Epitope and Paratope Mapping of PD-1/Nivolumab by Mass Spectrometry-Based Hydrogen–Deuterium Exchange, Cross-linking, and Molecular Docking

• Published in: Analytical chemistry (Washington) 2020-07, Vol.92 (13), p.9086-9094
• Main Authors: Zhang, Mengru Mira, Huang, Richard Y.-C, Beno, Brett R, Deyanova, Ekaterina G, Li, Jing, Chen, Guodong, Gross, Michael L
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino acids; Antibodies; Antigen-Antibody Complex - chemistry; Antigens; Apoptosis; Binding; Binding Sites; Cell death; Chemistry; Chromatography, High Pressure Liquid; Crosslinking; Crystal structure; Deuterium; Deuterium Exchange Measurement; Epitope mapping; Epitope Mapping - methods; Epitopes - analysis; Epitopes - chemistry; Epitopes - immunology; Exchanging; Humans; Hydrogen-deuterium exchange; Immune checkpoint; Immunotherapy; Mapping; Mass spectrometry; Mass spectroscopy; Molecular docking; Molecular Docking Simulation; Monoclonal antibodies; N-Terminus; Nivolumab - immunology; Nivolumab - metabolism; PD-1 protein; Programmed Cell Death 1 Receptor - chemistry; Programmed Cell Death 1 Receptor - immunology; Programmed Cell Death 1 Receptor - metabolism; Protein Binding; Protein Structure, Secondary; Scientific imaging; Spectroscopy; Tandem Mass Spectrometry
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/acs.analchem.0c01291

Programmed cell death-1 (PD-1), an antigen co-receptor on cell surfaces, is one of the conspicuous immune checkpoints. Nivolumab, a monoclonal antibody therapeutic approved by the FDA, binds to PD-1 and efficiently blocks its pathways. In this study, an integrated approach was developed to map the epitope/paratope of PD-1/nivolumab. The approach includes hydrogen–deuterium exchange mass spectrometry (HDX-MS) followed by electron-transfer dissociation (ETD), chemical cross-linking, and molecular docking. HDX-ETD offers some binding-site characterization with amino acid resolution. Chemical cross-linking provides complementary information on one additional epitope (i.e., the BC-loop) and a potential paratope at the N-terminus of the heavy chain. Furthermore, cross-linking identifies another loop region (i.e., the C’D-loop) that undergoes a remote conformational change. The distance restraints derived from the cross-links enable building high-confidence models of PD-1/nivolumab, evaluated with respect to a resolved crystal structure. This integrated strategy is an opportunity to characterize comprehensively other antigen–antibody interactions, to enable the understanding of binding mechanisms, and to design future antibody therapeutics.