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Probing Antibody Structures by Hydrogen/Deuterium Exchange Mass Spectrometry

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Mass Spectrometry-Based Proteomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2718))

Abstract

Hydrogen/deuterium exchange (HDX) followed by mass spectrometry detection (MS) provides a fast, reliable, and detailed solution for the assessment of a protein structure. It has been widely recognized as an indispensable tool and already approved by several regulatory agencies as a structural technique for the validation of protein biopharmaceuticals, including antibody-based drugs. Antibodies are of a key importance in life and medical sciences but considered to be challenging analytical targets because of their compact structure stabilized by disulfide bonds and due to the presence of glycosylation. Despite these difficulties, there are already numerous excellent studies describing MS-based antibody structure characterization. In this chapter, we describe a universal HDX-MS workflow. Deeper attention is paid to sample handling, optimization procedures, and feasibility stages, as these elements of the HDX experiment are crucial for obtaining reliable detailed and spatially well-resolved information.

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Acknowledgments

Support from Horizon 2020, EPIC-XS (project number—82383), and Czech Science Foundation 22-27695S is gratefully acknowledged. Additional support was obtained from EU/MEYS projects BioCeV (CZ.1.05/1.1.00/02.0109). L.F. also thanks GAUK project 359221.

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Authors and Affiliations

  1. BioCeV - Institute of Microbiology of the Czech Academy of Sciences, Vestec, Czech Republic

    Zuzana Kalaninová, Lukáš Fojtík, Petr Novák, Michael Volný & Petr Man

  2. Department of Biochemistry, Faculty of Science, Charles University, Prague, Czech Republic

    Zuzana Kalaninová & Lukáš Fojtík

  3. Institute of Microbiology of the Czech Academy of Sciences, Prague, Czech Republic

    Josef Chmelík

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  1. Zuzana Kalaninová
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  2. Lukáš Fojtík
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  3. Josef Chmelík
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  4. Petr Novák
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  5. Michael Volný
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  6. Petr Man
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Correspondence to Petr Man .

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  1. VIB-UGent Center for Medical Biotechnology, Ghent, Belgium

    Kris Gevaert

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Kalaninová, Z., Fojtík, L., Chmelík, J., Novák, P., Volný, M., Man, P. (2023). Probing Antibody Structures by Hydrogen/Deuterium Exchange Mass Spectrometry. In: Gevaert, K. (eds) Mass Spectrometry-Based Proteomics. Methods in Molecular Biology, vol 2718. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3457-8_17

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  • DOI: https://doi.org/10.1007/978-1-0716-3457-8_17

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