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2.
Anal Bioanal Chem ; 413(29): 7305-7318, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1460297

ABSTRACT

The spike protein of SARS-CoV-2, the virus responsible for the global pandemic of COVID-19, is an abundant, heavily glycosylated surface protein that plays a key role in receptor binding and host cell fusion, and is the focus of all current vaccine development efforts. Variants of concern are now circulating worldwide that exhibit mutations in the spike protein. Protein sequence and glycosylation variations of the spike may affect viral fitness, antigenicity, and immune evasion. Global surveillance of the virus currently involves genome sequencing, but tracking emerging variants should include quantitative measurement of changes in site-specific glycosylation as well. In this work, we used data-dependent acquisition (DDA) and data-independent acquisition (DIA) mass spectrometry to quantitatively characterize the five N-linked glycosylation sites of the glycoprotein standard alpha-1-acid glycoprotein (AGP), as well as the 22 sites of the SARS-CoV-2 spike protein. We found that DIA compared favorably to DDA in sensitivity, resulting in more assignments of low-abundance glycopeptides. However, the reproducibility across replicates of DIA-identified glycopeptides was lower than that of DDA, possibly due to the difficulty of reliably assigning low-abundance glycopeptides confidently. The differences in the data acquired between the two methods suggest that DIA outperforms DDA in terms of glycoprotein coverage but that overall performance is a balance of sensitivity, selectivity, and statistical confidence in glycoproteomics. We assert that these analytical and bioinformatics methods for assigning and quantifying glycoforms would benefit the process of tracking viral variants as well as for vaccine development.


Subject(s)
Glycomics/methods , Mass Spectrometry/methods , Proteomics/methods , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , COVID-19/virology , Glycosylation , Humans , Limit of Detection , Reproducibility of Results , Spike Glycoprotein, Coronavirus/metabolism
4.
STAR Protoc ; 1(3): 100214, 2020 12 18.
Article in English | MEDLINE | ID: covidwho-1386744

ABSTRACT

This protocol describes an integrated approach for analyzing site-specific N- and O-linked glycosylation of SARS-CoV-2 spike protein by mass spectrometry. Glycoproteomics analyzes intact glycopeptides to examine site-specific microheterogeneity of glycoproteins. Glycomics provides structural characterization on any glycan assignments by glycoproteomics. This procedure can be modified and applied to a variety of N- and/or O-linked glycoproteins. Combined with bioinformatics, the glycomics-informed glycoproteomics may be useful in generating 3D molecular dynamics simulations of certain glycoproteins alone or interacting with one another. For complete details on the use and execution of this protocol, please refer to Zhao et al. (2020).


Subject(s)
Glycomics/methods , Glycoproteins , Proteomics/methods , Spike Glycoprotein, Coronavirus , COVID-19/virology , Glycoproteins/analysis , Glycoproteins/chemistry , Glycoproteins/metabolism , Glycosylation , Humans , Spike Glycoprotein, Coronavirus/analysis , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
5.
Nat Methods ; 18(1): 28, 2021 01.
Article in English | MEDLINE | ID: covidwho-1065919
6.
J Biol Chem ; 296: 100375, 2021.
Article in English | MEDLINE | ID: covidwho-1062444

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged during the last months of 2019, spreading throughout the world as a highly transmissible infectious illness designated as COVID-19. Vaccines have now appeared, but the challenges in producing sufficient material and distributing them around the world means that effective treatments to limit infection and improve recovery are still urgently needed. This review focuses on the relevance of different glycobiological molecules that could potentially serve as or inspire therapeutic tools during SARS-CoV-2 infection. As such, we highlight the glycobiology of the SARS-CoV-2 infection process, where glycans on viral proteins and on host glycosaminoglycans have critical roles in efficient infection. We also take notice of the glycan-binding proteins involved in the infective capacity of virus and in human defense. In addition, we critically evaluate the glycobiological contribution of candidate drugs for COVID-19 therapy such as glycans for vaccines, anti-glycan antibodies, recombinant lectins, lectin inhibitors, glycosidase inhibitors, polysaccharides, and numerous glycosides, emphasizing some opportunities to repurpose FDA-approved drugs. For the next-generation drugs suggested here, biotechnological engineering of new probes to block the SARS-CoV-2 infection might be based on the essential glycobiological insight on glycosyltransferases, glycans, glycan-binding proteins, and glycosidases related to this pathology.


Subject(s)
Antiviral Agents/therapeutic use , COVID-19/prevention & control , Drug Repositioning , Glycoside Hydrolase Inhibitors/therapeutic use , Glycosyltransferases/antagonists & inhibitors , Viral Proteins/antagonists & inhibitors , Antibodies, Neutralizing/therapeutic use , Antiviral Agents/chemistry , COVID-19/epidemiology , COVID-19/immunology , COVID-19/virology , Drug Design , Drug Discovery , Gene Expression , Glycomics/methods , Glycosaminoglycans/chemistry , Glycosaminoglycans/immunology , Glycosaminoglycans/metabolism , Glycosyltransferases/chemistry , Glycosyltransferases/genetics , Glycosyltransferases/immunology , Host-Pathogen Interactions/drug effects , Host-Pathogen Interactions/genetics , Host-Pathogen Interactions/immunology , Humans , Lectins/chemistry , Lectins/immunology , Lectins/metabolism , Polysaccharides/chemistry , Polysaccharides/immunology , Polysaccharides/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Signal Transduction , Viral Proteins/chemistry , Viral Proteins/genetics , Viral Proteins/immunology
7.
J Am Soc Mass Spectrom ; 31(10): 2013-2024, 2020 Oct 07.
Article in English | MEDLINE | ID: covidwho-744341

ABSTRACT

As corona virus disease 2019 (COVID-19) is a rapidly growing public health crisis across the world, our knowledge of meaningful diagnostic tests and treatment for severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) is still evolving. This novel coronavirus disease COVID-19 can be diagnosed using RT-PCR, but inadequate access to reagents, equipment, and a nonspecific target has slowed disease detection and management. Precision medicine, individualized patient care, requires suitable diagnostics approaches to tackle the challenging aspects of viral outbreaks where many tests are needed in a rapid and deployable approach. Mass spectrometry (MS)-based technologies such as proteomics, glycomics, lipidomics, and metabolomics have been applied in disease outbreaks for identification of infectious disease agents such as virus and bacteria and the molecular phenomena associated with pathogenesis. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF/MS) is widely used in clinical diagnostics in the United States and Europe for bacterial pathogen identification. Paper spray ionization mass spectrometry (PSI-MS), a rapid ambient MS technique, has recently open a new opportunity for future clinical investigation to diagnose pathogens. Ultra-high-pressure liquid chromatography coupled high-resolution mass spectrometry (UHPLC-HRMS)-based metabolomics and lipidomics have been employed in large-scale biomedical research to discriminate infectious pathogens and uncover biomarkers associated with pathogenesis. PCR-MS has emerged as a new technology with the capability to directly identify known pathogens from the clinical specimens and the potential to identify genetic evidence of undiscovered pathogens. Moreover, miniaturized MS offers possible applications with relatively fast, highly sensitive, and potentially portable ways to analyze for viral compounds. However, beneficial aspects of these rapidly growing MS technologies in pandemics like COVID-19 outbreaks has been limited. Hence, this perspective gives a brief of the existing knowledge, current challenges, and opportunities for MS-based techniques as a promising avenue in studying emerging pathogen outbreaks such as COVID-19.


Subject(s)
Coronavirus Infections/etiology , Lipidomics/methods , Mass Spectrometry/methods , Metabolomics/methods , Pneumonia, Viral/etiology , Proteomics/methods , COVID-19 , COVID-19 Testing , Chromatography, High Pressure Liquid , Clinical Laboratory Techniques , Coronavirus Infections/diagnosis , Glycomics/methods , Humans , Pandemics , Polymerase Chain Reaction , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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