Novel enzyme-polymer conjugates for biotechnological applications

Autores
Romero, Oscar Eduardo; Rivero, Cintia Wanda; Guisán, José M.; Palomo, José M.
Año de publicación
2013
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
In the present research, a rapid, simple and efficient chemoselective method for the site-directed incorporation of tailor-made polymers into protein to create biocatalysts with excellent properties for pharmaceutical industrial purpose has been performed. First we focused on the protein engineering of the Geobacillus thermocatenulatus lipase 2 (BTL2) to replace the two cysteines (Cys65, Cys296) in the wild type enzyme (BTL-WT) by two serines. Then, by similar mode, a unique cysteine was introduced in the lid area of the protein. For the site-directed polymer incorporation, a set of different tailor-made thiol-ionic-polymers were synthesized and the protein cysteine was previously activated with 2,2-dithiodipyridine (2-PDS) to allow the disulfide exchange. The protected BTL variants were specifically modified with the different polymers in excellent yields, creating a small library of new biocatalysts. Different and important changes in the catalytic properties, possible caused by structural changes in the lid region, were observed. The different modified biocatalysts were tested in the synthesis of intermediates of antiviral and antitumor drugs, like nucleoside analogues and derivatives of phenylglutaric acid. In the hydrolysis of per-acetylated thymidine, the best biocatalyst was the BTL*-193-DextCOOH , where the activity was increased in 3-fold and the regioselectivity was improved, reaching a yield of 92% of 3’-O-acetyl-thymidine. In the case of the asymmetric hydrolysis of dimethyl phenylglutarate, the best result was found with BTL*-193-DextNH2-6000, where the enzyme activity was increased more than 5-fold and the enantiomeric excess was >99%.
Fil: Romero, Oscar Eduardo. Consejo Superior de Investigaciones Cientificas; España
Fil: Rivero, Cintia Wanda. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Biotecnología Sustentable; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Guisán, José M.. Consejo Superior de Investigaciones Cientificas; España
Fil: Palomo, José M.. Consejo Superior de Investigaciones Cientificas; España
Materia
Biocatalyst
Nucleosides
Lipase
Drugs precursor
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/23318
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/23318
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Novel enzyme-polymer conjugates for biotechnological applicationsRomero, Oscar EduardoRivero, Cintia WandaGuisán, José M.Palomo, José M.BiocatalystNucleosidesLipaseDrugs precursorhttps://purl.org/becyt/ford/2.9https://purl.org/becyt/ford/2In the present research, a rapid, simple and efficient chemoselective method for the site-directed incorporation of tailor-made polymers into protein to create biocatalysts with excellent properties for pharmaceutical industrial purpose has been performed. First we focused on the protein engineering of the Geobacillus thermocatenulatus lipase 2 (BTL2) to replace the two cysteines (Cys65, Cys296) in the wild type enzyme (BTL-WT) by two serines. Then, by similar mode, a unique cysteine was introduced in the lid area of the protein. For the site-directed polymer incorporation, a set of different tailor-made thiol-ionic-polymers were synthesized and the protein cysteine was previously activated with 2,2-dithiodipyridine (2-PDS) to allow the disulfide exchange. The protected BTL variants were specifically modified with the different polymers in excellent yields, creating a small library of new biocatalysts. Different and important changes in the catalytic properties, possible caused by structural changes in the lid region, were observed. The different modified biocatalysts were tested in the synthesis of intermediates of antiviral and antitumor drugs, like nucleoside analogues and derivatives of phenylglutaric acid. In the hydrolysis of per-acetylated thymidine, the best biocatalyst was the BTL*-193-DextCOOH , where the activity was increased in 3-fold and the regioselectivity was improved, reaching a yield of 92% of 3’-O-acetyl-thymidine. In the case of the asymmetric hydrolysis of dimethyl phenylglutarate, the best result was found with BTL*-193-DextNH2-6000, where the enzyme activity was increased more than 5-fold and the enantiomeric excess was >99%.Fil: Romero, Oscar Eduardo. Consejo Superior de Investigaciones Cientificas; EspañaFil: Rivero, Cintia Wanda. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Biotecnología Sustentable; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Guisán, José M.. Consejo Superior de Investigaciones Cientificas; EspañaFil: Palomo, José M.. Consejo Superior de Investigaciones Cientificas; EspañaPeerJ2013-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/23318Romero, Oscar Eduardo; Rivero, Cintia Wanda; Guisán, José M.; Palomo, José M.; Novel enzyme-polymer conjugates for biotechnological applications; PeerJ; PeerJ; 1-2013; 1-17; e272167-8359CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.7717/peerj.27info:eu-repo/semantics/altIdentifier/url/https://peerj.com/articles/27/info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:44:00Zoai:ri.conicet.gov.ar:11336/23318instacron: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:34982025-10-22 11:44:00.296CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Novel enzyme-polymer conjugates for biotechnological applications
title Novel enzyme-polymer conjugates for biotechnological applications
spellingShingle Novel enzyme-polymer conjugates for biotechnological applications
Romero, Oscar Eduardo
Biocatalyst
Nucleosides
Lipase
Drugs precursor
title_short Novel enzyme-polymer conjugates for biotechnological applications
title_full Novel enzyme-polymer conjugates for biotechnological applications
title_fullStr Novel enzyme-polymer conjugates for biotechnological applications
title_full_unstemmed Novel enzyme-polymer conjugates for biotechnological applications
title_sort Novel enzyme-polymer conjugates for biotechnological applications
dc.creator.none.fl_str_mv Romero, Oscar Eduardo
Rivero, Cintia Wanda
Guisán, José M.
Palomo, José M.
author Romero, Oscar Eduardo
author_facet Romero, Oscar Eduardo
Rivero, Cintia Wanda
Guisán, José M.
Palomo, José M.
author_role author
author2 Rivero, Cintia Wanda
Guisán, José M.
Palomo, José M.
author2_role author
author
author
dc.subject.none.fl_str_mv Biocatalyst
Nucleosides
Lipase
Drugs precursor
topic Biocatalyst
Nucleosides
Lipase
Drugs precursor
purl_subject.fl_str_mv https://purl.org/becyt/ford/2.9
https://purl.org/becyt/ford/2
dc.description.none.fl_txt_mv In the present research, a rapid, simple and efficient chemoselective method for the site-directed incorporation of tailor-made polymers into protein to create biocatalysts with excellent properties for pharmaceutical industrial purpose has been performed. First we focused on the protein engineering of the Geobacillus thermocatenulatus lipase 2 (BTL2) to replace the two cysteines (Cys65, Cys296) in the wild type enzyme (BTL-WT) by two serines. Then, by similar mode, a unique cysteine was introduced in the lid area of the protein. For the site-directed polymer incorporation, a set of different tailor-made thiol-ionic-polymers were synthesized and the protein cysteine was previously activated with 2,2-dithiodipyridine (2-PDS) to allow the disulfide exchange. The protected BTL variants were specifically modified with the different polymers in excellent yields, creating a small library of new biocatalysts. Different and important changes in the catalytic properties, possible caused by structural changes in the lid region, were observed. The different modified biocatalysts were tested in the synthesis of intermediates of antiviral and antitumor drugs, like nucleoside analogues and derivatives of phenylglutaric acid. In the hydrolysis of per-acetylated thymidine, the best biocatalyst was the BTL*-193-DextCOOH , where the activity was increased in 3-fold and the regioselectivity was improved, reaching a yield of 92% of 3’-O-acetyl-thymidine. In the case of the asymmetric hydrolysis of dimethyl phenylglutarate, the best result was found with BTL*-193-DextNH2-6000, where the enzyme activity was increased more than 5-fold and the enantiomeric excess was >99%.
Fil: Romero, Oscar Eduardo. Consejo Superior de Investigaciones Cientificas; España
Fil: Rivero, Cintia Wanda. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Biotecnología Sustentable; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Guisán, José M.. Consejo Superior de Investigaciones Cientificas; España
Fil: Palomo, José M.. Consejo Superior de Investigaciones Cientificas; España
description In the present research, a rapid, simple and efficient chemoselective method for the site-directed incorporation of tailor-made polymers into protein to create biocatalysts with excellent properties for pharmaceutical industrial purpose has been performed. First we focused on the protein engineering of the Geobacillus thermocatenulatus lipase 2 (BTL2) to replace the two cysteines (Cys65, Cys296) in the wild type enzyme (BTL-WT) by two serines. Then, by similar mode, a unique cysteine was introduced in the lid area of the protein. For the site-directed polymer incorporation, a set of different tailor-made thiol-ionic-polymers were synthesized and the protein cysteine was previously activated with 2,2-dithiodipyridine (2-PDS) to allow the disulfide exchange. The protected BTL variants were specifically modified with the different polymers in excellent yields, creating a small library of new biocatalysts. Different and important changes in the catalytic properties, possible caused by structural changes in the lid region, were observed. The different modified biocatalysts were tested in the synthesis of intermediates of antiviral and antitumor drugs, like nucleoside analogues and derivatives of phenylglutaric acid. In the hydrolysis of per-acetylated thymidine, the best biocatalyst was the BTL*-193-DextCOOH , where the activity was increased in 3-fold and the regioselectivity was improved, reaching a yield of 92% of 3’-O-acetyl-thymidine. In the case of the asymmetric hydrolysis of dimethyl phenylglutarate, the best result was found with BTL*-193-DextNH2-6000, where the enzyme activity was increased more than 5-fold and the enantiomeric excess was >99%.
publishDate 2013
dc.date.none.fl_str_mv 2013-01
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/11336/23318
Romero, Oscar Eduardo; Rivero, Cintia Wanda; Guisán, José M.; Palomo, José M.; Novel enzyme-polymer conjugates for biotechnological applications; PeerJ; PeerJ; 1-2013; 1-17; e27
2167-8359
CONICET Digital
CONICET
url http://hdl.handle.net/11336/23318
identifier_str_mv Romero, Oscar Eduardo; Rivero, Cintia Wanda; Guisán, José M.; Palomo, José M.; Novel enzyme-polymer conjugates for biotechnological applications; PeerJ; PeerJ; 1-2013; 1-17; e27
2167-8359
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.7717/peerj.27
info:eu-repo/semantics/altIdentifier/url/https://peerj.com/articles/27/
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv PeerJ
publisher.none.fl_str_mv PeerJ
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 12.982451

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