Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions

Bibliographic Details
Main Author: Gizardin-Fredon, Hugo
Publication Date: 2024
Other Authors: Santo, Paulo E., Chagot, Marie Eve, Charpentier, Bruno, Bandeiras, Tiago M., Manival, Xavier, Hernandez-Alba, Oscar, Cianférani, Sarah
Format: Article
Language: eng
Source: Repositórios Científicos de Acesso Aberto de Portugal (RCAAP)
Download full: http://hdl.handle.net/10362/172803
Summary: Funding Information: This work was supported by the CNRS, the University of Strasbourg, Lorraine University, the \u201CAgence National de la Recherche\u201D (ANR-16-CE11-0032-04 snoRNP assembly; ANR-23-CE44-0035 NAProt-XLMS) and the French Proteomics Infrastructure (ProFI; ANR-10-INBS-08-03). H.G.F. acknowledges the French Ministry for Education and Research for funding of his PhD. This work was funded by Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia/Minist\u00E9rio da Ci\u00EAncia, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020 and UIDP/04462/2020) and the Associate Laboratory LS4FUTURE (LA/P/0087/2020). Publisher Copyright: © The Author(s) 2024.
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spelling Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactionsChemistry(all)Biochemistry, Genetics and Molecular Biology(all)Physics and Astronomy(all)Funding Information: This work was supported by the CNRS, the University of Strasbourg, Lorraine University, the \u201CAgence National de la Recherche\u201D (ANR-16-CE11-0032-04 snoRNP assembly; ANR-23-CE44-0035 NAProt-XLMS) and the French Proteomics Infrastructure (ProFI; ANR-10-INBS-08-03). H.G.F. acknowledges the French Ministry for Education and Research for funding of his PhD. This work was funded by Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia/Minist\u00E9rio da Ci\u00EAncia, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020 and UIDP/04462/2020) and the Associate Laboratory LS4FUTURE (LA/P/0087/2020). Publisher Copyright: © The Author(s) 2024.Chemical cross-linking reactions (XL) are an important strategy for studying protein-protein interactions (PPIs), including low abundant sub-complexes, in structural biology. However, choosing XL reagents and conditions is laborious and mostly limited to analysis of protein assemblies that can be resolved using SDS-PAGE. To overcome these limitations, we develop here a denaturing mass photometry (dMP) method for fast, reliable and user-friendly optimization and monitoring of chemical XL reactions. The dMP is a robust 2-step protocol that ensures 95% of irreversible denaturation within only 5 min. We show that dMP provides accurate mass identification across a broad mass range (30 kDa–5 MDa) along with direct label-free relative quantification of all coexisting XL species (sub-complexes and aggregates). We compare dMP with SDS-PAGE and observe that, unlike the benchmark, dMP is time-efficient (3 min/triplicate), requires significantly less material (20–100×) and affords single molecule sensitivity. To illustrate its utility for routine structural biology applications, we show that dMP affords screening of 20 XL conditions in 1 h, accurately identifying and quantifying all coexisting species. Taken together, we anticipate that dMP will have an impact on ability to structurally characterize more PPIs and macromolecular assemblies, expected final complexes but also sub-complexes that form en route.Instituto de Tecnologia Química e Biológica António Xavier (ITQB)RUNGizardin-Fredon, HugoSanto, Paulo E.Chagot, Marie EveCharpentier, BrunoBandeiras, Tiago M.Manival, XavierHernandez-Alba, OscarCianférani, Sarah2024-10-01T22:22:41Z2024-04-252024-04-25T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10362/172803eng2041-1723PURE: 90361378https://doi.org/10.1038/s41467-024-47732-4info:eu-repo/semantics/openAccessreponame:Repositórios Científicos de Acesso Aberto de Portugal (RCAAP)instname:FCCN, serviços digitais da FCT – Fundação para a Ciência e a Tecnologiainstacron:RCAAP2024-10-07T01:39:53Zoai:run.unl.pt:10362/172803Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireinfo@rcaap.ptopendoar:https://opendoar.ac.uk/repository/71602025-05-28T18:55:22.857575Repositórios Científicos de Acesso Aberto de Portugal (RCAAP) - FCCN, serviços digitais da FCT – Fundação para a Ciência e a Tecnologiafalse
dc.title.none.fl_str_mv Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
title Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
spellingShingle Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
Gizardin-Fredon, Hugo
Chemistry(all)
Biochemistry, Genetics and Molecular Biology(all)
Physics and Astronomy(all)
title_short Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
title_full Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
title_fullStr Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
title_full_unstemmed Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
title_sort Denaturing mass photometry for rapid optimization of chemical protein-protein cross-linking reactions
author Gizardin-Fredon, Hugo
author_facet Gizardin-Fredon, Hugo
Santo, Paulo E.
Chagot, Marie Eve
Charpentier, Bruno
Bandeiras, Tiago M.
Manival, Xavier
Hernandez-Alba, Oscar
Cianférani, Sarah
author_role author
author2 Santo, Paulo E.
Chagot, Marie Eve
Charpentier, Bruno
Bandeiras, Tiago M.
Manival, Xavier
Hernandez-Alba, Oscar
Cianférani, Sarah
author2_role author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Instituto de Tecnologia Química e Biológica António Xavier (ITQB)
RUN
dc.contributor.author.fl_str_mv Gizardin-Fredon, Hugo
Santo, Paulo E.
Chagot, Marie Eve
Charpentier, Bruno
Bandeiras, Tiago M.
Manival, Xavier
Hernandez-Alba, Oscar
Cianférani, Sarah
dc.subject.por.fl_str_mv Chemistry(all)
Biochemistry, Genetics and Molecular Biology(all)
Physics and Astronomy(all)
topic Chemistry(all)
Biochemistry, Genetics and Molecular Biology(all)
Physics and Astronomy(all)
description Funding Information: This work was supported by the CNRS, the University of Strasbourg, Lorraine University, the \u201CAgence National de la Recherche\u201D (ANR-16-CE11-0032-04 snoRNP assembly; ANR-23-CE44-0035 NAProt-XLMS) and the French Proteomics Infrastructure (ProFI; ANR-10-INBS-08-03). H.G.F. acknowledges the French Ministry for Education and Research for funding of his PhD. This work was funded by Funda\u00E7\u00E3o para a Ci\u00EAncia e Tecnologia/Minist\u00E9rio da Ci\u00EAncia, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020 and UIDP/04462/2020) and the Associate Laboratory LS4FUTURE (LA/P/0087/2020). Publisher Copyright: © The Author(s) 2024.
publishDate 2024
dc.date.none.fl_str_mv 2024-10-01T22:22:41Z
2024-04-25
2024-04-25T00:00:00Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10362/172803
url http://hdl.handle.net/10362/172803
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv 2041-1723
PURE: 90361378
https://doi.org/10.1038/s41467-024-47732-4
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Repositórios Científicos de Acesso Aberto de Portugal (RCAAP)
instname:FCCN, serviços digitais da FCT – Fundação para a Ciência e a Tecnologia
instacron:RCAAP
instname_str FCCN, serviços digitais da FCT – Fundação para a Ciência e a Tecnologia
instacron_str RCAAP
institution RCAAP
reponame_str Repositórios Científicos de Acesso Aberto de Portugal (RCAAP)
collection Repositórios Científicos de Acesso Aberto de Portugal (RCAAP)
repository.name.fl_str_mv Repositórios Científicos de Acesso Aberto de Portugal (RCAAP) - FCCN, serviços digitais da FCT – Fundação para a Ciência e a Tecnologia
repository.mail.fl_str_mv info@rcaap.pt
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