Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity...

Autores
Moyano, Fernando; Setien, Evangelina; Silber, Juana J.; Correa, Nestor Mariano
Año de publicación
2013
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The reverse micelle (RM) media are very good as nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different water-dimethyl sulfoxide (DMSO) mixtures encapsulated in 1,4-bis-2-ethylhexylsulfosuccinate (AOT)/n-heptane RMs on the enzymatic hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide (Bz-Try-pNA) by α- chymotrypsin (α-CT). The reaction was first studied in homogeneous media at different DMSO-water mixture compositions and in DMSO-water/AOT/n-heptane RMs. The hydrolysis rates of Bz-Try-pNA catalyzed by α-CT were determined by UV−vis spectroscopy. The reaction follows the Michaelis-Menten mechanism and the kinetic parameters: kcat, KM, and kcat/KM were evaluated under different conditions. In this homogeneous media, DMSO plays an important role in the solubilization process of the peptide which is almost insoluble in water, but it has a tremendous impact on the inactivation of α-CT. It is shown that the enzyme dissolved in a 20% molar ratio of the DMSO-water mixture does not present enzymatic activity. Dynamic light scattering has been used to assess the formation of DMSO-water/AOT/heptane RMs at different DMSO compositions. The results also show that there is preferential solvation of the AOT RM interface by water molecules. To test the use of these RMs as nanoreactors, the kinetic parameters for the enzymatic reaction in these systems have been evaluated. The parameters were determined at fixed WS {WS = ([water] + [DMSO])/[AOT] = 20} at different DMSO-water compositions. The results show that the Michaelis−Menten mechanism is valid for α-CT in all the RM systems studied and that the reaction takes place at the RM interface. Surprisingly, it was observed that the enzyme encapsulated by the RMs show catalytic effects with similar kcat/KM values at any DMSO composition investigated, which evidence that DMSO molecules are localized far from the RM interface.
Fil: Moyano, Fernando. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Setien, Evangelina. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina
Fil: Silber, Juana J.. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina
Fil: Correa, Nestor Mariano. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Materia
Reverse Micelles
Α-Chymotrypsin
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:11336/23371
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/23371
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activityMoyano, FernandoSetien, EvangelinaSilber, Juana J.Correa, Nestor MarianoReverse MicellesΑ-Chymotrypsinhttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1The reverse micelle (RM) media are very good as nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different water-dimethyl sulfoxide (DMSO) mixtures encapsulated in 1,4-bis-2-ethylhexylsulfosuccinate (AOT)/n-heptane RMs on the enzymatic hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide (Bz-Try-pNA) by α- chymotrypsin (α-CT). The reaction was first studied in homogeneous media at different DMSO-water mixture compositions and in DMSO-water/AOT/n-heptane RMs. The hydrolysis rates of Bz-Try-pNA catalyzed by α-CT were determined by UV−vis spectroscopy. The reaction follows the Michaelis-Menten mechanism and the kinetic parameters: kcat, KM, and kcat/KM were evaluated under different conditions. In this homogeneous media, DMSO plays an important role in the solubilization process of the peptide which is almost insoluble in water, but it has a tremendous impact on the inactivation of α-CT. It is shown that the enzyme dissolved in a 20% molar ratio of the DMSO-water mixture does not present enzymatic activity. Dynamic light scattering has been used to assess the formation of DMSO-water/AOT/heptane RMs at different DMSO compositions. The results also show that there is preferential solvation of the AOT RM interface by water molecules. To test the use of these RMs as nanoreactors, the kinetic parameters for the enzymatic reaction in these systems have been evaluated. The parameters were determined at fixed WS {WS = ([water] + [DMSO])/[AOT] = 20} at different DMSO-water compositions. The results show that the Michaelis−Menten mechanism is valid for α-CT in all the RM systems studied and that the reaction takes place at the RM interface. Surprisingly, it was observed that the enzyme encapsulated by the RMs show catalytic effects with similar kcat/KM values at any DMSO composition investigated, which evidence that DMSO molecules are localized far from the RM interface.Fil: Moyano, Fernando. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Setien, Evangelina. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; ArgentinaFil: Silber, Juana J.. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; ArgentinaFil: Correa, Nestor Mariano. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaAmerican Chemical Society2013-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/23371Moyano, Fernando; Setien, Evangelina; Silber, Juana J.; Correa, Nestor Mariano; Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity; American Chemical Society; Langmuir; 29; 26; 6-2013; 8245-82540743-7463CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1021/la401103qinfo:eu-repo/semantics/altIdentifier/url/http://pubs.acs.org/doi/abs/10.1021/la401103qinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-10T13:24:33Zoai:ri.conicet.gov.ar:11336/23371instacron: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:34982025-09-10 13:24:33.331CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
title Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
spellingShingle Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
Moyano, Fernando
Reverse Micelles
Α-Chymotrypsin
title_short Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
title_full Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
title_fullStr Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
title_full_unstemmed Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
title_sort Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity
dc.creator.none.fl_str_mv Moyano, Fernando
Setien, Evangelina
Silber, Juana J.
Correa, Nestor Mariano
author Moyano, Fernando
author_facet Moyano, Fernando
Setien, Evangelina
Silber, Juana J.
Correa, Nestor Mariano
author_role author
author2 Setien, Evangelina
Silber, Juana J.
Correa, Nestor Mariano
author2_role author
author
author
dc.subject.none.fl_str_mv Reverse Micelles
Α-Chymotrypsin
topic Reverse Micelles
Α-Chymotrypsin
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv The reverse micelle (RM) media are very good as nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different water-dimethyl sulfoxide (DMSO) mixtures encapsulated in 1,4-bis-2-ethylhexylsulfosuccinate (AOT)/n-heptane RMs on the enzymatic hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide (Bz-Try-pNA) by α- chymotrypsin (α-CT). The reaction was first studied in homogeneous media at different DMSO-water mixture compositions and in DMSO-water/AOT/n-heptane RMs. The hydrolysis rates of Bz-Try-pNA catalyzed by α-CT were determined by UV−vis spectroscopy. The reaction follows the Michaelis-Menten mechanism and the kinetic parameters: kcat, KM, and kcat/KM were evaluated under different conditions. In this homogeneous media, DMSO plays an important role in the solubilization process of the peptide which is almost insoluble in water, but it has a tremendous impact on the inactivation of α-CT. It is shown that the enzyme dissolved in a 20% molar ratio of the DMSO-water mixture does not present enzymatic activity. Dynamic light scattering has been used to assess the formation of DMSO-water/AOT/heptane RMs at different DMSO compositions. The results also show that there is preferential solvation of the AOT RM interface by water molecules. To test the use of these RMs as nanoreactors, the kinetic parameters for the enzymatic reaction in these systems have been evaluated. The parameters were determined at fixed WS {WS = ([water] + [DMSO])/[AOT] = 20} at different DMSO-water compositions. The results show that the Michaelis−Menten mechanism is valid for α-CT in all the RM systems studied and that the reaction takes place at the RM interface. Surprisingly, it was observed that the enzyme encapsulated by the RMs show catalytic effects with similar kcat/KM values at any DMSO composition investigated, which evidence that DMSO molecules are localized far from the RM interface.
Fil: Moyano, Fernando. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Setien, Evangelina. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina
Fil: Silber, Juana J.. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina
Fil: Correa, Nestor Mariano. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
description The reverse micelle (RM) media are very good as nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different water-dimethyl sulfoxide (DMSO) mixtures encapsulated in 1,4-bis-2-ethylhexylsulfosuccinate (AOT)/n-heptane RMs on the enzymatic hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide (Bz-Try-pNA) by α- chymotrypsin (α-CT). The reaction was first studied in homogeneous media at different DMSO-water mixture compositions and in DMSO-water/AOT/n-heptane RMs. The hydrolysis rates of Bz-Try-pNA catalyzed by α-CT were determined by UV−vis spectroscopy. The reaction follows the Michaelis-Menten mechanism and the kinetic parameters: kcat, KM, and kcat/KM were evaluated under different conditions. In this homogeneous media, DMSO plays an important role in the solubilization process of the peptide which is almost insoluble in water, but it has a tremendous impact on the inactivation of α-CT. It is shown that the enzyme dissolved in a 20% molar ratio of the DMSO-water mixture does not present enzymatic activity. Dynamic light scattering has been used to assess the formation of DMSO-water/AOT/heptane RMs at different DMSO compositions. The results also show that there is preferential solvation of the AOT RM interface by water molecules. To test the use of these RMs as nanoreactors, the kinetic parameters for the enzymatic reaction in these systems have been evaluated. The parameters were determined at fixed WS {WS = ([water] + [DMSO])/[AOT] = 20} at different DMSO-water compositions. The results show that the Michaelis−Menten mechanism is valid for α-CT in all the RM systems studied and that the reaction takes place at the RM interface. Surprisingly, it was observed that the enzyme encapsulated by the RMs show catalytic effects with similar kcat/KM values at any DMSO composition investigated, which evidence that DMSO molecules are localized far from the RM interface.
publishDate 2013
dc.date.none.fl_str_mv 2013-06
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/11336/23371
Moyano, Fernando; Setien, Evangelina; Silber, Juana J.; Correa, Nestor Mariano; Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity; American Chemical Society; Langmuir; 29; 26; 6-2013; 8245-8254
0743-7463
CONICET Digital
CONICET
url http://hdl.handle.net/11336/23371
identifier_str_mv Moyano, Fernando; Setien, Evangelina; Silber, Juana J.; Correa, Nestor Mariano; Enzymatic Hydrolysis of N‑Benzoyl‑L‑Tyrosine p‑Nitroanilide by α‑Chymotrypsin in DMSO-Water/AOT/n‑Heptane Reverse Micelles: a unique interfacial effect on the enzymatic activity; American Chemical Society; Langmuir; 29; 26; 6-2013; 8245-8254
0743-7463
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1021/la401103q
info:eu-repo/semantics/altIdentifier/url/http://pubs.acs.org/doi/abs/10.1021/la401103q
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
dc.publisher.none.fl_str_mv American Chemical Society
publisher.none.fl_str_mv American Chemical Society
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 12.48226

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