host cells, which is expressed among betacoronaviruses in the common cold strains but not the virulent strains, SARS-CoV, SARS-CoV-2 and MERS. The arrangement and chemical composition of the glycans at the 22 N-glycosylation sites of SARS-CoV-2 spike protein and those at the sialoglycoprotein coating of RBCs allow exploration of specifics as to how virally induced RBC clumping may form. The in vitro and clinical testing of these possibilities can be sharpened by the incorporation of an existing anti-COVID-19 therapeutic that has been found in silico to competitively bind to multiple glycans on SARS-CoV-2 spike protein.


Subject(s)
COVID-19/metabolism , Erythrocytes/metabolism , SARS-CoV-2/metabolism , Sialoglycoproteins/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Basigin/metabolism , Binding Sites , COVID-19/virology , Glycosylation , Hemagglutination , Hemagglutinins, Viral/metabolism , Humans , N-Acetylneuraminic Acid/metabolism , Polysaccharides/metabolism , Protein Binding , SARS-CoV-2/physiology , Viral Fusion Proteins/metabolism , Virus Internalization
6.
Sci Rep ; 12(1): 2594, 2022 02 16.
Article in English | MEDLINE | ID: covidwho-1692553

ABSTRACT

Complex glycans decorate viral surface proteins and play a critical role in virus-host interactions. Viral surface glycans shield vulnerable protein epitopes from host immunity yet can also present distinct "glycoepitopes" that can be targeted by host antibodies such as the potent anti-HIV antibody 2G12 that binds high-mannose glycans on gp120. Two recent publications demonstrate 2G12 binding to high mannose glycans on SARS-CoV-2 and select Influenza A (Flu) H3N2 viruses. Previously, our lab observed 2G12 binding and functional inhibition of a range of Flu viruses that include H3N2 and H1N1 lineages. In this manuscript, we present these data alongside structural analyses to offer an expanded picture of 2G12-Flu interactions. Further, based on the remarkable breadth of 2G12 N-glycan recognition and the structural factors promoting glycoprotein oligomannosylation, we hypothesize that 2G12 glycoepitopes can be defined from protein structure alone according to N-glycan site topology. We develop a model describing 2G12 glycoepitopes based on N-glycan site topology, and apply the model to identify viruses within the Protein Data Bank presenting putative 2G12 glycoepitopes for 2G12 repurposing toward analytical, diagnostic, and therapeutic applications.


Subject(s)
Antibodies, Monoclonal/metabolism , Broadly Neutralizing Antibodies/metabolism , HIV Antibodies/metabolism , Influenza A Virus, H1N1 Subtype/immunology , Influenza A Virus, H3N2 Subtype/immunology , Models, Immunological , SARS-CoV-2/immunology , Animals , Dogs , Drug Repositioning , Epitopes , Hemagglutinin Glycoproteins, Influenza Virus/metabolism , Humans , Influenza A Virus, H1N1 Subtype/metabolism , Influenza A Virus, H3N2 Subtype/metabolism , Madin Darby Canine Kidney Cells , Molecular Targeted Therapy , Neutralization Tests , Polysaccharides/metabolism
7.
Viruses ; 14(2)2022 02 08.
Article in English | MEDLINE | ID: covidwho-1674831

ABSTRACT

This article aims to review all currently known interactions between animal and human coronaviruses and their cellular receptors. Over the past 20 years, three novel coronaviruses have emerged that have caused severe disease in humans, including SARS-CoV-2 (severe acute respiratory syndrome virus 2); therefore, a deeper understanding of coronavirus host-cell interactions is essential. Receptor-binding is the first stage in coronavirus entry prior to replication and can be altered by minor changes within the spike protein-the coronavirus surface glycoprotein responsible for the recognition of cell-surface receptors. The recognition of receptors by coronaviruses is also a major determinant in infection, tropism, and pathogenesis and acts as a key target for host-immune surveillance and other potential intervention strategies. We aim to highlight the need for a continued in-depth understanding of this subject area following on from the SARS-CoV-2 pandemic, with the possibility for more zoonotic transmission events. We also acknowledge the need for more targeted research towards glycan-coronavirus interactions as zoonotic spillover events from animals to humans, following an alteration in glycan-binding capability, have been well-documented for other viruses such as Influenza A.


Subject(s)
Host Microbial Interactions , Polysaccharides/metabolism , Receptors, Virus/metabolism , SARS-CoV-2/metabolism , Viral Tropism , Animals , COVID-19/transmission , COVID-19/virology , Humans , Protein Binding , SARS-CoV-2/immunology , Virus Internalization
8.
Int J Mol Sci ; 22(16)2021 Aug 05.
Article in English | MEDLINE | ID: covidwho-1662663

ABSTRACT

Coxsackievirus A24 variant (CVA24v) is the primary causative agent of the highly contagious eye infection designated acute hemorrhagic conjunctivitis (AHC). It is solely responsible for two pandemics and several recurring outbreaks of the disease over the last decades, thus affecting millions of individuals throughout the world. To date, no antiviral agents or vaccines are available for combating this disease, and treatment is mainly supportive. CVA24v utilizes Neu5Ac-containing glycans as attachment receptors facilitating entry into host cells. We have previously reported that pentavalent Neu5Ac conjugates based on a glucose-scaffold inhibit CVA24v infection of human corneal epithelial cells. In this study, we report on the design and synthesis of scaffold-replaced pentavalent Neu5Ac conjugates and their effect on CVA24v cell transduction and the use of cryogenic electron microscopy (cryo-EM) to study the binding of these multivalent conjugates to CVA24v. The results presented here provide insights into the development of Neu5Ac-based inhibitors of CVA24v and, most significantly, the first application of cryo-EM to study the binding of a multivalent ligand to a lectin.


Subject(s)
Antiviral Agents/pharmacology , Coxsackievirus Infections/diet therapy , Enterovirus C, Human/drug effects , N-Acetylneuraminic Acid/pharmacology , Conjunctivitis, Acute Hemorrhagic/drug therapy , Conjunctivitis, Acute Hemorrhagic/metabolism , Conjunctivitis, Acute Hemorrhagic/virology , Coxsackievirus Infections/metabolism , Coxsackievirus Infections/virology , Glucose/metabolism , Humans , Lectins/metabolism , Ligands , Polysaccharides/metabolism , Receptors, Virus/metabolism
9.
Cells ; 11(3)2022 01 20.
Article in English | MEDLINE | ID: covidwho-1643582

ABSTRACT

Pathogenic enveloped viruses are covered with a glycan shield that provides a dual function: the glycan structures contribute to virus protection as well as host cell recognition. The three classical types of N-glycans, in particular complex glycans, high-mannose glycans, and hybrid glycans, together with some O-glycans, participate in the glycan shield of the Ebola virus, influenza virus, human cytomegalovirus, herpes virus, human immunodeficiency virus, Lassa virus, and MERS-CoV, SARS-CoV, and SARS-CoV-2, which are responsible for respiratory syndromes. The glycans are linked to glycoproteins that occur as metastable prefusion glycoproteins on the surface of infectious virions such as gp120 of HIV, hemagglutinin of influenza, or spike proteins of beta-coronaviruses. Plant lectins with different carbohydrate-binding specificities and, especially, mannose-specific lectins from the Vicieae tribe, such as pea lectin and lentil lectin, can be used as glycan probes for targeting the glycan shield because of their specific interaction with the α1,6-fucosylated core Man3GlcNAc2, which predominantly occurs in complex and hybrid glycans. Other plant lectins with Neu5Ac specificity or GalNAc/T/Tn specificity can also serve as potential glycan probes for the often sialylated complex glycans and truncated O-glycans, respectively, which are abundantly distributed in the glycan shield of enveloped viruses. The biomedical and therapeutical potential of plant lectins as antiviral drugs is discussed.


Subject(s)
COVID-19/metabolism , Fabaceae/metabolism , Plant Lectins/metabolism , Polysaccharides/metabolism , SARS-CoV-2/metabolism , Viral Envelope/metabolism , COVID-19/epidemiology , COVID-19/virology , Humans , Mannose/metabolism , Protein Binding , SARS-CoV-2/physiology , Virion/metabolism , Virus Internalization
10.
Elife ; 102021 12 20.
Article in English | MEDLINE | ID: covidwho-1592091

ABSTRACT

Infection and viral entry of SARS-CoV-2 crucially depends on the binding of its Spike protein to angiotensin converting enzyme 2 (ACE2) presented on host cells. Glycosylation of both proteins is critical for this interaction. Recombinant soluble human ACE2 can neutralize SARS-CoV-2 and is currently undergoing clinical tests for the treatment of COVID-19. We used 3D structural models and molecular dynamics simulations to define the ACE2 N-glycans that critically influence Spike-ACE2 complex formation. Engineering of ACE2 N-glycosylation by site-directed mutagenesis or glycosidase treatment resulted in enhanced binding affinities and improved virus neutralization without notable deleterious effects on the structural stability and catalytic activity of the protein. Importantly, simultaneous removal of all accessible N-glycans from recombinant soluble human ACE2 yields a superior SARS-CoV-2 decoy receptor with promise as effective treatment for COVID-19 patients.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Molecular Dynamics Simulation , Polysaccharides/metabolism , Receptors, Virus/metabolism , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/genetics , COVID-19/prevention & control , COVID-19/virology , Glycosylation , Humans , Polysaccharides/chemistry , Protein Binding , Protein Engineering , Receptors, Virus/chemistry , Receptors, Virus/genetics , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Virus Internalization
11.
Glycobiology ; 32(1): 60-72, 2022 02 26.
Article in English | MEDLINE | ID: covidwho-1501077

ABSTRACT

Extensive glycosylation of the spike protein of severe acute respiratory syndrome coronavirus 2 virus not only shields the major part of it from host immune responses, but glycans at specific sites also act on its conformation dynamics and contribute to efficient host receptor binding, and hence infectivity. As variants of concern arise during the course of the coronavirus disease of 2019 pandemic, it is unclear if mutations accumulated within the spike protein would affect its site-specific glycosylation pattern. The Alpha variant derived from the D614G lineage is distinguished from others by having deletion mutations located right within an immunogenic supersite of the spike N-terminal domain (NTD) that make it refractory to most neutralizing antibodies directed against this domain. Despite maintaining an overall similar structural conformation, our mass spectrometry-based site-specific glycosylation analyses of similarly produced spike proteins with and without the D614G and Alpha variant mutations reveal a significant shift in the processing state of N-glycans on one specific NTD site. Its conversion to a higher proportion of complex type structures is indicative of altered spatial accessibility attributable to mutations specific to the Alpha variant that may impact its transmissibility. This and other more subtle changes in glycosylation features detected at other sites provide crucial missing information otherwise not apparent in the available cryogenic electron microscopy-derived structures of the spike protein variants.


Subject(s)
COVID-19/epidemiology , Glycopeptides/chemistry , Mutation , Polysaccharides/chemistry , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/transmission , COVID-19/virology , Carbohydrate Sequence , Datasets as Topic , Glycopeptides/genetics , Glycopeptides/metabolism , Glycosylation , HEK293 Cells , Humans , Mass Spectrometry , Peptide Mapping , Polysaccharides/metabolism , Protein Binding , Receptors, Virus/genetics , Receptors, Virus/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
12.
J Mol Biol ; 434(2): 167332, 2022 01 30.
Article in English | MEDLINE | ID: covidwho-1492301

ABSTRACT

Extensive glycosylation of viral glycoproteins is a key feature of the antigenic surface of viruses and yet glycan processing can also be influenced by the manner of their recombinant production. The low yields of the soluble form of the trimeric spike (S) glycoprotein from SARS-CoV-2 has prompted advances in protein engineering that have greatly enhanced the stability and yields of the glycoprotein. The latest expression-enhanced version of the spike incorporates six proline substitutions to stabilize the prefusion conformation (termed SARS-CoV-2 S HexaPro). Although the substitutions greatly enhanced expression whilst not compromising protein structure, the influence of these substitutions on glycan processing has not been explored. Here, we show that the site-specific N-linked glycosylation of the expression-enhanced HexaPro resembles that of an earlier version containing two proline substitutions (2P), and that both capture features of native viral glycosylation. However, there are site-specific differences in glycosylation of HexaPro when compared to 2P. Despite these discrepancies, analysis of the serological reactivity of clinical samples from infected individuals confirmed that both HexaPro and 2P protein are equally able to detect IgG, IgA, and IgM responses in all sera analysed. Moreover, we extend this observation to include an analysis of glycan engineered S protein, whereby all N-linked glycans were converted to oligomannose-type and conclude that serological activity is not impacted by large scale changes in glycosylation. These observations suggest that variations in glycan processing will not impact the serological assessments currently being performed across the globe.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , Mutation, Missense/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Antibodies, Viral/blood , Binding Sites/genetics , COVID-19/virology , Glycosylation , Humans , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , Mannose/metabolism , Mutation, Missense/genetics , Oligosaccharides/metabolism , Polysaccharides/metabolism , Proline/genetics , Proline/immunology , Proline/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/immunology , Recombinant Proteins/metabolism , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
13.
Int J Mol Sci ; 22(21)2021 Oct 26.
Article in English | MEDLINE | ID: covidwho-1488607

ABSTRACT

Emerging evidence suggests that males are more susceptible to severe infection by the SARS-CoV-2 virus than females. A variety of mechanisms may underlie the observed gender-related disparities including differences in sex hormones. However, the precise mechanisms by which female sex hormones may provide protection against SARS-CoV-2 infectivity remains unknown. Here we report new insights into the molecular basis of the interactions between the SARS-CoV-2 spike (S) protein and the human ACE2 receptor. We further report that glycosylation of the ACE2 receptor enhances SARS-CoV-2 infectivity. Importantly, estrogens can disrupt glycan-glycan interactions and glycan-protein interactions between the human ACE2 and the SARS-CoV-2 thereby blocking its entry into cells. In a mouse model of COVID-19, estrogens reduced ACE2 glycosylation and thereby alveolar uptake of the SARS-CoV-2 spike protein. These results shed light on a putative mechanism whereby female sex hormones may provide protection from developing severe infection and could inform the development of future therapies against COVID-19.


Subject(s)
Estrogens/chemistry , Estrogens/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/physiology , Virus Internalization/drug effects , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Animals , Biological Transport , COVID-19/drug therapy , COVID-19/metabolism , Disease Models, Animal , Estrogens/pharmacology , Glycosylation/drug effects , Human Umbilical Vein Endothelial Cells , Humans , Male , Mice, Inbred C57BL , Models, Molecular , Molecular Docking Simulation , Molecular Dynamics Simulation , Polysaccharides/chemistry , Polysaccharides/metabolism , SARS-CoV-2/drug effects , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Tunicamycin/pharmacology
14.
J Am Chem Soc ; 143(42): 17615-17621, 2021 10 27.
Article in English | MEDLINE | ID: covidwho-1467046

ABSTRACT

Cellular binding and entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are mediated by its spike glycoprotein (S protein), which binds with not only the human angiotensin-converting enzyme 2 (ACE2) receptor but also glycosaminoglycans such as heparin. Cell membrane-coated nanoparticles ("cellular nanosponges") mimic the host cells to attract and neutralize SARS-CoV-2 through natural cellular receptors, leading to a broad-spectrum antiviral strategy. Herein, we show that increasing surface heparin density on the cellular nanosponges can promote their inhibition against SARS-CoV-2. Specifically, cellular nanosponges are made with azido-expressing host cell membranes followed by conjugating heparin to the nanosponge surfaces. Cellular nanosponges with a higher heparin density have a larger binding capacity with viral S proteins and a significantly higher inhibition efficacy against SARS-CoV-2 infectivity. Overall, surface glycan engineering of host-mimicking cellular nanosponges is a facile method to enhance SARS-CoV-2 inhibition. This approach can be readily generalized to promote the inhibition of other glycan-dependent viruses.


Subject(s)
COVID-19/drug therapy , Heparin/administration & dosage , Nanostructures/therapeutic use , Polysaccharides/administration & dosage , SARS-CoV-2/metabolism , COVID-19/virology , Heparin/metabolism , Humans , Polysaccharides/metabolism
15.
Biochem Biophys Res Commun ; 579: 69-75, 2021 11 19.
Article in English | MEDLINE | ID: covidwho-1432975

ABSTRACT

N-glycosylation plays an important role in the pathogenesis of viral infections. However, the role of SARS-CoV-2 RBD N-glycosylation in viral entry remains elusive. In this study, we expressed and purified N331 and N343 N-glycosite mutants of SARS-CoV-2 RBD. We found that de-glycosylation at N331 and N343 drastically reduces the RBD binding to ACE2. More importantly, based on qualitative and quantitative virology research methods, we show that the mutation of RBD N-glycosites interfered with SARS-CoV-2 internalization rather than attachment potentially by decreasing RBD binding to the receptors. Also, the double N-glycosites mutant (N331 + N343) showed significantly increased sensitivity against the designated RBD neutralizing antibodies. Taken together, these results suggest that N-glycosylation of SARS-CoV-2 RBD is not only critical for viral internalization into respiratory epithelial cells but also shields the virus from neutralization. It may provide new insights into the biological process of early-stage SARS-CoV-2 infection with potential therapeutic implications.


Subject(s)
Polysaccharides/metabolism , Pulmonary Alveoli/cytology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Neutralizing , Binding Sites , COVID-19/metabolism , COVID-19/virology , Cell Line , Epithelial Cells , Glycosylation , Host-Pathogen Interactions/physiology , Humans , Mutation , Polysaccharides/chemistry , Pulmonary Alveoli/virology , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Virus Attachment
17.
J Biol Chem ; 297(4): 101207, 2021 10.
Article in English | MEDLINE | ID: covidwho-1415531

ABSTRACT

Certain sulfated glycans, including those from marine sources, can show potential effects against SARS-CoV-2. Here, a new fucosylated chondroitin sulfate (FucCS) from the sea cucumber Pentacta pygmaea (PpFucCS) (MW ∼10-60 kDa) was isolated and structurally characterized by NMR. PpFucCS is composed of {→3)-ß-GalNAcX-(1→4)-ß-GlcA-[(3→1)Y]-(1→}, where X = 4S (80%), 6S (10%) or nonsulfated (10%), Y = α-Fuc2,4S (40%), α-Fuc2,4S-(1→4)-α-Fuc (30%), or α-Fuc4S (30%), and S = SO3-. The anti-SARS-CoV-2 activity of PpFucCS and those of the FucCS and sulfated fucan isolated from Isostichopus badionotus (IbFucCS and IbSF) were compared with that of heparin. IC50 values demonstrated the activity of the three holothurian sulfated glycans to be ∼12 times more efficient than heparin, with no cytotoxic effects. The dissociation constant (KD) values obtained by surface plasmon resonance of the wildtype SARS-CoV-2 spike (S)-protein receptor-binding domain (RBD) and N501Y mutant RBD in interactions with the heparin-immobilized sensor chip were 94 and 1.8 × 103 nM, respectively. Competitive surface plasmon resonance inhibition analysis of PpFucCS, IbFucCS, and IbSF against heparin binding to wildtype S-protein showed IC50 values (in the nanomolar range) 6, 25, and 6 times more efficient than heparin, respectively. Data from computational simulations suggest an influence of the sulfation patterns of the Fuc units on hydrogen bonding with GlcA and that conformational change of some of the oligosaccharide structures occurs upon S-protein RBD binding. Compared with heparin, negligible anticoagulant action was observed for IbSF. Our results suggest that IbSF may represent a promising molecule for future investigations against SARS-CoV-2.


Subject(s)
Polysaccharides/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Sulfates/chemistry , Animals , Binding Sites , COVID-19/pathology , COVID-19/virology , Chondroitin Sulfates/chemistry , Chondroitin Sulfates/metabolism , Kinetics , Molecular Docking Simulation , Molecular Dynamics Simulation , Mutagenesis, Site-Directed , Partial Thromboplastin Time , Polysaccharides/chemistry , Protein Binding , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Sea Cucumbers/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Surface Plasmon Resonance
18.
Front Immunol ; 12: 638573, 2021.
Article in English | MEDLINE | ID: covidwho-1376694

ABSTRACT

Animal viruses are parasites of animal cells that have characteristics such as heredity and replication. Viruses can be divided into non-enveloped and enveloped viruses if a lipid bilayer membrane surrounds them or not. All the membrane proteins of enveloped viruses that function in attachment to target cells or membrane fusion are modified by glycosylation. Glycosylation is one of the most common post-translational modifications of proteins and plays an important role in many biological behaviors, such as protein folding and stabilization, virus attachment to target cell receptors and inhibition of antibody neutralization. Glycans of the host receptors can also regulate the attachment of the viruses and then influence the virus entry. With the development of glycosylation research technology, the research and development of novel virus vaccines and antiviral drugs based on glycan have received increasing attention. Here, we review the effects of host glycans and viral proteins on biological behaviors of viruses, and the opportunities for prevention and treatment of viral infectious diseases.


Subject(s)
Host-Parasite Interactions/physiology , Polysaccharides/metabolism , Receptors, Virus/metabolism , Virus Internalization , Viruses , Animals , Glycosylation , Humans , Immune Evasion , Viruses/immunology , Viruses/metabolism
19.
J Gen Virol ; 102(8)2021 08.
Article in English | MEDLINE | ID: covidwho-1368372

ABSTRACT

Infectious bronchitis virus (IBV) is an economically important coronavirus, causing damaging losses to the poultry industry worldwide as the causative agent of infectious bronchitis. The coronavirus spike (S) glycoprotein is a large type I membrane protein protruding from the surface of the virion, which facilitates attachment and entry into host cells. The IBV S protein is cleaved into two subunits, S1 and S2, the latter of which has been identified as a determinant of cellular tropism. Recent studies expressing coronavirus S proteins in mammalian and insect cells have identified a high level of glycosylation on the protein's surface. Here we used IBV propagated in embryonated hens' eggs to explore the glycan profile of viruses derived from infection in cells of the natural host, chickens. We identified multiple glycan types on the surface of the protein and found a strain-specific dependence on complex glycans for recognition of the S2 subunit by a monoclonal antibody in vitro, with no effect on viral replication following the chemical inhibition of complex glycosylation. Virus neutralization by monoclonal or polyclonal antibodies was not affected. Following analysis of predicted glycosylation sites for the S protein of four IBV strains, we confirmed glycosylation at 18 sites by mass spectrometry for the pathogenic laboratory strain M41-CK. Further characterization revealed heterogeneity among the glycans present at six of these sites, indicating a difference in the glycan profile of individual S proteins on the IBV virion. These results demonstrate a non-specific role for complex glycans in IBV replication, with an indication of an involvement in antibody recognition but not neutralisation.


Subject(s)
Coronavirus/physiology , Polysaccharides/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Alkaloids/chemistry , Alkaloids/pharmacology , Amino Acid Sequence , Animals , Binding Sites , Cells, Cultured , Chromatography, Liquid , Computational Biology/methods , Coronavirus/drug effects , Coronavirus Infections/veterinary , Gene Expression Regulation, Viral , Glycosylation/drug effects , Infectious bronchitis virus/physiology , Models, Molecular , Molecular Conformation , Molecular Weight , Neutralization Tests , Oligosaccharides/chemistry , Oligosaccharides/metabolism , Polysaccharides/chemistry , Poultry Diseases/virology , Protein Transport , Spectrometry, Mass, Electrospray Ionization , Spike Glycoprotein, Coronavirus/genetics , Structure-Activity Relationship , Virus Replication/drug effects
20.
Nat Chem ; 13(10): 963-968, 2021 10.
Article in English | MEDLINE | ID: covidwho-1366819

ABSTRACT

SARS-CoV-2 infection is controlled by the opening of the spike protein receptor binding domain (RBD), which transitions from a glycan-shielded 'down' to an exposed 'up' state to bind the human angiotensin-converting enzyme 2 receptor and infect cells. While snapshots of the 'up' and 'down' states have been obtained by cryo-electron microscopy and cryo-electron tomagraphy, details of the RBD-opening transition evade experimental characterization. Here over 130 µs of weighted ensemble simulations of the fully glycosylated spike ectodomain allow us to characterize more than 300 continuous, kinetically unbiased RBD-opening pathways. Together with ManifoldEM analysis of cryo-electron microscopy data and biolayer interferometry experiments, we reveal a gating role for the N-glycan at position N343, which facilitates RBD opening. Residues D405, R408 and D427 also participate. The atomic-level characterization of the glycosylated spike activation mechanism provided herein represents a landmark study for ensemble pathway simulations and offers a foundation for understanding the fundamental mechanisms of SARS-CoV-2 viral entry and infection.


Subject(s)
Polysaccharides/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Cryoelectron Microscopy , Humans , Molecular Dynamics Simulation
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