High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii

Autores
Chesini, Mariana; Plaza Cazón, Josefina del Carmen; Baruque, Diego Jorge; Vita, Carolina Elena; Cavalitto, Sebastian Fernando; Ghiringhelli, Pablo Daniel; Rojas, Natalia Lorena
Año de publicación
2018
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Exoinulinases—enzymes extensively studied in recent decades because of their industrial applications—need to be produced in suitable quantities in order to meet production demands. We describe here the production of an acid-stable recombinant inulinase from Aspergillus kawachii in the Pichia pastoris system and the recombinant enzyme's biochemical characteristics and potential application to industrial processes. After an appropriate cloning strategy, this genetically engineered inulinase was successfully overproduced in fed-batch fermentations, reaching up to 840 U/ml after a 72-h cultivation. The protein, purified to homogeneity by chromatographic techniques, was obtained at a 42% yield. The following biochemical characteristics were determined: the enzyme had an optimal pH of 3, was stable for at least 3 h at 55 °C, and was inhibited in catalytic activity almost completely by Hg+2. The respective Km and Vmax for the recombinant inulinase with inulin as substrate were 1.35 mM and 2673 μmol/min/mg. The recombinant enzyme is an exoinulinase but also possesses synthetic activity (i. e., fructosyl transferase). The high level of production of this recombinant plus its relevant biochemical properties would argue that the process presented here is a possible recourse for industrial applications in carbohydrate processing.
Fil: Chesini, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Plaza Cazón, Josefina del Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Baruque, Diego Jorge. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Vita, Carolina Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Cavalitto, Sebastian Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Ghiringhelli, Pablo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina
Fil: Rojas, Natalia Lorena. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Materia
EXOINULINASE
ASPERGILLUS KAWACHII
PICHIA PASTORIS
FRUCTOSYLTRANSFERASE
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:11336/94888
Acceder
id CONICETDig_b2e9d75fba5285b745bd7a49704d60cd
oai_identifier_str oai:ri.conicet.gov.ar:11336/94888
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling High level production of a recombinant acid stable exoinulinase from Aspergillus kawachiiChesini, MarianaPlaza Cazón, Josefina del CarmenBaruque, Diego JorgeVita, Carolina ElenaCavalitto, Sebastian FernandoGhiringhelli, Pablo DanielRojas, Natalia LorenaEXOINULINASEASPERGILLUS KAWACHIIPICHIA PASTORISFRUCTOSYLTRANSFERASEhttps://purl.org/becyt/ford/2.9https://purl.org/becyt/ford/2https://purl.org/becyt/ford/2.9https://purl.org/becyt/ford/2Exoinulinases—enzymes extensively studied in recent decades because of their industrial applications—need to be produced in suitable quantities in order to meet production demands. We describe here the production of an acid-stable recombinant inulinase from Aspergillus kawachii in the Pichia pastoris system and the recombinant enzyme's biochemical characteristics and potential application to industrial processes. After an appropriate cloning strategy, this genetically engineered inulinase was successfully overproduced in fed-batch fermentations, reaching up to 840 U/ml after a 72-h cultivation. The protein, purified to homogeneity by chromatographic techniques, was obtained at a 42% yield. The following biochemical characteristics were determined: the enzyme had an optimal pH of 3, was stable for at least 3 h at 55 °C, and was inhibited in catalytic activity almost completely by Hg+2. The respective Km and Vmax for the recombinant inulinase with inulin as substrate were 1.35 mM and 2673 μmol/min/mg. The recombinant enzyme is an exoinulinase but also possesses synthetic activity (i. e., fructosyl transferase). The high level of production of this recombinant plus its relevant biochemical properties would argue that the process presented here is a possible recourse for industrial applications in carbohydrate processing.Fil: Chesini, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; ArgentinaFil: Plaza Cazón, Josefina del Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Baruque, Diego Jorge. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Vita, Carolina Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; ArgentinaFil: Cavalitto, Sebastian Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; ArgentinaFil: Ghiringhelli, Pablo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; ArgentinaFil: Rojas, Natalia Lorena. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaAcademic Press Inc Elsevier Science2018-07info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/94888Chesini, Mariana; Plaza Cazón, Josefina del Carmen; Baruque, Diego Jorge; Vita, Carolina Elena; Cavalitto, Sebastian Fernando; et al.; High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii; Academic Press Inc Elsevier Science; Protein Expression and Purification; 147; 7-2018; 29-371046-5928CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S1046592817306927info:eu-repo/semantics/altIdentifier/doi/10.1016/j.pep.2018.02.007info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:42:33Zoai:ri.conicet.gov.ar:11336/94888instacron: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:34982025-09-29 09:42:34.146CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
title High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
spellingShingle High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
Chesini, Mariana
EXOINULINASE
ASPERGILLUS KAWACHII
PICHIA PASTORIS
FRUCTOSYLTRANSFERASE
title_short High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
title_full High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
title_fullStr High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
title_full_unstemmed High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
title_sort High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii
dc.creator.none.fl_str_mv Chesini, Mariana
Plaza Cazón, Josefina del Carmen
Baruque, Diego Jorge
Vita, Carolina Elena
Cavalitto, Sebastian Fernando
Ghiringhelli, Pablo Daniel
Rojas, Natalia Lorena
author Chesini, Mariana
author_facet Chesini, Mariana
Plaza Cazón, Josefina del Carmen
Baruque, Diego Jorge
Vita, Carolina Elena
Cavalitto, Sebastian Fernando
Ghiringhelli, Pablo Daniel
Rojas, Natalia Lorena
author_role author
author2 Plaza Cazón, Josefina del Carmen
Baruque, Diego Jorge
Vita, Carolina Elena
Cavalitto, Sebastian Fernando
Ghiringhelli, Pablo Daniel
Rojas, Natalia Lorena
author2_role author
author
author
author
author
author
dc.subject.none.fl_str_mv EXOINULINASE
ASPERGILLUS KAWACHII
PICHIA PASTORIS
FRUCTOSYLTRANSFERASE
topic EXOINULINASE
ASPERGILLUS KAWACHII
PICHIA PASTORIS
FRUCTOSYLTRANSFERASE
purl_subject.fl_str_mv https://purl.org/becyt/ford/2.9
https://purl.org/becyt/ford/2
https://purl.org/becyt/ford/2.9
https://purl.org/becyt/ford/2
dc.description.none.fl_txt_mv Exoinulinases—enzymes extensively studied in recent decades because of their industrial applications—need to be produced in suitable quantities in order to meet production demands. We describe here the production of an acid-stable recombinant inulinase from Aspergillus kawachii in the Pichia pastoris system and the recombinant enzyme's biochemical characteristics and potential application to industrial processes. After an appropriate cloning strategy, this genetically engineered inulinase was successfully overproduced in fed-batch fermentations, reaching up to 840 U/ml after a 72-h cultivation. The protein, purified to homogeneity by chromatographic techniques, was obtained at a 42% yield. The following biochemical characteristics were determined: the enzyme had an optimal pH of 3, was stable for at least 3 h at 55 °C, and was inhibited in catalytic activity almost completely by Hg+2. The respective Km and Vmax for the recombinant inulinase with inulin as substrate were 1.35 mM and 2673 μmol/min/mg. The recombinant enzyme is an exoinulinase but also possesses synthetic activity (i. e., fructosyl transferase). The high level of production of this recombinant plus its relevant biochemical properties would argue that the process presented here is a possible recourse for industrial applications in carbohydrate processing.
Fil: Chesini, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Plaza Cazón, Josefina del Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Baruque, Diego Jorge. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Vita, Carolina Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Cavalitto, Sebastian Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina
Fil: Ghiringhelli, Pablo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina
Fil: Rojas, Natalia Lorena. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
description Exoinulinases—enzymes extensively studied in recent decades because of their industrial applications—need to be produced in suitable quantities in order to meet production demands. We describe here the production of an acid-stable recombinant inulinase from Aspergillus kawachii in the Pichia pastoris system and the recombinant enzyme's biochemical characteristics and potential application to industrial processes. After an appropriate cloning strategy, this genetically engineered inulinase was successfully overproduced in fed-batch fermentations, reaching up to 840 U/ml after a 72-h cultivation. The protein, purified to homogeneity by chromatographic techniques, was obtained at a 42% yield. The following biochemical characteristics were determined: the enzyme had an optimal pH of 3, was stable for at least 3 h at 55 °C, and was inhibited in catalytic activity almost completely by Hg+2. The respective Km and Vmax for the recombinant inulinase with inulin as substrate were 1.35 mM and 2673 μmol/min/mg. The recombinant enzyme is an exoinulinase but also possesses synthetic activity (i. e., fructosyl transferase). The high level of production of this recombinant plus its relevant biochemical properties would argue that the process presented here is a possible recourse for industrial applications in carbohydrate processing.
publishDate 2018
dc.date.none.fl_str_mv 2018-07
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/11336/94888
Chesini, Mariana; Plaza Cazón, Josefina del Carmen; Baruque, Diego Jorge; Vita, Carolina Elena; Cavalitto, Sebastian Fernando; et al.; High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii; Academic Press Inc Elsevier Science; Protein Expression and Purification; 147; 7-2018; 29-37
1046-5928
CONICET Digital
CONICET
url http://hdl.handle.net/11336/94888
identifier_str_mv Chesini, Mariana; Plaza Cazón, Josefina del Carmen; Baruque, Diego Jorge; Vita, Carolina Elena; Cavalitto, Sebastian Fernando; et al.; High level production of a recombinant acid stable exoinulinase from Aspergillus kawachii; Academic Press Inc Elsevier Science; Protein Expression and Purification; 147; 7-2018; 29-37
1046-5928
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/S1046592817306927
info:eu-repo/semantics/altIdentifier/doi/10.1016/j.pep.2018.02.007
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
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Academic Press Inc Elsevier Science
publisher.none.fl_str_mv Academic Press Inc 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 13.070432

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