Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutage...

Autores
Ballicora, Miguel A.; Erben, Esteban Daniel; Yazaki, Terutaka; Bertolo, Ana L.; Demonte, Ana María Magdalena; Schmidt, Jennifer R.; Aleanzi, Mabel Cristina; Bejar, Clarisa M.; Figueroa, Carlos Maria; Fusari, Corina M.; Iglesias, Alberto Alvaro; Preiss, Jack
Año de publicación
2007
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
ADP-glucose pyrophosphorylase (ADP-Glc PPase) is the enzyme responsible for the regulation of bacterial glycogen synthesis. To perform a structure-function relationship study of the Escherichia coli ADP-Glc PPase enzyme, we studied the effects of pentapeptide insertions at different positions in the enzyme and analyzed the results with a homology model. We randomly inserted 15 bp in a plasmid with the ADP-Glc PPase gene. We obtained 140 modified plasmids with single insertions of which 21 were in the coding region of the enzyme. Fourteen of them generated insertions of five amino acids, whereas the other seven created a stop codon and produced truncations. Correlation of ADP-Glc PPase activity to these modifications validated the enzyme model. Six of the insertions and one truncation produced enzymes with sufficient activity for the E. coli cells to synthesize glycogen and stain in the presence of iodine vapor. These were in regions away from the substrate site, whereas the mutants that did not stain had alterations in critical areas of the protein. The enzyme with a pentapeptide insertion between Leu102 and Pro103 was catalytically competent but insensitive to activation. We postulate this region as critical for the allosteric regulation of the enzyme, participating in the communication between the catalytic and regulatory domains.
Fil: Ballicora, Miguel A.. Loyola University; Estados Unidos
Fil: Erben, Esteban Daniel. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina
Fil: Yazaki, Terutaka. Michigan State University; Estados Unidos
Fil: Bertolo, Ana L.. Michigan State University; Estados Unidos
Fil: Demonte, Ana María Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Schmidt, Jennifer R.. Michigan State University; Estados Unidos
Fil: Aleanzi, Mabel Cristina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Bejar, Clarisa M.. Michigan State University; Estados Unidos
Fil: Figueroa, Carlos Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Fusari, Corina M.. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Preiss, Jack. Michigan State University; Estados Unidos
Materia
Adp-Glucose Pyrophosphorylase
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:11336/79729
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/79729
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesisBallicora, Miguel A.Erben, Esteban DanielYazaki, TerutakaBertolo, Ana L.Demonte, Ana María MagdalenaSchmidt, Jennifer R.Aleanzi, Mabel CristinaBejar, Clarisa M.Figueroa, Carlos MariaFusari, Corina M.Iglesias, Alberto AlvaroPreiss, JackAdp-Glucose Pyrophosphorylasehttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1ADP-glucose pyrophosphorylase (ADP-Glc PPase) is the enzyme responsible for the regulation of bacterial glycogen synthesis. To perform a structure-function relationship study of the Escherichia coli ADP-Glc PPase enzyme, we studied the effects of pentapeptide insertions at different positions in the enzyme and analyzed the results with a homology model. We randomly inserted 15 bp in a plasmid with the ADP-Glc PPase gene. We obtained 140 modified plasmids with single insertions of which 21 were in the coding region of the enzyme. Fourteen of them generated insertions of five amino acids, whereas the other seven created a stop codon and produced truncations. Correlation of ADP-Glc PPase activity to these modifications validated the enzyme model. Six of the insertions and one truncation produced enzymes with sufficient activity for the E. coli cells to synthesize glycogen and stain in the presence of iodine vapor. These were in regions away from the substrate site, whereas the mutants that did not stain had alterations in critical areas of the protein. The enzyme with a pentapeptide insertion between Leu102 and Pro103 was catalytically competent but insensitive to activation. We postulate this region as critical for the allosteric regulation of the enzyme, participating in the communication between the catalytic and regulatory domains.Fil: Ballicora, Miguel A.. Loyola University; Estados UnidosFil: Erben, Esteban Daniel. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Yazaki, Terutaka. Michigan State University; Estados UnidosFil: Bertolo, Ana L.. Michigan State University; Estados UnidosFil: Demonte, Ana María Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; ArgentinaFil: Schmidt, Jennifer R.. Michigan State University; Estados UnidosFil: Aleanzi, Mabel Cristina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; ArgentinaFil: Bejar, Clarisa M.. Michigan State University; Estados UnidosFil: Figueroa, Carlos Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; ArgentinaFil: Fusari, Corina M.. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; ArgentinaFil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; ArgentinaFil: Preiss, Jack. Michigan State University; Estados UnidosAmerican Society for Microbiology2007-07info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/79729Ballicora, Miguel A.; Erben, Esteban Daniel; Yazaki, Terutaka; Bertolo, Ana L.; Demonte, Ana María Magdalena; et al.; Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis; American Society for Microbiology; Journal of Bacteriology; 189; 14; 7-2007; 5325-53330021-9193CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.ncbi.nlm.nih.gov/pubmed/17496097info:eu-repo/semantics/altIdentifier/doi/10.1128/JB.00481-07info:eu-repo/semantics/altIdentifier/url/https://jb.asm.org/content/189/14/5325info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:52:04Zoai:ri.conicet.gov.ar:11336/79729instacron: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 09:52:04.641CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
title Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
spellingShingle Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
Ballicora, Miguel A.
Adp-Glucose Pyrophosphorylase
title_short Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
title_full Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
title_fullStr Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
title_full_unstemmed Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
title_sort Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis
dc.creator.none.fl_str_mv Ballicora, Miguel A.
Erben, Esteban Daniel
Yazaki, Terutaka
Bertolo, Ana L.
Demonte, Ana María Magdalena
Schmidt, Jennifer R.
Aleanzi, Mabel Cristina
Bejar, Clarisa M.
Figueroa, Carlos Maria
Fusari, Corina M.
Iglesias, Alberto Alvaro
Preiss, Jack
author Ballicora, Miguel A.
author_facet Ballicora, Miguel A.
Erben, Esteban Daniel
Yazaki, Terutaka
Bertolo, Ana L.
Demonte, Ana María Magdalena
Schmidt, Jennifer R.
Aleanzi, Mabel Cristina
Bejar, Clarisa M.
Figueroa, Carlos Maria
Fusari, Corina M.
Iglesias, Alberto Alvaro
Preiss, Jack
author_role author
author2 Erben, Esteban Daniel
Yazaki, Terutaka
Bertolo, Ana L.
Demonte, Ana María Magdalena
Schmidt, Jennifer R.
Aleanzi, Mabel Cristina
Bejar, Clarisa M.
Figueroa, Carlos Maria
Fusari, Corina M.
Iglesias, Alberto Alvaro
Preiss, Jack
author2_role author
author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Adp-Glucose Pyrophosphorylase
topic Adp-Glucose Pyrophosphorylase
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv ADP-glucose pyrophosphorylase (ADP-Glc PPase) is the enzyme responsible for the regulation of bacterial glycogen synthesis. To perform a structure-function relationship study of the Escherichia coli ADP-Glc PPase enzyme, we studied the effects of pentapeptide insertions at different positions in the enzyme and analyzed the results with a homology model. We randomly inserted 15 bp in a plasmid with the ADP-Glc PPase gene. We obtained 140 modified plasmids with single insertions of which 21 were in the coding region of the enzyme. Fourteen of them generated insertions of five amino acids, whereas the other seven created a stop codon and produced truncations. Correlation of ADP-Glc PPase activity to these modifications validated the enzyme model. Six of the insertions and one truncation produced enzymes with sufficient activity for the E. coli cells to synthesize glycogen and stain in the presence of iodine vapor. These were in regions away from the substrate site, whereas the mutants that did not stain had alterations in critical areas of the protein. The enzyme with a pentapeptide insertion between Leu102 and Pro103 was catalytically competent but insensitive to activation. We postulate this region as critical for the allosteric regulation of the enzyme, participating in the communication between the catalytic and regulatory domains.
Fil: Ballicora, Miguel A.. Loyola University; Estados Unidos
Fil: Erben, Esteban Daniel. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina
Fil: Yazaki, Terutaka. Michigan State University; Estados Unidos
Fil: Bertolo, Ana L.. Michigan State University; Estados Unidos
Fil: Demonte, Ana María Magdalena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Schmidt, Jennifer R.. Michigan State University; Estados Unidos
Fil: Aleanzi, Mabel Cristina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Bejar, Clarisa M.. Michigan State University; Estados Unidos
Fil: Figueroa, Carlos Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Fusari, Corina M.. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Enzimología Molecular; Argentina
Fil: Preiss, Jack. Michigan State University; Estados Unidos
description ADP-glucose pyrophosphorylase (ADP-Glc PPase) is the enzyme responsible for the regulation of bacterial glycogen synthesis. To perform a structure-function relationship study of the Escherichia coli ADP-Glc PPase enzyme, we studied the effects of pentapeptide insertions at different positions in the enzyme and analyzed the results with a homology model. We randomly inserted 15 bp in a plasmid with the ADP-Glc PPase gene. We obtained 140 modified plasmids with single insertions of which 21 were in the coding region of the enzyme. Fourteen of them generated insertions of five amino acids, whereas the other seven created a stop codon and produced truncations. Correlation of ADP-Glc PPase activity to these modifications validated the enzyme model. Six of the insertions and one truncation produced enzymes with sufficient activity for the E. coli cells to synthesize glycogen and stain in the presence of iodine vapor. These were in regions away from the substrate site, whereas the mutants that did not stain had alterations in critical areas of the protein. The enzyme with a pentapeptide insertion between Leu102 and Pro103 was catalytically competent but insensitive to activation. We postulate this region as critical for the allosteric regulation of the enzyme, participating in the communication between the catalytic and regulatory domains.
publishDate 2007
dc.date.none.fl_str_mv 2007-07
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/11336/79729
Ballicora, Miguel A.; Erben, Esteban Daniel; Yazaki, Terutaka; Bertolo, Ana L.; Demonte, Ana María Magdalena; et al.; Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis; American Society for Microbiology; Journal of Bacteriology; 189; 14; 7-2007; 5325-5333
0021-9193
CONICET Digital
CONICET
url http://hdl.handle.net/11336/79729
identifier_str_mv Ballicora, Miguel A.; Erben, Esteban Daniel; Yazaki, Terutaka; Bertolo, Ana L.; Demonte, Ana María Magdalena; et al.; Identification of regions critically affecting kinetics and allosteric regulation of the Escherichia coli ADP-glucose pyrophosphorylase by modeling and pentapeptide-scanning mutagenesis; American Society for Microbiology; Journal of Bacteriology; 189; 14; 7-2007; 5325-5333
0021-9193
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.ncbi.nlm.nih.gov/pubmed/17496097
info:eu-repo/semantics/altIdentifier/doi/10.1128/JB.00481-07
info:eu-repo/semantics/altIdentifier/url/https://jb.asm.org/content/189/14/5325
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
application/pdf
dc.publisher.none.fl_str_mv American Society for Microbiology
publisher.none.fl_str_mv American Society for Microbiology
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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