Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes

Autores
Mangialavori, Irene Cecilia; Ferreira Gomes, Mariela Soledad; Pignataro, María Florencia; Strehler, Emanuel E.; Rossi, Juan Pablo Francisco
Año de publicación
2010
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The purpose of this work was to obtain information about conformational changes of the plasma membrane Ca2+-pump (PMCA) in the membrane region upon interaction with Ca2+, calmodulin (CaM) and acidic phospholipids. To this end, we have quantified labeling of PMCA with the photoactivatable phosphatidylcholine analog [125I]TID-PC/16, measuring the shift of conformation E2 to the auto-inhibited conformation E1I and to the activated E1A state, titrating the effect of Ca2+ under different conditions. Using a similar approach, we also determined the CaM-PMCA dissociation constant. The results indicate that the PMCA possesses a high affinity site for Ca2+ regardless of the presence or absence of activators. Modulation of pump activity is exerted through the C-terminal domain, which induces an apparent auto-inhibited conformation for Ca2+ transport but does not modify the affinity for Ca2+ at the transmembrane domain. The C-terminal domain is affected by CaM and CaM-like treatments driving the auto-inhibited conformation E1I to the activated E1A conformation and thus modulating the transport of Ca2+. This is reflected in the different apparent constants for Ca2+ in the absence of CaM (calculated by Ca2+-ATPase activity) that sharply contrast with the lack of variation of the affinity for the Ca2+ site at equilibrium. This is the first time that equilibrium constants for the dissociation of Ca2+ and CaM ligands from PMCA complexes are measured through the change of transmembrane conformations of the pump. The data further suggest that the transmembrane domain of the PMCA undergoes major rearrangements resulting in altered lipid accessibility upon Ca2+ binding and activation.
Fil: Mangialavori, Irene Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Ferreira Gomes, Mariela Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Pignataro, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Strehler, Emanuel E.. Mayo Clinic College of Medicine; Estados Unidos
Fil: Rossi, Juan Pablo Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Materia
Plasma Membrane Calcium Pump
Dissociation Constants
Conformational Changes
Transmembrane Domain
Equilibrium Constants
Membrane Proteins
Photoactivatable Phospholipids
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:11336/18238
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/18238
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changesMangialavori, Irene CeciliaFerreira Gomes, Mariela SoledadPignataro, María FlorenciaStrehler, Emanuel E.Rossi, Juan Pablo FranciscoPlasma Membrane Calcium PumpDissociation ConstantsConformational ChangesTransmembrane DomainEquilibrium ConstantsMembrane ProteinsPhotoactivatable Phospholipidshttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1The purpose of this work was to obtain information about conformational changes of the plasma membrane Ca2+-pump (PMCA) in the membrane region upon interaction with Ca2+, calmodulin (CaM) and acidic phospholipids. To this end, we have quantified labeling of PMCA with the photoactivatable phosphatidylcholine analog [125I]TID-PC/16, measuring the shift of conformation E2 to the auto-inhibited conformation E1I and to the activated E1A state, titrating the effect of Ca2+ under different conditions. Using a similar approach, we also determined the CaM-PMCA dissociation constant. The results indicate that the PMCA possesses a high affinity site for Ca2+ regardless of the presence or absence of activators. Modulation of pump activity is exerted through the C-terminal domain, which induces an apparent auto-inhibited conformation for Ca2+ transport but does not modify the affinity for Ca2+ at the transmembrane domain. The C-terminal domain is affected by CaM and CaM-like treatments driving the auto-inhibited conformation E1I to the activated E1A conformation and thus modulating the transport of Ca2+. This is reflected in the different apparent constants for Ca2+ in the absence of CaM (calculated by Ca2+-ATPase activity) that sharply contrast with the lack of variation of the affinity for the Ca2+ site at equilibrium. This is the first time that equilibrium constants for the dissociation of Ca2+ and CaM ligands from PMCA complexes are measured through the change of transmembrane conformations of the pump. The data further suggest that the transmembrane domain of the PMCA undergoes major rearrangements resulting in altered lipid accessibility upon Ca2+ binding and activation.Fil: Mangialavori, Irene Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; ArgentinaFil: Ferreira Gomes, Mariela Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; ArgentinaFil: Pignataro, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; ArgentinaFil: Strehler, Emanuel E.. Mayo Clinic College of Medicine; Estados UnidosFil: Rossi, Juan Pablo Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; ArgentinaAmerican Society for Biochemistry and Molecular Biology2010-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/18238Mangialavori, Irene Cecilia; Ferreira Gomes, Mariela Soledad; Pignataro, María Florencia; Strehler, Emanuel E.; Rossi, Juan Pablo Francisco; Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes; American Society for Biochemistry and Molecular Biology; Journal of Biological Chemistry; 285; 1; 1-2010; 123-1300021-9258CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1074/jbc.M109.076679info:eu-repo/semantics/altIdentifier/url/http://www.jbc.org/content/285/1/123.longinfo:eu-repo/semantics/altIdentifier/url/https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2804156/info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T09:45:34Zoai:ri.conicet.gov.ar:11336/18238instacron: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:34982025-09-03 09:45:34.683CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
title Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
spellingShingle Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
Mangialavori, Irene Cecilia
Plasma Membrane Calcium Pump
Dissociation Constants
Conformational Changes
Transmembrane Domain
Equilibrium Constants
Membrane Proteins
Photoactivatable Phospholipids
title_short Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
title_full Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
title_fullStr Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
title_full_unstemmed Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
title_sort Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes
dc.creator.none.fl_str_mv Mangialavori, Irene Cecilia
Ferreira Gomes, Mariela Soledad
Pignataro, María Florencia
Strehler, Emanuel E.
Rossi, Juan Pablo Francisco
author Mangialavori, Irene Cecilia
author_facet Mangialavori, Irene Cecilia
Ferreira Gomes, Mariela Soledad
Pignataro, María Florencia
Strehler, Emanuel E.
Rossi, Juan Pablo Francisco
author_role author
author2 Ferreira Gomes, Mariela Soledad
Pignataro, María Florencia
Strehler, Emanuel E.
Rossi, Juan Pablo Francisco
author2_role author
author
author
author
dc.subject.none.fl_str_mv Plasma Membrane Calcium Pump
Dissociation Constants
Conformational Changes
Transmembrane Domain
Equilibrium Constants
Membrane Proteins
Photoactivatable Phospholipids
topic Plasma Membrane Calcium Pump
Dissociation Constants
Conformational Changes
Transmembrane Domain
Equilibrium Constants
Membrane Proteins
Photoactivatable Phospholipids
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv The purpose of this work was to obtain information about conformational changes of the plasma membrane Ca2+-pump (PMCA) in the membrane region upon interaction with Ca2+, calmodulin (CaM) and acidic phospholipids. To this end, we have quantified labeling of PMCA with the photoactivatable phosphatidylcholine analog [125I]TID-PC/16, measuring the shift of conformation E2 to the auto-inhibited conformation E1I and to the activated E1A state, titrating the effect of Ca2+ under different conditions. Using a similar approach, we also determined the CaM-PMCA dissociation constant. The results indicate that the PMCA possesses a high affinity site for Ca2+ regardless of the presence or absence of activators. Modulation of pump activity is exerted through the C-terminal domain, which induces an apparent auto-inhibited conformation for Ca2+ transport but does not modify the affinity for Ca2+ at the transmembrane domain. The C-terminal domain is affected by CaM and CaM-like treatments driving the auto-inhibited conformation E1I to the activated E1A conformation and thus modulating the transport of Ca2+. This is reflected in the different apparent constants for Ca2+ in the absence of CaM (calculated by Ca2+-ATPase activity) that sharply contrast with the lack of variation of the affinity for the Ca2+ site at equilibrium. This is the first time that equilibrium constants for the dissociation of Ca2+ and CaM ligands from PMCA complexes are measured through the change of transmembrane conformations of the pump. The data further suggest that the transmembrane domain of the PMCA undergoes major rearrangements resulting in altered lipid accessibility upon Ca2+ binding and activation.
Fil: Mangialavori, Irene Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Ferreira Gomes, Mariela Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Pignataro, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
Fil: Strehler, Emanuel E.. Mayo Clinic College of Medicine; Estados Unidos
Fil: Rossi, Juan Pablo Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina
description The purpose of this work was to obtain information about conformational changes of the plasma membrane Ca2+-pump (PMCA) in the membrane region upon interaction with Ca2+, calmodulin (CaM) and acidic phospholipids. To this end, we have quantified labeling of PMCA with the photoactivatable phosphatidylcholine analog [125I]TID-PC/16, measuring the shift of conformation E2 to the auto-inhibited conformation E1I and to the activated E1A state, titrating the effect of Ca2+ under different conditions. Using a similar approach, we also determined the CaM-PMCA dissociation constant. The results indicate that the PMCA possesses a high affinity site for Ca2+ regardless of the presence or absence of activators. Modulation of pump activity is exerted through the C-terminal domain, which induces an apparent auto-inhibited conformation for Ca2+ transport but does not modify the affinity for Ca2+ at the transmembrane domain. The C-terminal domain is affected by CaM and CaM-like treatments driving the auto-inhibited conformation E1I to the activated E1A conformation and thus modulating the transport of Ca2+. This is reflected in the different apparent constants for Ca2+ in the absence of CaM (calculated by Ca2+-ATPase activity) that sharply contrast with the lack of variation of the affinity for the Ca2+ site at equilibrium. This is the first time that equilibrium constants for the dissociation of Ca2+ and CaM ligands from PMCA complexes are measured through the change of transmembrane conformations of the pump. The data further suggest that the transmembrane domain of the PMCA undergoes major rearrangements resulting in altered lipid accessibility upon Ca2+ binding and activation.
publishDate 2010
dc.date.none.fl_str_mv 2010-01
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/11336/18238
Mangialavori, Irene Cecilia; Ferreira Gomes, Mariela Soledad; Pignataro, María Florencia; Strehler, Emanuel E.; Rossi, Juan Pablo Francisco; Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes; American Society for Biochemistry and Molecular Biology; Journal of Biological Chemistry; 285; 1; 1-2010; 123-130
0021-9258
CONICET Digital
CONICET
url http://hdl.handle.net/11336/18238
identifier_str_mv Mangialavori, Irene Cecilia; Ferreira Gomes, Mariela Soledad; Pignataro, María Florencia; Strehler, Emanuel E.; Rossi, Juan Pablo Francisco; Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes; American Society for Biochemistry and Molecular Biology; Journal of Biological Chemistry; 285; 1; 1-2010; 123-130
0021-9258
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1074/jbc.M109.076679
info:eu-repo/semantics/altIdentifier/url/http://www.jbc.org/content/285/1/123.long
info:eu-repo/semantics/altIdentifier/url/https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2804156/
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
dc.publisher.none.fl_str_mv American Society for Biochemistry and Molecular Biology
publisher.none.fl_str_mv American Society for Biochemistry and Molecular Biology
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 13.13397

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