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Signature of antibody domain exchange by native mass spectrometry and collision-induced unfolding

DOI: 10.1021/acs.analchem.8b00573 DOI Help

Authors: Yasunori Watanabe (Oxford Glycobiology Institute, University of Oxford; University of Southampton; Wellcome Centre for Human Genetics) , Snezana Vasiljevic (Oxford Glycobiology Institute, University of Oxford) , Joel D. Allen (University of Southampton) , Gemma E. Seabright (Oxford Glycobiology Institute, University of Oxford; University of Southampton) , Helen M. E. Duyvesteyn (University of Oxford; Diamond Light Source) , Katie J. Doores (King’s College London) , Max Crispin (University of Southampton) , Weston B. Struwe (Oxford Glycobiology Institute, University of Oxford)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Analytical Chemistry

State: Published (Approved)
Published: May 2018

Abstract: The development of domain-exchanged antibodies offers a route to high-affinity targeting to clustered multivalent epitopes, such as those associated with viral infections and many cancers. One strategy to generate these antibodies is to introduce mutations into target antibodies to drive domain exchange using the only known naturally occurring domain-exchanged anti-HIV (anti-human immunodeficiency virus) IgG1 antibody, 2G12, as a template. Here, we show that domain exchange can be sensitively monitored by ion-mobility mass spectrometry and gas-phase collision-induced unfolding. Using native 2G12 and a mutated form that disrupts domain exchange such that it has a canonical IgG1 architecture (2G12 I19R), we show that the two forms can be readily distinguished by their unfolding profiles. Importantly, the same signature of domain exchange is observed for both intact antibody and isolated Fab fragments. The development of a mass spectrometric method to detect antibody domain exchange will enable rapid screening and selection of candidate antibodies engineered to exhibit this and other unusual quaternary antibody architectures.

Subject Areas: Biology and Bio-materials, Chemistry

Technical Areas: