A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis

Autores
Paggi, Roberto Alejandro; Gimenez, Maria Ines; de Castro, Rosana Esther; Cesari, Andreina
Año de publicación
2014
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Proteins present in the archaeal cell envelope play key roles in a variety of processes necessary for survival in extreme environments. The haloarchaeon Haloferax volcanii is a good model for membrane proteomic studies because its genome sequence is known, it can be genetically manipulated, and a number of studies at the “omics” level have been performed in this organism. This work reports an easy strategy to improve the resolution of acidic membrane proteins from H. volcanii by 2DE. The method is based on the solubilization, delipidation, and salt removal from membrane proteins. Due to the abundance of the S-layer glycoprotein (SLG) in membrane protein extracts, other proteins from the envelope are consequently underrepresented. Thus, a protocol to reduce the amount of the SLG by EDTA treatment was applied and 11 cm narrow range pH (3.9–5.1) IPG strips were used to fractionate the remaining proteins. Using this method, horizontal streaking was substantially decreased and at least 75 defined spots (20% of the predicted membrane proteome within this pI/Mw range) were reproducibly detected. Two of these spots were identified as thermosome subunit 1 and NADH dehydrogenase from H. volcanii, confirming that proteins from the membrane fraction were enriched. Removal of the SLG from membrane protein extracts can be applied to increase protein load for 2DE as well as for other proteomic methods.
Fil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: Cesari, Andreina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Materia
2d Electrophoresis
Haloarchaea
Acidic Membrane Proteins
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:11336/12877
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/12877
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresisPaggi, Roberto AlejandroGimenez, Maria Inesde Castro, Rosana EstherCesari, Andreina2d ElectrophoresisHaloarchaeaAcidic Membrane Proteinshttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Proteins present in the archaeal cell envelope play key roles in a variety of processes necessary for survival in extreme environments. The haloarchaeon Haloferax volcanii is a good model for membrane proteomic studies because its genome sequence is known, it can be genetically manipulated, and a number of studies at the “omics” level have been performed in this organism. This work reports an easy strategy to improve the resolution of acidic membrane proteins from H. volcanii by 2DE. The method is based on the solubilization, delipidation, and salt removal from membrane proteins. Due to the abundance of the S-layer glycoprotein (SLG) in membrane protein extracts, other proteins from the envelope are consequently underrepresented. Thus, a protocol to reduce the amount of the SLG by EDTA treatment was applied and 11 cm narrow range pH (3.9–5.1) IPG strips were used to fractionate the remaining proteins. Using this method, horizontal streaking was substantially decreased and at least 75 defined spots (20% of the predicted membrane proteome within this pI/Mw range) were reproducibly detected. Two of these spots were identified as thermosome subunit 1 and NADH dehydrogenase from H. volcanii, confirming that proteins from the membrane fraction were enriched. Removal of the SLG from membrane protein extracts can be applied to increase protein load for 2DE as well as for other proteomic methods.Fil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: Cesari, Andreina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaWiley VCH Verlag2014-12info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/12877Paggi, Roberto Alejandro; Gimenez, Maria Ines; de Castro, Rosana Esther; Cesari, Andreina; A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis; Wiley VCH Verlag; Electrophoresis; 35; 24; 12-2014; 3518-35220173-08351522-2683enginfo:eu-repo/semantics/altIdentifier/url/http://onlinelibrary.wiley.com/doi/10.1002/elps.201400407/abstractinfo:eu-repo/semantics/altIdentifier/doi/10.1002/elps.201400407info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-15T14:51:09Zoai:ri.conicet.gov.ar:11336/12877instacron: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:34982025-10-15 14:51:09.377CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
title A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
spellingShingle A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
Paggi, Roberto Alejandro
2d Electrophoresis
Haloarchaea
Acidic Membrane Proteins
title_short A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
title_full A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
title_fullStr A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
title_full_unstemmed A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
title_sort A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
dc.creator.none.fl_str_mv Paggi, Roberto Alejandro
Gimenez, Maria Ines
de Castro, Rosana Esther
Cesari, Andreina
author Paggi, Roberto Alejandro
author_facet Paggi, Roberto Alejandro
Gimenez, Maria Ines
de Castro, Rosana Esther
Cesari, Andreina
author_role author
author2 Gimenez, Maria Ines
de Castro, Rosana Esther
Cesari, Andreina
author2_role author
author
author
dc.subject.none.fl_str_mv 2d Electrophoresis
Haloarchaea
Acidic Membrane Proteins
topic 2d Electrophoresis
Haloarchaea
Acidic Membrane Proteins
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Proteins present in the archaeal cell envelope play key roles in a variety of processes necessary for survival in extreme environments. The haloarchaeon Haloferax volcanii is a good model for membrane proteomic studies because its genome sequence is known, it can be genetically manipulated, and a number of studies at the “omics” level have been performed in this organism. This work reports an easy strategy to improve the resolution of acidic membrane proteins from H. volcanii by 2DE. The method is based on the solubilization, delipidation, and salt removal from membrane proteins. Due to the abundance of the S-layer glycoprotein (SLG) in membrane protein extracts, other proteins from the envelope are consequently underrepresented. Thus, a protocol to reduce the amount of the SLG by EDTA treatment was applied and 11 cm narrow range pH (3.9–5.1) IPG strips were used to fractionate the remaining proteins. Using this method, horizontal streaking was substantially decreased and at least 75 defined spots (20% of the predicted membrane proteome within this pI/Mw range) were reproducibly detected. Two of these spots were identified as thermosome subunit 1 and NADH dehydrogenase from H. volcanii, confirming that proteins from the membrane fraction were enriched. Removal of the SLG from membrane protein extracts can be applied to increase protein load for 2DE as well as for other proteomic methods.
Fil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
Fil: Cesari, Andreina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentina
description Proteins present in the archaeal cell envelope play key roles in a variety of processes necessary for survival in extreme environments. The haloarchaeon Haloferax volcanii is a good model for membrane proteomic studies because its genome sequence is known, it can be genetically manipulated, and a number of studies at the “omics” level have been performed in this organism. This work reports an easy strategy to improve the resolution of acidic membrane proteins from H. volcanii by 2DE. The method is based on the solubilization, delipidation, and salt removal from membrane proteins. Due to the abundance of the S-layer glycoprotein (SLG) in membrane protein extracts, other proteins from the envelope are consequently underrepresented. Thus, a protocol to reduce the amount of the SLG by EDTA treatment was applied and 11 cm narrow range pH (3.9–5.1) IPG strips were used to fractionate the remaining proteins. Using this method, horizontal streaking was substantially decreased and at least 75 defined spots (20% of the predicted membrane proteome within this pI/Mw range) were reproducibly detected. Two of these spots were identified as thermosome subunit 1 and NADH dehydrogenase from H. volcanii, confirming that proteins from the membrane fraction were enriched. Removal of the SLG from membrane protein extracts can be applied to increase protein load for 2DE as well as for other proteomic methods.
publishDate 2014
dc.date.none.fl_str_mv 2014-12
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/11336/12877
Paggi, Roberto Alejandro; Gimenez, Maria Ines; de Castro, Rosana Esther; Cesari, Andreina; A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis; Wiley VCH Verlag; Electrophoresis; 35; 24; 12-2014; 3518-3522
0173-0835
1522-2683
url http://hdl.handle.net/11336/12877
identifier_str_mv Paggi, Roberto Alejandro; Gimenez, Maria Ines; de Castro, Rosana Esther; Cesari, Andreina; A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis; Wiley VCH Verlag; Electrophoresis; 35; 24; 12-2014; 3518-3522
0173-0835
1522-2683
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://onlinelibrary.wiley.com/doi/10.1002/elps.201400407/abstract
info:eu-repo/semantics/altIdentifier/doi/10.1002/elps.201400407
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 Wiley VCH Verlag
publisher.none.fl_str_mv Wiley VCH Verlag
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 13.22299

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