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Mass spectrometry-based mapping of conformational epitopes on SARS-CoV-2 antigens targeted by monoclonal antibodies.

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Τίτλος: Mass spectrometry-based mapping of conformational epitopes on SARS-CoV-2 antigens targeted by monoclonal antibodies.
Συγγραφείς: Pianpaktr P; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: p.pianpaktr@gmail.com., Ramanandana P; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Department of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Huachiew Chalermprakiet University, Bang Phli, Samut Prakan, 10540, Thailand. Electronic address: waew2.pr@gmail.com., Nanthawong S; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: saharat.nanthawongb@gmail.com., Wongtangprasert T; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; The Excellence Chulalongkorn Comprehensive Cancer Center, King Chulalongkorn Memorial Hospital, Pathumwan, Bangkok, 10330, Thailand. Electronic address: cassavara@gmail.com., Saelao P; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: p.saelao11@gmail.com., Muanwien P; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: phijitra.jame7@gmail.com., Boonkrai C; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: ctkorn@gmail.com., Cicek H; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: hasan.cicek@saromics.com., Focht D; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: dorota.focht@saromics.com., Kimbung R; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: Raymond.kimbung@saromics.com., Welin M; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: martin.welin@saromics.com., Stahl VL; Department of Chemistry, University of Massachusetts Amherst, Amherst, MA, 01003, United States. Electronic address: vstahl@umass.edu., Graban EM; QuarryBio Inc., Bloomington, IN, 47404, United States. Electronic address: eric.graban@reclaim-rx.com., Vachet RW; Department of Chemistry, University of Massachusetts Amherst, Amherst, MA, 01003, United States. Electronic address: rwvachet@chem.umass.edu., Pisitkun T; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Division of Research Affairs, Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: trairak@gmail.com., Limpikirati PK; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: patanachai.l@pharm.chula.ac.th.
Πηγή: International journal of biological macromolecules [Int J Biol Macromol] 2026 Aug; Vol. 372, pp. 153102. Date of Electronic Publication: 2026 Jun 21.
Τύπος έκδοσης: Journal Article
Γλώσσα: English
Στοιχεία περιοδικού: Publisher: Elsevier Country of Publication: Netherlands NLM ID: 7909578 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1879-0003 (Electronic) Linking ISSN: 01418130 NLM ISO Abbreviation: Int J Biol Macromol Subsets: MEDLINE
Imprint Name(s): Publication: Amsterdam : Elsevier
Original Publication: Guildford, Eng., IPC Science and Technology Press.
Ιατρικοί όροι (MeSH): Antibodies, Monoclonal*/immunology , Antibodies, Monoclonal*/chemistry , Epitope Mapping*/methods , Spike Glycoprotein, Coronavirus*/immunology , Spike Glycoprotein, Coronavirus*/chemistry , Epitopes*/immunology , Epitopes*/chemistry , SARS-CoV-2*/immunology , SARS-CoV-2*/chemistry , Antigens, Viral*/immunology , Antigens, Viral*/chemistry , Antibodies, Viral*/immunology, Mass Spectrometry/methods ; Humans ; Tandem Mass Spectrometry ; Models, Molecular
Περίληψη: The spike glycoprotein of SARS-CoV-2, particularly its receptor-binding domain (RBD), is a key target for therapeutic monoclonal antibodies (mAbs). Epitope mapping is therefore critical for the development of effective antiviral therapeutics. In this study, diethylpyrocarbonate covalent labeling mass spectrometry (DEPC CL-MS) was applied to map epitopes on the beta (B.1.351) and omicron (B.1.1.529) variants of the SARS-CoV-2 RBD, as well as on the original SARS-CoV-2 spike S1 subunit, in complex with anti-SARS-CoV-2 mAbs. Combined with bottom-up LC-MS/MS, DEPC labeling enabled site-specific identification of residues exhibiting significant modification changes. Clustering of these residues on the protein surface identified potential epitopes for the 1D1 mAb on the RBDs and for the 1D3 mAb on the C-terminal domain of the S1 subunit. Structural interpretation was supported by available experimental data, with AlphaFold models providing supplementary context where needed. Thus, "theory guides, but experiment decides": AlphaFold aided epitope identification when high-resolution antigen-antibody complex structures were unavailable, but conclusions were ultimately resolved experimentally. Together, these findings establish DEPC CL-MS as a useful and complementary approach for epitope mapping, providing residue-level insights into antigen-antibody interactions and advancing structural understanding for antiviral mAb development.
(Copyright © 2026 The Authors. Published by Elsevier B.V. All rights reserved.)
Competing Interests: Declaration of competing interest Eric M. Graban is an employee of QuarryBio Inc., which is commercializing DEPC CL-MS methods to characterize the higher order structure of protein therapeutics.
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Grant Information: R35 GM145272 United States GM NIGMS NIH HHS
Contributed Indexing: Keywords: Covalent labeling mass spectrometry; Epitope mapping; SARS-CoV-2 spike protein
Substance Nomenclature: 0 (Antibodies, Monoclonal)
0 (Spike Glycoprotein, Coronavirus)
0 (Epitopes)
0 (spike protein, SARS-CoV-2)
0 (Antigens, Viral)
0 (Antibodies, Viral)
Entry Date(s): Date Created: 20260621 Date Completed: 20260723 Latest Revision: 20260922
Update Code: 20260922
PubMed Central ID: PMC13359033
DOI: 10.1016/j.ijbiomac.2026.153102
PMID: 42324012
Βάση Δεδομένων: MEDLINE
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  Data: Mass spectrometry-based mapping of conformational epitopes on SARS-CoV-2 antigens targeted by monoclonal antibodies.
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  Data: <searchLink fieldCode="AU" term="%22Pianpaktr+P%22">Pianpaktr P</searchLink>; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: p.pianpaktr@gmail.com.<br /><searchLink fieldCode="AU" term="%22Ramanandana+P%22">Ramanandana P</searchLink>; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Department of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Huachiew Chalermprakiet University, Bang Phli, Samut Prakan, 10540, Thailand. Electronic address: waew2.pr@gmail.com.<br /><searchLink fieldCode="AU" term="%22Nanthawong+S%22">Nanthawong S</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: saharat.nanthawongb@gmail.com.<br /><searchLink fieldCode="AU" term="%22Wongtangprasert+T%22">Wongtangprasert T</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; The Excellence Chulalongkorn Comprehensive Cancer Center, King Chulalongkorn Memorial Hospital, Pathumwan, Bangkok, 10330, Thailand. Electronic address: cassavara@gmail.com.<br /><searchLink fieldCode="AU" term="%22Saelao+P%22">Saelao P</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: p.saelao11@gmail.com.<br /><searchLink fieldCode="AU" term="%22Muanwien+P%22">Muanwien P</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: phijitra.jame7@gmail.com.<br /><searchLink fieldCode="AU" term="%22Boonkrai+C%22">Boonkrai C</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: ctkorn@gmail.com.<br /><searchLink fieldCode="AU" term="%22Cicek+H%22">Cicek H</searchLink>; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: hasan.cicek@saromics.com.<br /><searchLink fieldCode="AU" term="%22Focht+D%22">Focht D</searchLink>; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: dorota.focht@saromics.com.<br /><searchLink fieldCode="AU" term="%22Kimbung+R%22">Kimbung R</searchLink>; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: Raymond.kimbung@saromics.com.<br /><searchLink fieldCode="AU" term="%22Welin+M%22">Welin M</searchLink>; SARomics Biostructures AB, Medicon Village, Lund, 223 81, Sweden. Electronic address: martin.welin@saromics.com.<br /><searchLink fieldCode="AU" term="%22Stahl+VL%22">Stahl VL</searchLink>; Department of Chemistry, University of Massachusetts Amherst, Amherst, MA, 01003, United States. Electronic address: vstahl@umass.edu.<br /><searchLink fieldCode="AU" term="%22Graban+EM%22">Graban EM</searchLink>; QuarryBio Inc., Bloomington, IN, 47404, United States. Electronic address: eric.graban@reclaim-rx.com.<br /><searchLink fieldCode="AU" term="%22Vachet+RW%22">Vachet RW</searchLink>; Department of Chemistry, University of Massachusetts Amherst, Amherst, MA, 01003, United States. Electronic address: rwvachet@chem.umass.edu.<br /><searchLink fieldCode="AU" term="%22Pisitkun+T%22">Pisitkun T</searchLink>; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Division of Research Affairs, Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: trairak@gmail.com.<br /><searchLink fieldCode="AU" term="%22Limpikirati+PK%22">Limpikirati PK</searchLink>; Department of Food and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Pharmaceutical Sciences and Technology (PST) Graduate Program, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand; Center of Excellence in Systems Biology (CUSB), Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand. Electronic address: patanachai.l@pharm.chula.ac.th.
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  Data: <searchLink fieldCode="MM" term="%22Antibodies%2C+Monoclonal%22">Antibodies, Monoclonal*</searchLink>/<searchLink fieldCode="MM" term="%22Antibodies%2C+Monoclonal+immunology%22">immunology</searchLink> <br /><searchLink fieldCode="MM" term="%22Antibodies%2C+Monoclonal%22">Antibodies, Monoclonal*</searchLink>/<searchLink fieldCode="MM" term="%22Antibodies%2C+Monoclonal+chemistry%22">chemistry</searchLink> <br /><searchLink fieldCode="MM" term="%22Epitope+Mapping%22">Epitope Mapping*</searchLink>/<searchLink fieldCode="MM" term="%22Epitope+Mapping+methods%22">methods</searchLink> <br /><searchLink fieldCode="MM" term="%22Spike+Glycoprotein%2C+Coronavirus%22">Spike Glycoprotein, Coronavirus*</searchLink>/<searchLink fieldCode="MM" term="%22Spike+Glycoprotein%2C+Coronavirus+immunology%22">immunology</searchLink> <br /><searchLink fieldCode="MM" term="%22Spike+Glycoprotein%2C+Coronavirus%22">Spike Glycoprotein, Coronavirus*</searchLink>/<searchLink fieldCode="MM" term="%22Spike+Glycoprotein%2C+Coronavirus+chemistry%22">chemistry</searchLink> <br /><searchLink fieldCode="MM" term="%22Epitopes%22">Epitopes*</searchLink>/<searchLink fieldCode="MM" term="%22Epitopes+immunology%22">immunology</searchLink> <br /><searchLink fieldCode="MM" term="%22Epitopes%22">Epitopes*</searchLink>/<searchLink fieldCode="MM" term="%22Epitopes+chemistry%22">chemistry</searchLink> <br /><searchLink fieldCode="MM" term="%22SARS-CoV-2%22">SARS-CoV-2*</searchLink>/<searchLink fieldCode="MM" term="%22SARS-CoV-2+immunology%22">immunology</searchLink> <br /><searchLink fieldCode="MM" term="%22SARS-CoV-2%22">SARS-CoV-2*</searchLink>/<searchLink fieldCode="MM" term="%22SARS-CoV-2+chemistry%22">chemistry</searchLink> <br /><searchLink fieldCode="MM" term="%22Antigens%2C+Viral%22">Antigens, Viral*</searchLink>/<searchLink fieldCode="MM" term="%22Antigens%2C+Viral+immunology%22">immunology</searchLink> <br /><searchLink fieldCode="MM" term="%22Antigens%2C+Viral%22">Antigens, Viral*</searchLink>/<searchLink fieldCode="MM" term="%22Antigens%2C+Viral+chemistry%22">chemistry</searchLink> <br /><searchLink fieldCode="MM" term="%22Antibodies%2C+Viral%22">Antibodies, Viral*</searchLink>/<searchLink fieldCode="MM" term="%22Antibodies%2C+Viral+immunology%22">immunology</searchLink><br /><searchLink fieldCode="MH" term="%22Mass+Spectrometry%22">Mass Spectrometry</searchLink>/<searchLink fieldCode="MH" term="%22Mass+Spectrometry+methods%22">methods</searchLink> ; <searchLink fieldCode="MH" term="%22Humans%22">Humans</searchLink> ; <searchLink fieldCode="MH" term="%22Tandem+Mass+Spectrometry%22">Tandem Mass Spectrometry</searchLink> ; <searchLink fieldCode="MH" term="%22Models%2C+Molecular%22">Models, Molecular</searchLink>
– Name: Abstract
  Label: Abstract
  Group: Ab
  Data: The spike glycoprotein of SARS-CoV-2, particularly its receptor-binding domain (RBD), is a key target for therapeutic monoclonal antibodies (mAbs). Epitope mapping is therefore critical for the development of effective antiviral therapeutics. In this study, diethylpyrocarbonate covalent labeling mass spectrometry (DEPC CL-MS) was applied to map epitopes on the beta (B.1.351) and omicron (B.1.1.529) variants of the SARS-CoV-2 RBD, as well as on the original SARS-CoV-2 spike S<subscript>1</subscript> subunit, in complex with anti-SARS-CoV-2 mAbs. Combined with bottom-up LC-MS/MS, DEPC labeling enabled site-specific identification of residues exhibiting significant modification changes. Clustering of these residues on the protein surface identified potential epitopes for the 1D1 mAb on the RBDs and for the 1D3 mAb on the C-terminal domain of the S<subscript>1</subscript> subunit. Structural interpretation was supported by available experimental data, with AlphaFold models providing supplementary context where needed. Thus, "theory guides, but experiment decides": AlphaFold aided epitope identification when high-resolution antigen-antibody complex structures were unavailable, but conclusions were ultimately resolved experimentally. Together, these findings establish DEPC CL-MS as a useful and complementary approach for epitope mapping, providing residue-level insights into antigen-antibody interactions and advancing structural understanding for antiviral mAb development.<br /> (Copyright © 2026 The Authors. Published by Elsevier B.V. All rights reserved.)
– Name: Abstract
  Label: Competing Interests
  Group: Ab
  Data: Declaration of competing interest Eric M. Graban is an employee of QuarryBio Inc., which is commercializing DEPC CL-MS methods to characterize the higher order structure of protein therapeutics.
– Name: Ref
  Label: References
  Group: RefInfo
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  Data: <i>Keywords: </i>Covalent labeling mass spectrometry; Epitope mapping; SARS-CoV-2 spike protein
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  Data: 10.1016/j.ijbiomac.2026.153102
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        Text: English
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      – SubjectFull: Mass Spectrometry methods
        Type: general
      – SubjectFull: Humans
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      – SubjectFull: Tandem Mass Spectrometry
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      – SubjectFull: Spike Glycoprotein, Coronavirus chemistry
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      – SubjectFull: Epitopes immunology
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              Text: 2026 Aug
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            – TitleFull: International journal of biological macromolecules
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