Immunocomplexed Antigen Capture and Identification by Native Top-Down Mass Spectrometry

John P. McGee, Rafael D. Melani, Ben Des Soye, Derek Croote, Valerie Winton, Stephen R. Quake, Jared O. Kafader, Neil L. Kelleher*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Antibody-antigen interactions are central to the immune response. Variation of protein antigens such as isoforms and post-translational modifications can alter their antibody binding sites. To directly connect the recognition of protein antigens with their molecular composition, we probed antibody-antigen complexes by using native tandem mass spectrometry. Specifically, we characterized the prominent peanut allergen Ara h 2 and a convergent IgE variable region discovered in patients who are allergic to peanuts. In addition to measuring the antigen-induced dimerization of IgE antibodies, we demonstrated how immunocomplexes can be isolated in the gas phase and activated to eject, identify, and characterize proteoforms of their bound antigens. Using tandem experiments, we isolated the ejected antigens and then fragmented them to identify their chemical composition. These results establish native top-down mass spectrometry as a viable platform for precise and thorough characterization of immunocomplexes to relate structure to function and enable the discovery of antigen proteoforms and their binding sites.

Original languageEnglish (US)
Pages (from-to)2093-2097
Number of pages5
JournalJournal of the American Society for Mass Spectrometry
Volume34
Issue number10
DOIs
StatePublished - Oct 4 2023

Keywords

  • Orbitrap
  • antibody
  • antigen
  • complex-up
  • native
  • top-down

ASJC Scopus subject areas

  • Structural Biology
  • Spectroscopy

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