Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC)
Autor(a) principal: | |
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Data de Publicação: | 2022 |
Outros Autores: | , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/10362/150714 |
Resumo: | Funding Information: N. Nayak acknowledges the Education, Audiovisual, and Culture Executive Agency (EACEA) for the Ph.D. grant under the Program “Erasmus Mundus Doctorate in Membrane Engineering”—EUDIME (FPA 2011–2014, http://www.eudime.unical.it , accessed on 7 June 2022). The authors would like also to thank POR Calabria FESR-FSE 2014/2020 for the financial support to the SmartMatter project in the ERA.NET Call 2019 and the project Nanomax (N-CHEM): “Nanomax-integrable sensors for pathological biomarkers diagnosis” from the Programma Nazionale della Ricerca, 2011–2013 (Progetto Bandiera). Publisher Copyright: © 2022 by the authors. |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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7160 |
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Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC)APTES functionalizationchromatographyimmunoglobulin G purificationmonolithsprotein Gprotein immobilizationprotein-binding kineticsAnalytical ChemistryChemistry (miscellaneous)Molecular MedicinePharmaceutical ScienceDrug DiscoveryPhysical and Theoretical ChemistryOrganic ChemistryFunding Information: N. Nayak acknowledges the Education, Audiovisual, and Culture Executive Agency (EACEA) for the Ph.D. grant under the Program “Erasmus Mundus Doctorate in Membrane Engineering”—EUDIME (FPA 2011–2014, http://www.eudime.unical.it , accessed on 7 June 2022). The authors would like also to thank POR Calabria FESR-FSE 2014/2020 for the financial support to the SmartMatter project in the ERA.NET Call 2019 and the project Nanomax (N-CHEM): “Nanomax-integrable sensors for pathological biomarkers diagnosis” from the Programma Nazionale della Ricerca, 2011–2013 (Progetto Bandiera). Publisher Copyright: © 2022 by the authors.This work aims at understanding the attachment mechanisms and stability of proteins on a chromatography medium to develop more efficient functionalization methodologies, which can be exploited in affinity chromatography. In particular, the study was focused on the understanding of the attachment mechanisms of bovine serum albumin (BSA), used as a ligand model, and protein G on novel amine-modified alumina monoliths as a stationary phase. Protein G was used to develop a column for antibody purification. The results showed that, at lower protein concentrations (i.e., 0.5 to 1.0 mg·mL−1), protein attachment follows a 1st-order kinetics compatible with the presence of covalent binding between the monolith and the protein. At higher protein concentrations (i.e., up to 10 mg·mL−1), the data preferably fit a 2nd-order kinetics. Such a change reflects a different mechanism in the protein attachment which, at higher concentrations, seems to be governed by physical adsorption resulting in a multilayered protein formation, due to the presence of ligand aggregates. The threshold condition for the prevalence of physical adsorption of BSA was found at a concentration higher than 1.0 mg·mL−1. Based on this result, protein concentrations of 0.7 and 1.0 mg·mL−1 were used for the functionalization of monoliths with protein G, allowing a maximum attachment of 1.43 mg of protein G/g of monolith. This column was then used for IgG binding–elution experiments, which resulted in an antibody attachment of 73.5% and, subsequently, elution of 86%, in acidic conditions. This proved the potential of the amine-functionalized monoliths for application in affinity chromatography.LAQV@REQUIMTEDQ - Departamento de QuímicaRUNNayak, NayanMazzei, RosalindaGiorno, LidiettaCrespo, João G.Portugal, Carla A.M.Poerio, Teresa2023-03-16T22:36:51Z2022-072022-07-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/article14application/pdfhttp://hdl.handle.net/10362/150714eng1420-3049PURE: 56072894https://doi.org/10.3390/molecules27144496info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-03-11T05:33:02Zoai:run.unl.pt:10362/150714Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:54:16.366693Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
title |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
spellingShingle |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) Nayak, Nayan APTES functionalization chromatography immunoglobulin G purification monoliths protein G protein immobilization protein-binding kinetics Analytical Chemistry Chemistry (miscellaneous) Molecular Medicine Pharmaceutical Science Drug Discovery Physical and Theoretical Chemistry Organic Chemistry |
title_short |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
title_full |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
title_fullStr |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
title_full_unstemmed |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
title_sort |
Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC) |
author |
Nayak, Nayan |
author_facet |
Nayak, Nayan Mazzei, Rosalinda Giorno, Lidietta Crespo, João G. Portugal, Carla A.M. Poerio, Teresa |
author_role |
author |
author2 |
Mazzei, Rosalinda Giorno, Lidietta Crespo, João G. Portugal, Carla A.M. Poerio, Teresa |
author2_role |
author author author author author |
dc.contributor.none.fl_str_mv |
LAQV@REQUIMTE DQ - Departamento de Química RUN |
dc.contributor.author.fl_str_mv |
Nayak, Nayan Mazzei, Rosalinda Giorno, Lidietta Crespo, João G. Portugal, Carla A.M. Poerio, Teresa |
dc.subject.por.fl_str_mv |
APTES functionalization chromatography immunoglobulin G purification monoliths protein G protein immobilization protein-binding kinetics Analytical Chemistry Chemistry (miscellaneous) Molecular Medicine Pharmaceutical Science Drug Discovery Physical and Theoretical Chemistry Organic Chemistry |
topic |
APTES functionalization chromatography immunoglobulin G purification monoliths protein G protein immobilization protein-binding kinetics Analytical Chemistry Chemistry (miscellaneous) Molecular Medicine Pharmaceutical Science Drug Discovery Physical and Theoretical Chemistry Organic Chemistry |
description |
Funding Information: N. Nayak acknowledges the Education, Audiovisual, and Culture Executive Agency (EACEA) for the Ph.D. grant under the Program “Erasmus Mundus Doctorate in Membrane Engineering”—EUDIME (FPA 2011–2014, http://www.eudime.unical.it , accessed on 7 June 2022). The authors would like also to thank POR Calabria FESR-FSE 2014/2020 for the financial support to the SmartMatter project in the ERA.NET Call 2019 and the project Nanomax (N-CHEM): “Nanomax-integrable sensors for pathological biomarkers diagnosis” from the Programma Nazionale della Ricerca, 2011–2013 (Progetto Bandiera). Publisher Copyright: © 2022 by the authors. |
publishDate |
2022 |
dc.date.none.fl_str_mv |
2022-07 2022-07-01T00:00:00Z 2023-03-16T22:36:51Z |
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/10362/150714 |
url |
http://hdl.handle.net/10362/150714 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
1420-3049 PURE: 56072894 https://doi.org/10.3390/molecules27144496 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
14 application/pdf |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
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Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
instacron_str |
RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
collection |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository.name.fl_str_mv |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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1799138131920814080 |