Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)

STD NMR spectroscopy is a powerful ligand-observed NMR tool for screening and characterizing the interactions of small molecules and low molecular weight fragments with a given macromolecule, identifying the main intermolecular contacts in the bound state. It is also a powerful analytical technique...

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Autores: Rocha, Gabriel, Ramírez-Cárdenas, Jonathan, Padilla-Pérez, M Carmen, Walpole, Samuel, Nepravishta, Ridvan, García-Moreno, M. Isabel, Sánchez-Fernández, Elena M., Ortiz-Mellet, Carmen, Angulo, Jesús, Muñoz-García, Juan C.
Formato: artículo
Estado:Versión publicada
Fecha de publicación:2024
País:España
Recursos:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/356374
Acesso em linha:http://hdl.handle.net/10261/356374
https://api.elsevier.com/content/abstract/scopus_id/85181558542
Access Level:acceso abierto
Palavra-chave:Ligands
Magnetic Resonance Imaging
Magnetic Resonance Spectroscopy
Epitope
Mapping
Protein Binding Proteins
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dc.title.none.fl_str_mv Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
title Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
spellingShingle Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
Rocha, Gabriel
Ligands
Magnetic Resonance Imaging
Magnetic Resonance Spectroscopy
Epitope
Mapping
Protein Binding Proteins
title_short Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
title_full Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
title_fullStr Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
title_full_unstemmed Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
title_sort Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)
dc.creator.none.fl_str_mv Rocha, Gabriel
Ramírez-Cárdenas, Jonathan
Padilla-Pérez, M Carmen
Walpole, Samuel
Nepravishta, Ridvan
García-Moreno, M. Isabel
Sánchez-Fernández, Elena M.
Ortiz-Mellet, Carmen
Angulo, Jesús
Muñoz-García, Juan C.
author Rocha, Gabriel
author_facet Rocha, Gabriel
Ramírez-Cárdenas, Jonathan
Padilla-Pérez, M Carmen
Walpole, Samuel
Nepravishta, Ridvan
García-Moreno, M. Isabel
Sánchez-Fernández, Elena M.
Ortiz-Mellet, Carmen
Angulo, Jesús
Muñoz-García, Juan C.
author_role author
author2 Ramírez-Cárdenas, Jonathan
Padilla-Pérez, M Carmen
Walpole, Samuel
Nepravishta, Ridvan
García-Moreno, M. Isabel
Sánchez-Fernández, Elena M.
Ortiz-Mellet, Carmen
Angulo, Jesús
Muñoz-García, Juan C.
author2_role author
author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Agencia Estatal de Investigación (España)
Ministerio de Ciencia y Tecnología (España)
Norwich Research Park
European Commission
Universidad de Sevilla
Rocha, Gabriel [0009-0004-5673-5943]
Ramírez-Cárdenas, Jonathan [0000-0003-4357-228X]
Padilla-Pérez, M Carmen [0000-0002-1280-3623]
Walpole, Samuel [0000-0002-3360-5204]
Nepravishta, Ridvan [0000-0002-0051-762X]
García-Moreno, M. Isabel [0000-0003-4273-3010]
#NODATA#
Ortiz-Mellet, Carmen [0000-0002-7676-7721]
Angulo, Jesús [0000-0001-7250-5639]
Muñoz-García, Juan C.[0000-0003-2246-3236]
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Ligands
Magnetic Resonance Imaging
Magnetic Resonance Spectroscopy
Epitope
Mapping
Protein Binding Proteins
topic Ligands
Magnetic Resonance Imaging
Magnetic Resonance Spectroscopy
Epitope
Mapping
Protein Binding Proteins
description STD NMR spectroscopy is a powerful ligand-observed NMR tool for screening and characterizing the interactions of small molecules and low molecular weight fragments with a given macromolecule, identifying the main intermolecular contacts in the bound state. It is also a powerful analytical technique for the accurate determination of protein-ligand dissociation constants (KD) of medium-to-weak affinity, of interest in the pharmaceutical industry. However, accurate KD determination and epitope mapping requires a long series of experiments at increasing saturation times to carry out a full analysis using the so-called STD NMR build-up curve approach and apply the "initial slopes approximation". Here, we have developed a new protocol to bypass this important limitation, which allows us to obtain initial slopes by using just two saturation times and, hence, to very quickly determine precise protein-ligand dissociation constants by STD NMR.
publishDate 2024
dc.date.none.fl_str_mv 2024
2024
2024
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Publisher's version
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/356374
https://api.elsevier.com/content/abstract/scopus_id/85181558542
url http://hdl.handle.net/10261/356374
https://api.elsevier.com/content/abstract/scopus_id/85181558542
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
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info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2017-2020/PID2019-109395GB-I00
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2024/PID2022-142879NB-I00
info:eu-repo/grantAgreement/EC/H2020/101064251
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2017-2020/PID2019-105858RB-I00
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PID2022-141034OB-C21
https://pubs.acs.org/doi/full/10.1021/acs.analchem.3c03980
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dc.publisher.none.fl_str_mv ACS Publications
publisher.none.fl_str_mv ACS Publications
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
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spelling Speeding-up the Determination of Protein-Ligand Affinities by STD NMR: The Reduced Data Set STD NMR Approach (rd-STD NMR)Rocha, GabrielRamírez-Cárdenas, JonathanPadilla-Pérez, M CarmenWalpole, SamuelNepravishta, RidvanGarcía-Moreno, M. IsabelSánchez-Fernández, Elena M.Ortiz-Mellet, CarmenAngulo, JesúsMuñoz-García, Juan C.LigandsMagnetic Resonance ImagingMagnetic Resonance SpectroscopyEpitopeMappingProtein Binding ProteinsSTD NMR spectroscopy is a powerful ligand-observed NMR tool for screening and characterizing the interactions of small molecules and low molecular weight fragments with a given macromolecule, identifying the main intermolecular contacts in the bound state. It is also a powerful analytical technique for the accurate determination of protein-ligand dissociation constants (KD) of medium-to-weak affinity, of interest in the pharmaceutical industry. However, accurate KD determination and epitope mapping requires a long series of experiments at increasing saturation times to carry out a full analysis using the so-called STD NMR build-up curve approach and apply the "initial slopes approximation". Here, we have developed a new protocol to bypass this important limitation, which allows us to obtain initial slopes by using just two saturation times and, hence, to very quickly determine precise protein-ligand dissociation constants by STD NMR.J.A., J.R.-C., and G.R. acknowledge the Ministerio de Ciencia e Innovación for funding through the grants PID2019-109395GB-I00 and PID2022-142879NB-I00. J.R.-C. also acknowledges the associated scholarship PRE2020-092754. J.A. and R.N. acknowledge support of BBSRC, grant BB/P010660/1. S.W. was funded by Biotechnology and Biological Sciences Research Council (BBSRC) Norwich Research Park Doctoral Training Grant BB/M011216/1. J.C.M.-G. thanks the European Commission for a HORIZON-TMA-MSCA-PF-EF grant (Sweet2Gel, ID 101064251). This work was supported by grants from the Ministerio de Ciencia e Innovacion, the Agencia Estatal de Investigacion and the European Regional Development Funds: AEI/10.13039/501100011033 (PID2019-105858RB-I00 to C.O.M and PID2022-141034OB-C21 to E.M.S.-F. and C.O.M). M.C.P.-P. is an FPU fellow (Grant number FPU19/04361). E.M.S.-F. thanks the VII Plan Propio de Investigación y Transferencia from the University of Seville (VIIPPIT-2022-V.1, Modality A2). Technical assistance from the research support services of the University of Seville (CITIUS) is also acknowledged.Peer reviewedACS PublicationsAgencia Estatal de Investigación (España)Ministerio de Ciencia y Tecnología (España)Norwich Research ParkEuropean CommissionUniversidad de SevillaRocha, Gabriel [0009-0004-5673-5943]Ramírez-Cárdenas, Jonathan [0000-0003-4357-228X]Padilla-Pérez, M Carmen [0000-0002-1280-3623]Walpole, Samuel [0000-0002-3360-5204]Nepravishta, Ridvan [0000-0002-0051-762X]García-Moreno, M. Isabel [0000-0003-4273-3010]#NODATA#Ortiz-Mellet, Carmen [0000-0002-7676-7721]Angulo, Jesús [0000-0001-7250-5639]Muñoz-García, Juan C.[0000-0003-2246-3236]Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]202420242024info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Publisher's versioninfo:eu-repo/semantics/publishedVersionapplication/pdfhttp://hdl.handle.net/10261/356374https://api.elsevier.com/content/abstract/scopus_id/85181558542reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Inglés#PLACEHOLDER_PARENT_METADATA_VALUE##PLACEHOLDER_PARENT_METADATA_VALUE##PLACEHOLDER_PARENT_METADATA_VALUE##PLACEHOLDER_PARENT_METADATA_VALUE##PLACEHOLDER_PARENT_METADATA_VALUE#info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2017-2020/PID2019-109395GB-I00info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2024/PID2022-142879NB-I00info:eu-repo/grantAgreement/EC/H2020/101064251info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2017-2020/PID2019-105858RB-I00info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PID2022-141034OB-C21https://pubs.acs.org/doi/full/10.1021/acs.analchem.3c03980Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/3563742026-05-22T06:33:51Z
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