Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis

Autores
Muchut, Robertino José; Piattoni, Claudia Vanesa; Margarit, Ezequiel; Tripodi, Karina Eva Josefina; Podesta, Florencio Esteban; Iglesias, Alberto Alvaro
Año de publicación
2019
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
This work reports the molecular cloning and heterologous expression of the genes coding for α and β subunits of pyrophosphate-dependent phosphofructokinase (PPi-PFK) from orange. When expressed individually, both recombinant subunits were produced as highly purified monomeric proteins able to phosphorylate fructose-6-phosphate at the expenses of PPi (specific activity of 0.075 and 0.017 units. mg −1 for α and β subunits, respectively). On the other hand, co-expression rendered a α 3 β 3 hexamer with specific activity three orders of magnitude higher than the single subunits. All the conformations of the enzyme were characterized with respect to its kinetic properties and sensitivity to the regulator fructose-2,6-bisphosphate. A thorough review of current knowledge on the matter indicates that this is the first report of the recombinant production of active plant PPi-PFK and the characterization of its different conformations. This is a main contribution for future studies focused to better understand the enzyme properties and how it accomplishes its relevant role in plant metabolism.
Fil: Muchut, Robertino José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
Fil: Piattoni, Claudia Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
Fil: Margarit, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Tripodi, Karina Eva Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Podesta, Florencio Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
Materia
CITRUS SINENSIS
FRUCTOSE-2,6-BISPHOSPHATE
HETERO-HEXAMER
ORANGE
PPI-DEPENDENT PHOSPHOFRUCTOKINASE
RECOMBINANT PROTEIN
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/148468
Acceder
id CONICETDig_10283f9d36872c6c9add6d81f36000ad
oai_identifier_str oai:ri.conicet.gov.ar:11336/148468
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensisMuchut, Robertino JoséPiattoni, Claudia VanesaMargarit, EzequielTripodi, Karina Eva JosefinaPodesta, Florencio EstebanIglesias, Alberto AlvaroCITRUS SINENSISFRUCTOSE-2,6-BISPHOSPHATEHETERO-HEXAMERORANGEPPI-DEPENDENT PHOSPHOFRUCTOKINASERECOMBINANT PROTEINhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1This work reports the molecular cloning and heterologous expression of the genes coding for α and β subunits of pyrophosphate-dependent phosphofructokinase (PPi-PFK) from orange. When expressed individually, both recombinant subunits were produced as highly purified monomeric proteins able to phosphorylate fructose-6-phosphate at the expenses of PPi (specific activity of 0.075 and 0.017 units. mg −1 for α and β subunits, respectively). On the other hand, co-expression rendered a α 3 β 3 hexamer with specific activity three orders of magnitude higher than the single subunits. All the conformations of the enzyme were characterized with respect to its kinetic properties and sensitivity to the regulator fructose-2,6-bisphosphate. A thorough review of current knowledge on the matter indicates that this is the first report of the recombinant production of active plant PPi-PFK and the characterization of its different conformations. This is a main contribution for future studies focused to better understand the enzyme properties and how it accomplishes its relevant role in plant metabolism.Fil: Muchut, Robertino José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Piattoni, Claudia Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Margarit, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; ArgentinaFil: Tripodi, Karina Eva Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; ArgentinaFil: Podesta, Florencio Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; ArgentinaFil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaElsevier Ireland2019-03info: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/11336/148468Muchut, Robertino José; Piattoni, Claudia Vanesa; Margarit, Ezequiel; Tripodi, Karina Eva Josefina; Podesta, Florencio Esteban; et al.; Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis; Elsevier Ireland; Plant Science; 280; 3-2019; 348-3540168-9452CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/abs/pii/S0168945218311075info:eu-repo/semantics/altIdentifier/doi/10.1016/j.plantsci.2018.12.012info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-nd/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:13:31Zoai:ri.conicet.gov.ar:11336/148468instacron: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 11:13:31.264CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
title Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
spellingShingle Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
Muchut, Robertino José
CITRUS SINENSIS
FRUCTOSE-2,6-BISPHOSPHATE
HETERO-HEXAMER
ORANGE
PPI-DEPENDENT PHOSPHOFRUCTOKINASE
RECOMBINANT PROTEIN
title_short Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
title_full Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
title_fullStr Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
title_full_unstemmed Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
title_sort Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis
dc.creator.none.fl_str_mv Muchut, Robertino José
Piattoni, Claudia Vanesa
Margarit, Ezequiel
Tripodi, Karina Eva Josefina
Podesta, Florencio Esteban
Iglesias, Alberto Alvaro
author Muchut, Robertino José
author_facet Muchut, Robertino José
Piattoni, Claudia Vanesa
Margarit, Ezequiel
Tripodi, Karina Eva Josefina
Podesta, Florencio Esteban
Iglesias, Alberto Alvaro
author_role author
author2 Piattoni, Claudia Vanesa
Margarit, Ezequiel
Tripodi, Karina Eva Josefina
Podesta, Florencio Esteban
Iglesias, Alberto Alvaro
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv CITRUS SINENSIS
FRUCTOSE-2,6-BISPHOSPHATE
HETERO-HEXAMER
ORANGE
PPI-DEPENDENT PHOSPHOFRUCTOKINASE
RECOMBINANT PROTEIN
topic CITRUS SINENSIS
FRUCTOSE-2,6-BISPHOSPHATE
HETERO-HEXAMER
ORANGE
PPI-DEPENDENT PHOSPHOFRUCTOKINASE
RECOMBINANT PROTEIN
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv This work reports the molecular cloning and heterologous expression of the genes coding for α and β subunits of pyrophosphate-dependent phosphofructokinase (PPi-PFK) from orange. When expressed individually, both recombinant subunits were produced as highly purified monomeric proteins able to phosphorylate fructose-6-phosphate at the expenses of PPi (specific activity of 0.075 and 0.017 units. mg −1 for α and β subunits, respectively). On the other hand, co-expression rendered a α 3 β 3 hexamer with specific activity three orders of magnitude higher than the single subunits. All the conformations of the enzyme were characterized with respect to its kinetic properties and sensitivity to the regulator fructose-2,6-bisphosphate. A thorough review of current knowledge on the matter indicates that this is the first report of the recombinant production of active plant PPi-PFK and the characterization of its different conformations. This is a main contribution for future studies focused to better understand the enzyme properties and how it accomplishes its relevant role in plant metabolism.
Fil: Muchut, Robertino José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
Fil: Piattoni, Claudia Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
Fil: Margarit, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Tripodi, Karina Eva Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Podesta, Florencio Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina
Fil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina
description This work reports the molecular cloning and heterologous expression of the genes coding for α and β subunits of pyrophosphate-dependent phosphofructokinase (PPi-PFK) from orange. When expressed individually, both recombinant subunits were produced as highly purified monomeric proteins able to phosphorylate fructose-6-phosphate at the expenses of PPi (specific activity of 0.075 and 0.017 units. mg −1 for α and β subunits, respectively). On the other hand, co-expression rendered a α 3 β 3 hexamer with specific activity three orders of magnitude higher than the single subunits. All the conformations of the enzyme were characterized with respect to its kinetic properties and sensitivity to the regulator fructose-2,6-bisphosphate. A thorough review of current knowledge on the matter indicates that this is the first report of the recombinant production of active plant PPi-PFK and the characterization of its different conformations. This is a main contribution for future studies focused to better understand the enzyme properties and how it accomplishes its relevant role in plant metabolism.
publishDate 2019
dc.date.none.fl_str_mv 2019-03
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/11336/148468
Muchut, Robertino José; Piattoni, Claudia Vanesa; Margarit, Ezequiel; Tripodi, Karina Eva Josefina; Podesta, Florencio Esteban; et al.; Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis; Elsevier Ireland; Plant Science; 280; 3-2019; 348-354
0168-9452
CONICET Digital
CONICET
url http://hdl.handle.net/11336/148468
identifier_str_mv Muchut, Robertino José; Piattoni, Claudia Vanesa; Margarit, Ezequiel; Tripodi, Karina Eva Josefina; Podesta, Florencio Esteban; et al.; Heterologous expression and kinetic characterization of the α β and αβ blend of the PPi-dependent phosphofructokinase from Citrus sinensis; Elsevier Ireland; Plant Science; 280; 3-2019; 348-354
0168-9452
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/abs/pii/S0168945218311075
info:eu-repo/semantics/altIdentifier/doi/10.1016/j.plantsci.2018.12.012
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-nd/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 Elsevier Ireland
publisher.none.fl_str_mv Elsevier Ireland
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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