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Glycosylation analysis of SARS-CoV-2 S and hCoV-HKU1 S

ABSTRACT: The rapid global spread of SARS-CoV-2 and resultant mortality and social disruption have highlighted the need to better understand coronaviral immunity to expedite vaccine development efforts. Multiple candidate vaccines with the goal of eliciting protective neutralising antibodies targeting the viral spike glycoprotein are rapidly advancing to clinical trial. However, the immunogenic properties of the spike in human populations are unresolved. To address this, we undertook an in-depth characterisation of humoral and cellular immunity against SARS-CoV-2 spike in humans following mild to moderate SARS-CoV-2 infection. We find serological antibody responses against spike are routinely elicited by infection and correlate with plasma neutralising and ACE2 inhibitory activity. Expanded populations of spike-specific memory B cells and circulating T follicular helper cells (cTFH) were detected within convalescent donors, while responses to receptor binding domain constitute a minor fraction. Using regression analysis, we find high serum neutralisation activity was associated with increased spike-specific antibody, but notably also with the relative distribution of some spike-specific cTFH subsets. Thus both qualitative and quantitative features of B and T cell immunity to spike constitute informative biomarkers to assess protective potential of novel SARS-CoV-2 vaccines

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Coronavirus Predict Cov-19 (ncbitaxon:2042545)

SUBMITTER: Adam Wheatley

PROVIDER: MSV000085758 | MassIVE | Wed Jul 15 11:00:00 BST 2020

SECONDARY ACCESSION(S): PXD019163

REPOSITORIES: MassIVE

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Project description:Despite the clinical success of anti-spike vaccines, the effectiveness of neutralizing antibodies and vaccines by rapidly spreading SARS-CoV-2 variants has been compromised. Viruses can hijack the glycosylation machinery of host cells to shield themselves from the host’s immune response and attenuate antibody efficiency. However, it still remains unclear whether targeting glycosylation on spike can impair SARS-CoV-2 and its variants infectivity. Methods: To assess the binding ability of glycosylated or deglycosylated spike with ACE2, we performed flow cytometry, ELISA, and BioLayer Interferometry methods. Viral entry ability was determined by luciferase intensity, immunoblotting, and immunofluorescence assay. A genome-wide association study (GWAS) was performed to identify the relationship of STT3A and COVID-19 severity. N-glycosylation regulated by NF-kB/STT3A axis was investigated by knockdown approach, chromatin immunoprecipitation, and promoter assay. To specifically target SARS-CoV-2 infected cells, we developed an antibody-drug conjugate coupling non-neutralization anti-spike antibody with NGI-1 (4G10-ADC) on inhibitory effects of SARS-CoV-2 infection. Results: We found receptor binding domain and three SARS-CoV-2 distinct surface Nglycosylation sites in 57,311 spikes retrieved from NCBI-Virus-database are highly evolutionarily conserved (99.67%) and involved in ACE2 interaction. We further identified STT3A as a key glycosyltransferase that catalyzed spike glycosylation and positively correlated with COVID-19 severity. Inhibition of STT3A by N-linked glycosylation inhibitor-1 (NGI-1) impaired SARS-CoV-2 and its variants (B.1.1.7, and B.1.351) infectivity. Most importantly, 4G10-ADC internalized SARS-CoV-2 infected cells and subsequently released NGI-1 to deglycosylate spike protein. Thereby, it reinforces the neutralizing abilities in antibodies, vaccines, or convalescent sera, inhibiting SARS-CoV-2 and its variants’ infectivity. Our results suggest targeting STT3A-mediated evolution conserved glycosylation via ADC can provide a widespread impact on SARS-CoV-2 variants infection. Together, we identified a novel deglycosylation method to eradicate SARS-CoV-2 variants infection.

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Glycosylation analysis of SARS-CoV-2 S and hCoV-HKU1 S

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