Protein Attachment Mechanism for Improved Functionalization of Affinity Monolith Chromatography (AMC)

Detalhes bibliográficos
Autor(a) principal: Nayak, Nayan
Data de Publicação: 2022
Outros Autores: Mazzei, Rosalinda, Giorno, Lidietta, Crespo, João G., Portugal, Carla A.M., Poerio, Teresa
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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spelling 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
instname_str 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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