Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)

Autores
Martín, Mariana; Wayllace, Nahuel Zenón; Valdez, Hugo Alberto; Gomez Casati, Diego Fabian; Busi, María Victoria
Año de publicación
2013
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Glycogen and starch, the major storage carbohydrate in most living organisms, result mainly from the action of starch or glycogen synthases (SS or GS, respectively, EC 2.4.1.21). SSIII from Arabidopsis thaliana is an SS isoform with a particular modular organization: the C-terminal highly conserved glycosyltransferase domain is preceded by a unique specific region (SSIII-SD) which contains three in tandem starch binding domains (SBDs, named D1, D2 and D3) characteristic of polysaccharide degrading enzymes. N-terminal SBDs have a probed regulatory role in SSIII activity, showing starch binding ability and modulating the catalytic properties of the enzyme. On the other hand, GS from Agrobacterium tumefaciens has a simple primary structure organization, characterized only by the highly conserved glycosyltransferase domain and lacking SBDs. To further investigate the functional role of A. thaliana SSIII-SD, three chimeric proteins were constructed combining the SBDs from A. thaliana with the GS from A. tumefaciens. Recombinant proteins were expressed in and purified to homogeneity from Escherichia coli cells in order to be kinetically characterized. Furthermore, we tested the ability to restore in vivo glycogen biosynthesis in transformed E. coli glgA− cells, deficient in GS. Results show that the D3-GS chimeric enzyme showed increased capacity of glycogen synthesis in vivo with minor changes in its kinetics parameters compared to GS.
Fil: Martín, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Wayllace, Nahuel Zenón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Valdez, Hugo Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina
Fil: Gomez Casati, Diego Fabian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Busi, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Materia
Glycogen
Enzyme
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:11336/7834
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/7834
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)Martín, MarianaWayllace, Nahuel ZenónValdez, Hugo AlbertoGomez Casati, Diego FabianBusi, María VictoriaGlycogenEnzymehttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Glycogen and starch, the major storage carbohydrate in most living organisms, result mainly from the action of starch or glycogen synthases (SS or GS, respectively, EC 2.4.1.21). SSIII from Arabidopsis thaliana is an SS isoform with a particular modular organization: the C-terminal highly conserved glycosyltransferase domain is preceded by a unique specific region (SSIII-SD) which contains three in tandem starch binding domains (SBDs, named D1, D2 and D3) characteristic of polysaccharide degrading enzymes. N-terminal SBDs have a probed regulatory role in SSIII activity, showing starch binding ability and modulating the catalytic properties of the enzyme. On the other hand, GS from Agrobacterium tumefaciens has a simple primary structure organization, characterized only by the highly conserved glycosyltransferase domain and lacking SBDs. To further investigate the functional role of A. thaliana SSIII-SD, three chimeric proteins were constructed combining the SBDs from A. thaliana with the GS from A. tumefaciens. Recombinant proteins were expressed in and purified to homogeneity from Escherichia coli cells in order to be kinetically characterized. Furthermore, we tested the ability to restore in vivo glycogen biosynthesis in transformed E. coli glgA− cells, deficient in GS. Results show that the D3-GS chimeric enzyme showed increased capacity of glycogen synthesis in vivo with minor changes in its kinetics parameters compared to GS.Fil: Martín, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); ArgentinaFil: Wayllace, Nahuel Zenón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); ArgentinaFil: Valdez, Hugo Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); ArgentinaFil: Gomez Casati, Diego Fabian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); ArgentinaFil: Busi, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); ArgentinaElsevier Masson2013-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/7834Martín, Mariana; Wayllace, Nahuel Zenón; Valdez, Hugo Alberto; Gomez Casati, Diego Fabian; Busi, María Victoria; Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs); Elsevier Masson; Biochimie; 95; 10; 6-2013; 1865-18700300-9084enginfo:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0300908413001892info:eu-repo/semantics/altIdentifier/doi/10.1016/j.biochi.2013.06.009info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-22T11:44:37Zoai:ri.conicet.gov.ar:11336/7834instacron: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 11:44:38.073CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
title Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
spellingShingle Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
Martín, Mariana
Glycogen
Enzyme
title_short Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
title_full Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
title_fullStr Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
title_full_unstemmed Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
title_sort Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs)
dc.creator.none.fl_str_mv Martín, Mariana
Wayllace, Nahuel Zenón
Valdez, Hugo Alberto
Gomez Casati, Diego Fabian
Busi, María Victoria
author Martín, Mariana
author_facet Martín, Mariana
Wayllace, Nahuel Zenón
Valdez, Hugo Alberto
Gomez Casati, Diego Fabian
Busi, María Victoria
author_role author
author2 Wayllace, Nahuel Zenón
Valdez, Hugo Alberto
Gomez Casati, Diego Fabian
Busi, María Victoria
author2_role author
author
author
author
dc.subject.none.fl_str_mv Glycogen
Enzyme
topic Glycogen
Enzyme
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Glycogen and starch, the major storage carbohydrate in most living organisms, result mainly from the action of starch or glycogen synthases (SS or GS, respectively, EC 2.4.1.21). SSIII from Arabidopsis thaliana is an SS isoform with a particular modular organization: the C-terminal highly conserved glycosyltransferase domain is preceded by a unique specific region (SSIII-SD) which contains three in tandem starch binding domains (SBDs, named D1, D2 and D3) characteristic of polysaccharide degrading enzymes. N-terminal SBDs have a probed regulatory role in SSIII activity, showing starch binding ability and modulating the catalytic properties of the enzyme. On the other hand, GS from Agrobacterium tumefaciens has a simple primary structure organization, characterized only by the highly conserved glycosyltransferase domain and lacking SBDs. To further investigate the functional role of A. thaliana SSIII-SD, three chimeric proteins were constructed combining the SBDs from A. thaliana with the GS from A. tumefaciens. Recombinant proteins were expressed in and purified to homogeneity from Escherichia coli cells in order to be kinetically characterized. Furthermore, we tested the ability to restore in vivo glycogen biosynthesis in transformed E. coli glgA− cells, deficient in GS. Results show that the D3-GS chimeric enzyme showed increased capacity of glycogen synthesis in vivo with minor changes in its kinetics parameters compared to GS.
Fil: Martín, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Wayllace, Nahuel Zenón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Valdez, Hugo Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina
Fil: Gomez Casati, Diego Fabian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
Fil: Busi, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Centro de Estudios Fotosintéticos y Bioquímicos (i); Argentina
description Glycogen and starch, the major storage carbohydrate in most living organisms, result mainly from the action of starch or glycogen synthases (SS or GS, respectively, EC 2.4.1.21). SSIII from Arabidopsis thaliana is an SS isoform with a particular modular organization: the C-terminal highly conserved glycosyltransferase domain is preceded by a unique specific region (SSIII-SD) which contains three in tandem starch binding domains (SBDs, named D1, D2 and D3) characteristic of polysaccharide degrading enzymes. N-terminal SBDs have a probed regulatory role in SSIII activity, showing starch binding ability and modulating the catalytic properties of the enzyme. On the other hand, GS from Agrobacterium tumefaciens has a simple primary structure organization, characterized only by the highly conserved glycosyltransferase domain and lacking SBDs. To further investigate the functional role of A. thaliana SSIII-SD, three chimeric proteins were constructed combining the SBDs from A. thaliana with the GS from A. tumefaciens. Recombinant proteins were expressed in and purified to homogeneity from Escherichia coli cells in order to be kinetically characterized. Furthermore, we tested the ability to restore in vivo glycogen biosynthesis in transformed E. coli glgA− cells, deficient in GS. Results show that the D3-GS chimeric enzyme showed increased capacity of glycogen synthesis in vivo with minor changes in its kinetics parameters compared to GS.
publishDate 2013
dc.date.none.fl_str_mv 2013-06
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/11336/7834
Martín, Mariana; Wayllace, Nahuel Zenón; Valdez, Hugo Alberto; Gomez Casati, Diego Fabian; Busi, María Victoria; Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs); Elsevier Masson; Biochimie; 95; 10; 6-2013; 1865-1870
0300-9084
url http://hdl.handle.net/11336/7834
identifier_str_mv Martín, Mariana; Wayllace, Nahuel Zenón; Valdez, Hugo Alberto; Gomez Casati, Diego Fabian; Busi, María Victoria; Improving the glycosyltransferase activity of Agrobacterium tumefaciens glycogen synthase by fusion of N-terminal starch binding domains (SBDs); Elsevier Masson; Biochimie; 95; 10; 6-2013; 1865-1870
0300-9084
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0300908413001892
info:eu-repo/semantics/altIdentifier/doi/10.1016/j.biochi.2013.06.009
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
dc.publisher.none.fl_str_mv Elsevier Masson
publisher.none.fl_str_mv Elsevier Masson
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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