Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species

Abstract The understanding and characterization of protein interactions is crucial for elucidation of complicated biomolecular processes as well as for the development of new biopharmaceutical therapies. Often, protein interactions involve multiple binding, avidity, oligomerization, and are dependen...

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Autores principales:	Morten E. Pedersen, Ragna M. S. Haegebaert, Jesper Østergaard, Henrik Jensen
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/11861b6c148c485c88c61ddeddb22054
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spelling	oai:doaj.org-article:11861b6c148c485c88c61ddeddb220542021-12-02T13:35:05ZSize-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species10.1038/s41598-021-84113-z2045-2322https://doaj.org/article/11861b6c148c485c88c61ddeddb220542021-02-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-84113-zhttps://doaj.org/toc/2045-2322Abstract The understanding and characterization of protein interactions is crucial for elucidation of complicated biomolecular processes as well as for the development of new biopharmaceutical therapies. Often, protein interactions involve multiple binding, avidity, oligomerization, and are dependent on the local environment. Current analytical methodologies are unable to provide a detailed mechanistic characterization considering all these parameters, since they often rely on surface immobilization, cannot measure under biorelevant conditions, or do not feature a structurally-related readout for indicating formation of multiple bound species. In this work, we report the use of flow induced dispersion analysis (FIDA) for in-solution characterization of complex protein interactions under in vivo like conditions. FIDA is an immobilization-free ligand binding methodology employing Taylor dispersion analysis for measuring the hydrodynamic radius (size) of biomolecular complexes. Here, the FIDA technology is utilized for a size-based characterization of the interaction between TNF-α and adalimumab. We report concentration-dependent complex sizes, binding affinities (K d), kinetics, and higher order stoichiometries, thus providing essential information on the TNF-α–adalimumab binding mechanism. Furthermore, it is shown that the avidity stabilized complexes involving formation of multiple non-covalent bonds are formed on a longer timescale than the primary complexes formed in a simple 1 to 1 binding event.Morten E. PedersenRagna M. S. HaegebaertJesper ØstergaardHenrik JensenNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-10 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Morten E. Pedersen Ragna M. S. Haegebaert Jesper Østergaard Henrik Jensen Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
description	Abstract The understanding and characterization of protein interactions is crucial for elucidation of complicated biomolecular processes as well as for the development of new biopharmaceutical therapies. Often, protein interactions involve multiple binding, avidity, oligomerization, and are dependent on the local environment. Current analytical methodologies are unable to provide a detailed mechanistic characterization considering all these parameters, since they often rely on surface immobilization, cannot measure under biorelevant conditions, or do not feature a structurally-related readout for indicating formation of multiple bound species. In this work, we report the use of flow induced dispersion analysis (FIDA) for in-solution characterization of complex protein interactions under in vivo like conditions. FIDA is an immobilization-free ligand binding methodology employing Taylor dispersion analysis for measuring the hydrodynamic radius (size) of biomolecular complexes. Here, the FIDA technology is utilized for a size-based characterization of the interaction between TNF-α and adalimumab. We report concentration-dependent complex sizes, binding affinities (K d), kinetics, and higher order stoichiometries, thus providing essential information on the TNF-α–adalimumab binding mechanism. Furthermore, it is shown that the avidity stabilized complexes involving formation of multiple non-covalent bonds are formed on a longer timescale than the primary complexes formed in a simple 1 to 1 binding event.
format	article
author	Morten E. Pedersen Ragna M. S. Haegebaert Jesper Østergaard Henrik Jensen
author_facet	Morten E. Pedersen Ragna M. S. Haegebaert Jesper Østergaard Henrik Jensen
author_sort	Morten E. Pedersen
title	Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
title_short	Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
title_full	Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
title_fullStr	Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
title_full_unstemmed	Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
title_sort	size-based characterization of adalimumab and tnf-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/11861b6c148c485c88c61ddeddb22054
work_keys_str_mv	AT mortenepedersen sizebasedcharacterizationofadalimumabandtnfainteractionsusingflowinduceddispersionanalysisassessmentofaviditystabilizedmultipleboundspecies AT ragnamshaegebaert sizebasedcharacterizationofadalimumabandtnfainteractionsusingflowinduceddispersionanalysisassessmentofaviditystabilizedmultipleboundspecies AT jesperøstergaard sizebasedcharacterizationofadalimumabandtnfainteractionsusingflowinduceddispersionanalysisassessmentofaviditystabilizedmultipleboundspecies AT henrikjensen sizebasedcharacterizationofadalimumabandtnfainteractionsusingflowinduceddispersionanalysisassessmentofaviditystabilizedmultipleboundspecies
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Size-based characterization of adalimumab and TNF-α interactions using flow induced dispersion analysis: assessment of avidity-stabilized multiple bound species

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