Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.

BACKGROUND In a multicellular organism, the extracellular matrix (ECM) provides a cell-supporting scaffold and helps maintaining the biophysical integrity of tissues and organs. At the same time it plays crucial roles in cellular communication and signalling, with implications in spatial organisatio...

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Autores: Faria Oliveira, Fábio, Carvalho, Joana, Belmiro, Celso LR, Martinez Gomariz, Montserrat, Hernaez, Maria Luisa, Pavão, Mauro, Gil, Concha, Lucas, Cândida, Ferreira, Célia
Tipo de recurso: artículo
Fecha de publicación:2014
País:España
Institución:Universidad Complutense de Madrid (UCM)
Repositorio:Docta Complutense
Idioma:inglés
OAI Identifier:oai:docta.ucm.es:20.500.14352/34911
Acceso en línea:https://hdl.handle.net/20.500.14352/34911
Access Level:acceso abierto
Palabra clave:579
Microbiología (Farmacia)
3302.03 Microbiología Industrial
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spelling Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.Faria Oliveira, FábioCarvalho, JoanaBelmiro, Celso LRMartinez Gomariz, MontserratHernaez, Maria LuisaPavão, MauroGil, ConchaLucas, CândidaFerreira, Célia579Microbiología (Farmacia)3302.03 Microbiología IndustrialBACKGROUND In a multicellular organism, the extracellular matrix (ECM) provides a cell-supporting scaffold and helps maintaining the biophysical integrity of tissues and organs. At the same time it plays crucial roles in cellular communication and signalling, with implications in spatial organisation, motility and differentiation. Similarly, the presence of an ECM-like extracellular polymeric substance is known to support and protect bacterial and fungal multicellular aggregates, such as biofilms or colonies. However, the roles and composition of this microbial ECM are still poorly understood. RESULTS This work presents a protocol to produce S. cerevisiae and C. albicans ECM in an equally highly reproducible manner. Additionally, methodologies for the extraction and fractionation into protein and glycosidic analytical pure fractions were improved. These were subjected to analytical procedures, respectively SDS-PAGE, 2-DE, MALDI-TOF-MS and LC-MS/MS, and DAE and FPLC. Additional chemical methods were also used to test for uronic acids and sulphation. CONCLUSIONS The methodologies hereby presented were equally efficiently applied to extract high amounts of ECM material from S. cerevisiae and C. albicans mats, therefore showing their robustness and reproducibility for yECM molecular and structural characterization. yECM from S. cerevisiae and C. albicans displayed a different proteome and glycoside fractions. S. cerevisiae yECM presented two well-defined polysaccharides with different mass/charge, and C. albicans ECM presented a single different one. The chemical methods further suggested the presence of uronic acids, and chemical modification, possibly through sulphate substitution. All taken, the procedures herein described present the first sensible and concise approach to the molecular and chemical characterisation of the yeast ECM, opening the way to the in-depth study of the microbe multicellular aggregates structure and life-style.BioMed CentralUniversidad Complutense de Madrid20142014-01-0120142014-01-01journal articlehttp://purl.org/coar/resource_type/c_6501info:eu-repo/semantics/articleapplication/pdfhttps://hdl.handle.net/20.500.14352/34911reponame:Docta Complutenseinstname:Universidad Complutense de Madrid (UCM)Inglésengopen accesshttp://purl.org/coar/access_right/c_abf2Atribución 3.0 Españahttps://creativecommons.org/licenses/by/3.0/es/info:eu-repo/semantics/openAccessoai:docta.ucm.es:20.500.14352/349112026-06-02T12:44:21Z
dc.title.none.fl_str_mv Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
title Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
spellingShingle Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
Faria Oliveira, Fábio
579
Microbiología (Farmacia)
3302.03 Microbiología Industrial
title_short Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
title_full Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
title_fullStr Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
title_full_unstemmed Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
title_sort Methodologies to generate, extract, purify and fractionate yeast ECM for analytical use in proteomics and glycomics.
dc.creator.none.fl_str_mv Faria Oliveira, Fábio
Carvalho, Joana
Belmiro, Celso LR
Martinez Gomariz, Montserrat
Hernaez, Maria Luisa
Pavão, Mauro
Gil, Concha
Lucas, Cândida
Ferreira, Célia
author Faria Oliveira, Fábio
author_facet Faria Oliveira, Fábio
Carvalho, Joana
Belmiro, Celso LR
Martinez Gomariz, Montserrat
Hernaez, Maria Luisa
Pavão, Mauro
Gil, Concha
Lucas, Cândida
Ferreira, Célia
author_role author
author2 Carvalho, Joana
Belmiro, Celso LR
Martinez Gomariz, Montserrat
Hernaez, Maria Luisa
Pavão, Mauro
Gil, Concha
Lucas, Cândida
Ferreira, Célia
author2_role author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Universidad Complutense de Madrid
dc.subject.none.fl_str_mv 579
Microbiología (Farmacia)
3302.03 Microbiología Industrial
topic 579
Microbiología (Farmacia)
3302.03 Microbiología Industrial
description BACKGROUND In a multicellular organism, the extracellular matrix (ECM) provides a cell-supporting scaffold and helps maintaining the biophysical integrity of tissues and organs. At the same time it plays crucial roles in cellular communication and signalling, with implications in spatial organisation, motility and differentiation. Similarly, the presence of an ECM-like extracellular polymeric substance is known to support and protect bacterial and fungal multicellular aggregates, such as biofilms or colonies. However, the roles and composition of this microbial ECM are still poorly understood. RESULTS This work presents a protocol to produce S. cerevisiae and C. albicans ECM in an equally highly reproducible manner. Additionally, methodologies for the extraction and fractionation into protein and glycosidic analytical pure fractions were improved. These were subjected to analytical procedures, respectively SDS-PAGE, 2-DE, MALDI-TOF-MS and LC-MS/MS, and DAE and FPLC. Additional chemical methods were also used to test for uronic acids and sulphation. CONCLUSIONS The methodologies hereby presented were equally efficiently applied to extract high amounts of ECM material from S. cerevisiae and C. albicans mats, therefore showing their robustness and reproducibility for yECM molecular and structural characterization. yECM from S. cerevisiae and C. albicans displayed a different proteome and glycoside fractions. S. cerevisiae yECM presented two well-defined polysaccharides with different mass/charge, and C. albicans ECM presented a single different one. The chemical methods further suggested the presence of uronic acids, and chemical modification, possibly through sulphate substitution. All taken, the procedures herein described present the first sensible and concise approach to the molecular and chemical characterisation of the yeast ECM, opening the way to the in-depth study of the microbe multicellular aggregates structure and life-style.
publishDate 2014
dc.date.none.fl_str_mv 2014
2014-01-01
2014
2014-01-01
dc.type.none.fl_str_mv journal article
http://purl.org/coar/resource_type/c_6501
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://hdl.handle.net/20.500.14352/34911
url https://hdl.handle.net/20.500.14352/34911
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
Atribución 3.0 España
https://creativecommons.org/licenses/by/3.0/es/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
Atribución 3.0 España
https://creativecommons.org/licenses/by/3.0/es/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv BioMed Central
publisher.none.fl_str_mv BioMed Central
dc.source.none.fl_str_mv reponame:Docta Complutense
instname:Universidad Complutense de Madrid (UCM)
instname_str Universidad Complutense de Madrid (UCM)
reponame_str Docta Complutense
collection Docta Complutense
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,198674

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