Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulat...

Autores
Ballicora, Miguel A.; Sesma, Juliana; Iglesias, Alberto Alvaro; Preiss, Jack
Año de publicación
2002
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
ADPglucose pyrophosphorylase catalyzes the regulatory step in the pathway for bacterial glycogen synthesis. The enzymes from different organisms exhibit distinctive regulatory properties related to the main carbon metabolic pathway. Escherichia coli ADPglucose pyrophosphorylase is mainly activated by fructose 1,6-bisphosphate (FBP), whereas the Agrobacterium tumefaciens enzyme is activated by fructose 6-phosphate (F6P) and pyruvate. Little is known about the regions determining the specificity for the allosteric regulator. To study the function of different domains, two chimeric enzymes were constructed. "AE" contains the N-terminus (271 amino acids) of the A. tumefaciens ADPglucose pyrophosphorylase and the C-terminus (153 residues) of the E. coli enzyme, and "EA", the inverse construction. Expression of the recombinant wild-type and chimeric enzymes was performed using derivatives of the pET24a plasmid. Characterization of the purified chimeric enzymes showed that the C-terminus of the E. coli enzyme is relevant for the selectivity by FBP. However, this region seems to be less important for the specificity by F6P in the A. tumefaciens enzyme. The chimeric enzyme AE is activated by both FBP and F6P, neither of which affect EA. Pyruvate activates EA with higher apparent affinity than AE, suggesting that the C-terminus of the A. tumefaciens enzyme plays a role in the binding of this effector. The allosteric inhibitor site is apparently disrupted, as a marked desensitization toward AMP was observed in the chimeric enzymes.
Fil: Ballicora, Miguel A.. Michigan State University; Estados Unidos
Fil: Sesma, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; 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
Fil: Preiss, Jack. Michigan State University; Estados Unidos
Materia
ADPglucose pyrophosphorylase
glycogen synthesis
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:11336/85682
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/85682
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulatorsBallicora, Miguel A.Sesma, JulianaIglesias, Alberto AlvaroPreiss, JackADPglucose pyrophosphorylaseglycogen synthesishttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1ADPglucose pyrophosphorylase catalyzes the regulatory step in the pathway for bacterial glycogen synthesis. The enzymes from different organisms exhibit distinctive regulatory properties related to the main carbon metabolic pathway. Escherichia coli ADPglucose pyrophosphorylase is mainly activated by fructose 1,6-bisphosphate (FBP), whereas the Agrobacterium tumefaciens enzyme is activated by fructose 6-phosphate (F6P) and pyruvate. Little is known about the regions determining the specificity for the allosteric regulator. To study the function of different domains, two chimeric enzymes were constructed. "AE" contains the N-terminus (271 amino acids) of the A. tumefaciens ADPglucose pyrophosphorylase and the C-terminus (153 residues) of the E. coli enzyme, and "EA", the inverse construction. Expression of the recombinant wild-type and chimeric enzymes was performed using derivatives of the pET24a plasmid. Characterization of the purified chimeric enzymes showed that the C-terminus of the E. coli enzyme is relevant for the selectivity by FBP. However, this region seems to be less important for the specificity by F6P in the A. tumefaciens enzyme. The chimeric enzyme AE is activated by both FBP and F6P, neither of which affect EA. Pyruvate activates EA with higher apparent affinity than AE, suggesting that the C-terminus of the A. tumefaciens enzyme plays a role in the binding of this effector. The allosteric inhibitor site is apparently disrupted, as a marked desensitization toward AMP was observed in the chimeric enzymes.Fil: Ballicora, Miguel A.. Michigan State University; Estados UnidosFil: Sesma, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; 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; ArgentinaFil: Preiss, Jack. Michigan State University; Estados UnidosAmerican Chemical Society2002-07info: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/11336/85682Ballicora, Miguel A.; Sesma, Juliana; Iglesias, Alberto Alvaro; Preiss, Jack; Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators; American Chemical Society; Biochemistry; 41; 30; 7-2002; 9431-94370006-2960CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://www.ncbi.nlm.nih.gov/pubmed/?term=Characterization+of+chimeric+ADP-glucose+pyrophosphorylases+of+Escherichia+coli+and+Agrobacterium+tumefaciens.+Importance+of+the+C-terminus+on+the+slectivity+for+allosteric+regulatorsinfo:eu-repo/semantics/altIdentifier/doi/10.1021/bi025793binfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:52:36Zoai:ri.conicet.gov.ar:11336/85682instacron: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 11:52:36.99CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
title Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
spellingShingle Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
Ballicora, Miguel A.
ADPglucose pyrophosphorylase
glycogen synthesis
title_short Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
title_full Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
title_fullStr Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
title_full_unstemmed Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
title_sort Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators
dc.creator.none.fl_str_mv Ballicora, Miguel A.
Sesma, Juliana
Iglesias, Alberto Alvaro
Preiss, Jack
author Ballicora, Miguel A.
author_facet Ballicora, Miguel A.
Sesma, Juliana
Iglesias, Alberto Alvaro
Preiss, Jack
author_role author
author2 Sesma, Juliana
Iglesias, Alberto Alvaro
Preiss, Jack
author2_role author
author
author
dc.subject.none.fl_str_mv ADPglucose pyrophosphorylase
glycogen synthesis
topic ADPglucose pyrophosphorylase
glycogen synthesis
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv ADPglucose pyrophosphorylase catalyzes the regulatory step in the pathway for bacterial glycogen synthesis. The enzymes from different organisms exhibit distinctive regulatory properties related to the main carbon metabolic pathway. Escherichia coli ADPglucose pyrophosphorylase is mainly activated by fructose 1,6-bisphosphate (FBP), whereas the Agrobacterium tumefaciens enzyme is activated by fructose 6-phosphate (F6P) and pyruvate. Little is known about the regions determining the specificity for the allosteric regulator. To study the function of different domains, two chimeric enzymes were constructed. "AE" contains the N-terminus (271 amino acids) of the A. tumefaciens ADPglucose pyrophosphorylase and the C-terminus (153 residues) of the E. coli enzyme, and "EA", the inverse construction. Expression of the recombinant wild-type and chimeric enzymes was performed using derivatives of the pET24a plasmid. Characterization of the purified chimeric enzymes showed that the C-terminus of the E. coli enzyme is relevant for the selectivity by FBP. However, this region seems to be less important for the specificity by F6P in the A. tumefaciens enzyme. The chimeric enzyme AE is activated by both FBP and F6P, neither of which affect EA. Pyruvate activates EA with higher apparent affinity than AE, suggesting that the C-terminus of the A. tumefaciens enzyme plays a role in the binding of this effector. The allosteric inhibitor site is apparently disrupted, as a marked desensitization toward AMP was observed in the chimeric enzymes.
Fil: Ballicora, Miguel A.. Michigan State University; Estados Unidos
Fil: Sesma, Juliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; 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
Fil: Preiss, Jack. Michigan State University; Estados Unidos
description ADPglucose pyrophosphorylase catalyzes the regulatory step in the pathway for bacterial glycogen synthesis. The enzymes from different organisms exhibit distinctive regulatory properties related to the main carbon metabolic pathway. Escherichia coli ADPglucose pyrophosphorylase is mainly activated by fructose 1,6-bisphosphate (FBP), whereas the Agrobacterium tumefaciens enzyme is activated by fructose 6-phosphate (F6P) and pyruvate. Little is known about the regions determining the specificity for the allosteric regulator. To study the function of different domains, two chimeric enzymes were constructed. "AE" contains the N-terminus (271 amino acids) of the A. tumefaciens ADPglucose pyrophosphorylase and the C-terminus (153 residues) of the E. coli enzyme, and "EA", the inverse construction. Expression of the recombinant wild-type and chimeric enzymes was performed using derivatives of the pET24a plasmid. Characterization of the purified chimeric enzymes showed that the C-terminus of the E. coli enzyme is relevant for the selectivity by FBP. However, this region seems to be less important for the specificity by F6P in the A. tumefaciens enzyme. The chimeric enzyme AE is activated by both FBP and F6P, neither of which affect EA. Pyruvate activates EA with higher apparent affinity than AE, suggesting that the C-terminus of the A. tumefaciens enzyme plays a role in the binding of this effector. The allosteric inhibitor site is apparently disrupted, as a marked desensitization toward AMP was observed in the chimeric enzymes.
publishDate 2002
dc.date.none.fl_str_mv 2002-07
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/11336/85682
Ballicora, Miguel A.; Sesma, Juliana; Iglesias, Alberto Alvaro; Preiss, Jack; Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators; American Chemical Society; Biochemistry; 41; 30; 7-2002; 9431-9437
0006-2960
CONICET Digital
CONICET
url http://hdl.handle.net/11336/85682
identifier_str_mv Ballicora, Miguel A.; Sesma, Juliana; Iglesias, Alberto Alvaro; Preiss, Jack; Characterization of chimeric ADPglucose pyrophosphorylases of Escherichia coli and Agrobacterium tumefaciens. Importance of the C-terminus on the selectivity for allosteric regulators; American Chemical Society; Biochemistry; 41; 30; 7-2002; 9431-9437
0006-2960
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://www.ncbi.nlm.nih.gov/pubmed/?term=Characterization+of+chimeric+ADP-glucose+pyrophosphorylases+of+Escherichia+coli+and+Agrobacterium+tumefaciens.+Importance+of+the+C-terminus+on+the+slectivity+for+allosteric+regulators
info:eu-repo/semantics/altIdentifier/doi/10.1021/bi025793b
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 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 12.982451

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