Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques

Autores
Braia, Mauricio Javier; Tubio, Gisela; Nerli, Bibiana Beatriz; Loh, Watson; Romanini, Diana
Año de publicación
2012
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Flexible-chain polymers with charge (polyelectrolytes) can interact with globular proteins with a net charge opposite to the charge of the polymers forming insoluble complexes polymer?protein. In this work, the interaction between the basic protein trypsin and the anionic polyelectrolyte Eudragit® L100 was studied by using isothermal calorimetric titrations and differential scanning calorimetry. Turbidimetric assays allowed determining that protein?polymer complex was insoluble at pH below 5 and the trypsin and Eudragit® L100 concentrations required forming the insoluble complex. DSC measurements showed that the Tm and denaturalization heat of trypsin increased in the polymer presence and the complex unfolded according to a two-state model. DH y DS binding parameters obtained by ITC were positives agree with hydrophobic interaction between trypsin and polymer. However, ionic strength of 1.0 M modified the insoluble complex formation. We propose a mechanism of interaction between Eudragit® L100 and trypsin molecules that involves both hydrophobic and electrostatic interactions. Kinetic studies of complex formation showed that the interaction requires less than 1 min achieving the maximum quantity of complex. Finally, a high percentage of active trypsin was precipitated (approximately 76% of the total mass of protein). These findings could be useful in different protocols such as a protein isolation strategy, immobilization or purification of a target protein.
Fil: Braia, Mauricio Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Tubio, Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Nerli, Bibiana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Loh, Watson. Universidade Estadual de Campinas; Brasil
Fil: Romanini, Diana. Universidad Nacional de Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina
Materia
DIFFERENTIAL SCANNING CALORIMETRY
ISOTHERMAL CALORIMETRIC TITRATIONS
PROTEIN-POLYMER INTERACTION
TRYPSIN
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/196175
Acceder
id CONICETDig_d1149370526d134cb2b214ed7f056df4
oai_identifier_str oai:ri.conicet.gov.ar:11336/196175
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniquesBraia, Mauricio JavierTubio, GiselaNerli, Bibiana BeatrizLoh, WatsonRomanini, DianaDIFFERENTIAL SCANNING CALORIMETRYISOTHERMAL CALORIMETRIC TITRATIONSPROTEIN-POLYMER INTERACTIONTRYPSINhttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1Flexible-chain polymers with charge (polyelectrolytes) can interact with globular proteins with a net charge opposite to the charge of the polymers forming insoluble complexes polymer?protein. In this work, the interaction between the basic protein trypsin and the anionic polyelectrolyte Eudragit® L100 was studied by using isothermal calorimetric titrations and differential scanning calorimetry. Turbidimetric assays allowed determining that protein?polymer complex was insoluble at pH below 5 and the trypsin and Eudragit® L100 concentrations required forming the insoluble complex. DSC measurements showed that the Tm and denaturalization heat of trypsin increased in the polymer presence and the complex unfolded according to a two-state model. DH y DS binding parameters obtained by ITC were positives agree with hydrophobic interaction between trypsin and polymer. However, ionic strength of 1.0 M modified the insoluble complex formation. We propose a mechanism of interaction between Eudragit® L100 and trypsin molecules that involves both hydrophobic and electrostatic interactions. Kinetic studies of complex formation showed that the interaction requires less than 1 min achieving the maximum quantity of complex. Finally, a high percentage of active trypsin was precipitated (approximately 76% of the total mass of protein). These findings could be useful in different protocols such as a protein isolation strategy, immobilization or purification of a target protein.Fil: Braia, Mauricio Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; ArgentinaFil: Tubio, Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; ArgentinaFil: Nerli, Bibiana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; ArgentinaFil: Loh, Watson. Universidade Estadual de Campinas; BrasilFil: Romanini, Diana. Universidad Nacional de Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaElsevier Science2012-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/196175Braia, Mauricio Javier; Tubio, Gisela; Nerli, Bibiana Beatriz; Loh, Watson; Romanini, Diana; Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques; Elsevier Science; International Journal of Biological Macromolecules; 50; 1; 1-2012; 180-1860141-8130CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0141813011004065info:eu-repo/semantics/altIdentifier/doi/10.1016/j.ijbiomac.2011.10.016info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T10:02:17Zoai:ri.conicet.gov.ar:11336/196175instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-03 10:02:17.616CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
title Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
spellingShingle Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
Braia, Mauricio Javier
DIFFERENTIAL SCANNING CALORIMETRY
ISOTHERMAL CALORIMETRIC TITRATIONS
PROTEIN-POLYMER INTERACTION
TRYPSIN
title_short Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
title_full Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
title_fullStr Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
title_full_unstemmed Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
title_sort Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques
dc.creator.none.fl_str_mv Braia, Mauricio Javier
Tubio, Gisela
Nerli, Bibiana Beatriz
Loh, Watson
Romanini, Diana
author Braia, Mauricio Javier
author_facet Braia, Mauricio Javier
Tubio, Gisela
Nerli, Bibiana Beatriz
Loh, Watson
Romanini, Diana
author_role author
author2 Tubio, Gisela
Nerli, Bibiana Beatriz
Loh, Watson
Romanini, Diana
author2_role author
author
author
author
dc.subject.none.fl_str_mv DIFFERENTIAL SCANNING CALORIMETRY
ISOTHERMAL CALORIMETRIC TITRATIONS
PROTEIN-POLYMER INTERACTION
TRYPSIN
topic DIFFERENTIAL SCANNING CALORIMETRY
ISOTHERMAL CALORIMETRIC TITRATIONS
PROTEIN-POLYMER INTERACTION
TRYPSIN
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Flexible-chain polymers with charge (polyelectrolytes) can interact with globular proteins with a net charge opposite to the charge of the polymers forming insoluble complexes polymer?protein. In this work, the interaction between the basic protein trypsin and the anionic polyelectrolyte Eudragit® L100 was studied by using isothermal calorimetric titrations and differential scanning calorimetry. Turbidimetric assays allowed determining that protein?polymer complex was insoluble at pH below 5 and the trypsin and Eudragit® L100 concentrations required forming the insoluble complex. DSC measurements showed that the Tm and denaturalization heat of trypsin increased in the polymer presence and the complex unfolded according to a two-state model. DH y DS binding parameters obtained by ITC were positives agree with hydrophobic interaction between trypsin and polymer. However, ionic strength of 1.0 M modified the insoluble complex formation. We propose a mechanism of interaction between Eudragit® L100 and trypsin molecules that involves both hydrophobic and electrostatic interactions. Kinetic studies of complex formation showed that the interaction requires less than 1 min achieving the maximum quantity of complex. Finally, a high percentage of active trypsin was precipitated (approximately 76% of the total mass of protein). These findings could be useful in different protocols such as a protein isolation strategy, immobilization or purification of a target protein.
Fil: Braia, Mauricio Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Tubio, Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Nerli, Bibiana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario; Argentina
Fil: Loh, Watson. Universidade Estadual de Campinas; Brasil
Fil: Romanini, Diana. Universidad Nacional de Rosario; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina
description Flexible-chain polymers with charge (polyelectrolytes) can interact with globular proteins with a net charge opposite to the charge of the polymers forming insoluble complexes polymer?protein. In this work, the interaction between the basic protein trypsin and the anionic polyelectrolyte Eudragit® L100 was studied by using isothermal calorimetric titrations and differential scanning calorimetry. Turbidimetric assays allowed determining that protein?polymer complex was insoluble at pH below 5 and the trypsin and Eudragit® L100 concentrations required forming the insoluble complex. DSC measurements showed that the Tm and denaturalization heat of trypsin increased in the polymer presence and the complex unfolded according to a two-state model. DH y DS binding parameters obtained by ITC were positives agree with hydrophobic interaction between trypsin and polymer. However, ionic strength of 1.0 M modified the insoluble complex formation. We propose a mechanism of interaction between Eudragit® L100 and trypsin molecules that involves both hydrophobic and electrostatic interactions. Kinetic studies of complex formation showed that the interaction requires less than 1 min achieving the maximum quantity of complex. Finally, a high percentage of active trypsin was precipitated (approximately 76% of the total mass of protein). These findings could be useful in different protocols such as a protein isolation strategy, immobilization or purification of a target protein.
publishDate 2012
dc.date.none.fl_str_mv 2012-01
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/196175
Braia, Mauricio Javier; Tubio, Gisela; Nerli, Bibiana Beatriz; Loh, Watson; Romanini, Diana; Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques; Elsevier Science; International Journal of Biological Macromolecules; 50; 1; 1-2012; 180-186
0141-8130
CONICET Digital
CONICET
url http://hdl.handle.net/11336/196175
identifier_str_mv Braia, Mauricio Javier; Tubio, Gisela; Nerli, Bibiana Beatriz; Loh, Watson; Romanini, Diana; Analysis of the interactions between Eudragit® L100 and porcine pancreatic trypsin by calorimetric techniques; Elsevier Science; International Journal of Biological Macromolecules; 50; 1; 1-2012; 180-186
0141-8130
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0141813011004065
info:eu-repo/semantics/altIdentifier/doi/10.1016/j.ijbiomac.2011.10.016
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Science
publisher.none.fl_str_mv Elsevier Science
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.13397

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