Journal
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
Volume 32, Issue 7, Pages 1567-1574Publisher
AMER CHEMICAL SOC
DOI: 10.1021/jasms.0c00399
Keywords
NKG2A; higher-order structure; epitope mapping; HDX-MS; FPOP; protein footprinting
Funding
- Bristol Myers Squibb
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NKG2A, an immune checkpoint inhibitor, forms a heterodimer with CD94 and recognizes HLA-E antigen. Mass spectrometry techniques can provide insights into the structure of NKG2A/CD94 heterodimer and the mechanisms of antibody interactions with it.
NK group 2 member A (NKG2A), an immune checkpoint inhibitor, is an emerging therapeutic target in immunooncology. NKG2A forms a heterodimer with CD94 on the cell surface of NK and a subset of T cells and recognizes the nonclassical human leukocyte antigen (HLA-E) in humans. Therapeutic blocking antibodies that block the ligation between HLA-E and NKG2A/CD94 have been shown to enhance antitumor immunity in mice and humans. In this study, we illustrate the practical utilities of mass spectrometry (MS)-based protein footprinting in areas from reagent characterization to antibody epitope mapping. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) in the higher-order structure characterization of NKG2A in complex with CD94 provides novel insights into the conformational dynamics of NKG2A/CD94 heterodimer. To fully understand antibody/target interactions, we employed complementary protein footprinting methods, including HDX-MS and fast photochemical oxidation of proteins (FPOP)-MS, to determine the binding epitopes of therapeutic monoclonal antibodies targeting NKG2A. Such a combination approach provides molecular insights into the binding mechanisms of antibodies to NKG2A with high specificity, demonstrating the blockade of NKG2A/HLA-E interaction.
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