Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )

Autores
Placenti, Maria Agueda; Roman, Ernesto Andres; Gonzalez Flecha, Francisco Luis; Gonzalez-Lebrero, Rodolfo Martin
Año de publicación
2021
Idioma
inglés
Tipo de recurso
documento de conferencia
Estado
versión publicada
Descripción
P-type ATPases are a family of membrane proteins which couple ATPhydrolysis to the transport of substrates across biological membranes. Withinthem, Cu + -ATPases are the most widespread and conserved heavy metal iontransporting ATPases (PIB-ATPases). Its reaction cycle is assumed to bedescribed by the so-called Albers-Post model postulated for the most studied P-ATPases such as the Na + ,K + -ATPase or the Ca 2+ -ATPases. However, as somestructural and functional particularities arise for Cu + -ATPases, several authorsposit some doubts about their reaction cycle mechanism. The aim of our work isto perform a functional characterization of Legionella pneumophila Cu⁺-ATPase(LpCopA) by measuring steady state ATPase activity. Cu + -ATPase activity of theenzyme presents a maximum at ∼37ºC and pH 6.6-6.8. Phospholipids enhanceLpCopA Cu + -ATPase activity in a non-essential mode where optimal activity isachieved at an asolectin mole fraction of 0.15 and an amphiphile-protein ratio of~30000. As described for other P-ATPases, Mg 2+ acts as an essential activator.When evaluating the role of ATP and Cu + in the reaction cycle of LpCopA weobserved that ATPase activity increases as Cu + concentration increases with afunctional dependence that can be described by a sum of two hyperboles. Onthe other hand, the increment on ATP concentration in the reaction mediaproduces an increment of ATPase activity that can be described by a hyperbolaplus a constant value. Based on that, and the [Cu + ] and [ATP] dependencies ofthe best fitting parameters of the functions pointed above, we propose aminimal reaction scheme for LpCopA catalytic mechanism that contemplatestwo enzyme conformations with different affinities for ATP, enzymephosphorylation and binding of at least two Cu + ions with different affinities. Thismodel is compatible with the structural information available and the maincharacteristics of the reaction cycle models for the most characterized P-TypeATPases.
Fil: Placenti, Maria Agueda. 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: Roman, Ernesto Andres. 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: Gonzalez Flecha, Francisco Luis. 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: Gonzalez-Lebrero, Rodolfo Martin. 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
20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences Congress
Brasil
Sociedade Brasileira de Bioquímica e Biología Molecular
Materia
Cu(I) transport ATPase
P-ATPase
Kinetic mechanism
reaction cycle
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:11336/173666
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/173666
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )Placenti, Maria AguedaRoman, Ernesto AndresGonzalez Flecha, Francisco LuisGonzalez-Lebrero, Rodolfo MartinCu(I) transport ATPaseP-ATPaseKinetic mechanismreaction cyclehttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1P-type ATPases are a family of membrane proteins which couple ATPhydrolysis to the transport of substrates across biological membranes. Withinthem, Cu + -ATPases are the most widespread and conserved heavy metal iontransporting ATPases (PIB-ATPases). Its reaction cycle is assumed to bedescribed by the so-called Albers-Post model postulated for the most studied P-ATPases such as the Na + ,K + -ATPase or the Ca 2+ -ATPases. However, as somestructural and functional particularities arise for Cu + -ATPases, several authorsposit some doubts about their reaction cycle mechanism. The aim of our work isto perform a functional characterization of Legionella pneumophila Cu⁺-ATPase(LpCopA) by measuring steady state ATPase activity. Cu + -ATPase activity of theenzyme presents a maximum at ∼37ºC and pH 6.6-6.8. Phospholipids enhanceLpCopA Cu + -ATPase activity in a non-essential mode where optimal activity isachieved at an asolectin mole fraction of 0.15 and an amphiphile-protein ratio of~30000. As described for other P-ATPases, Mg 2+ acts as an essential activator.When evaluating the role of ATP and Cu + in the reaction cycle of LpCopA weobserved that ATPase activity increases as Cu + concentration increases with afunctional dependence that can be described by a sum of two hyperboles. Onthe other hand, the increment on ATP concentration in the reaction mediaproduces an increment of ATPase activity that can be described by a hyperbolaplus a constant value. Based on that, and the [Cu + ] and [ATP] dependencies ofthe best fitting parameters of the functions pointed above, we propose aminimal reaction scheme for LpCopA catalytic mechanism that contemplatestwo enzyme conformations with different affinities for ATP, enzymephosphorylation and binding of at least two Cu + ions with different affinities. Thismodel is compatible with the structural information available and the maincharacteristics of the reaction cycle models for the most characterized P-TypeATPases.Fil: Placenti, Maria Agueda. 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: Roman, Ernesto Andres. 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: Gonzalez Flecha, Francisco Luis. 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: Gonzalez-Lebrero, Rodolfo Martin. 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; Argentina20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences CongressBrasilSociedade Brasileira de Bioquímica e Biología MolecularSociedade Brasileira de Bioquímica e Biología Molecular2021info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectCongresoBookhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/173666Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP ); 20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences Congress; Brasil; 2021; 128-128CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://easyapp.ekmf.com.br/sg/uploads/eventos/evento8/documentos/abstract_book.pdfInternacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-10T13:01:04Zoai:ri.conicet.gov.ar:11336/173666instacron: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:34982025-09-10 13:01:04.849CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
title Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
spellingShingle Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
Placenti, Maria Agueda
Cu(I) transport ATPase
P-ATPase
Kinetic mechanism
reaction cycle
title_short Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
title_full Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
title_fullStr Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
title_full_unstemmed Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
title_sort Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP )
dc.creator.none.fl_str_mv Placenti, Maria Agueda
Roman, Ernesto Andres
Gonzalez Flecha, Francisco Luis
Gonzalez-Lebrero, Rodolfo Martin
author Placenti, Maria Agueda
author_facet Placenti, Maria Agueda
Roman, Ernesto Andres
Gonzalez Flecha, Francisco Luis
Gonzalez-Lebrero, Rodolfo Martin
author_role author
author2 Roman, Ernesto Andres
Gonzalez Flecha, Francisco Luis
Gonzalez-Lebrero, Rodolfo Martin
author2_role author
author
author
dc.subject.none.fl_str_mv Cu(I) transport ATPase
P-ATPase
Kinetic mechanism
reaction cycle
topic Cu(I) transport ATPase
P-ATPase
Kinetic mechanism
reaction cycle
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv P-type ATPases are a family of membrane proteins which couple ATPhydrolysis to the transport of substrates across biological membranes. Withinthem, Cu + -ATPases are the most widespread and conserved heavy metal iontransporting ATPases (PIB-ATPases). Its reaction cycle is assumed to bedescribed by the so-called Albers-Post model postulated for the most studied P-ATPases such as the Na + ,K + -ATPase or the Ca 2+ -ATPases. However, as somestructural and functional particularities arise for Cu + -ATPases, several authorsposit some doubts about their reaction cycle mechanism. The aim of our work isto perform a functional characterization of Legionella pneumophila Cu⁺-ATPase(LpCopA) by measuring steady state ATPase activity. Cu + -ATPase activity of theenzyme presents a maximum at ∼37ºC and pH 6.6-6.8. Phospholipids enhanceLpCopA Cu + -ATPase activity in a non-essential mode where optimal activity isachieved at an asolectin mole fraction of 0.15 and an amphiphile-protein ratio of~30000. As described for other P-ATPases, Mg 2+ acts as an essential activator.When evaluating the role of ATP and Cu + in the reaction cycle of LpCopA weobserved that ATPase activity increases as Cu + concentration increases with afunctional dependence that can be described by a sum of two hyperboles. Onthe other hand, the increment on ATP concentration in the reaction mediaproduces an increment of ATPase activity that can be described by a hyperbolaplus a constant value. Based on that, and the [Cu + ] and [ATP] dependencies ofthe best fitting parameters of the functions pointed above, we propose aminimal reaction scheme for LpCopA catalytic mechanism that contemplatestwo enzyme conformations with different affinities for ATP, enzymephosphorylation and binding of at least two Cu + ions with different affinities. Thismodel is compatible with the structural information available and the maincharacteristics of the reaction cycle models for the most characterized P-TypeATPases.
Fil: Placenti, Maria Agueda. 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: Roman, Ernesto Andres. 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: Gonzalez Flecha, Francisco Luis. 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: Gonzalez-Lebrero, Rodolfo Martin. 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
20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences Congress
Brasil
Sociedade Brasileira de Bioquímica e Biología Molecular
description P-type ATPases are a family of membrane proteins which couple ATPhydrolysis to the transport of substrates across biological membranes. Withinthem, Cu + -ATPases are the most widespread and conserved heavy metal iontransporting ATPases (PIB-ATPases). Its reaction cycle is assumed to bedescribed by the so-called Albers-Post model postulated for the most studied P-ATPases such as the Na + ,K + -ATPase or the Ca 2+ -ATPases. However, as somestructural and functional particularities arise for Cu + -ATPases, several authorsposit some doubts about their reaction cycle mechanism. The aim of our work isto perform a functional characterization of Legionella pneumophila Cu⁺-ATPase(LpCopA) by measuring steady state ATPase activity. Cu + -ATPase activity of theenzyme presents a maximum at ∼37ºC and pH 6.6-6.8. Phospholipids enhanceLpCopA Cu + -ATPase activity in a non-essential mode where optimal activity isachieved at an asolectin mole fraction of 0.15 and an amphiphile-protein ratio of~30000. As described for other P-ATPases, Mg 2+ acts as an essential activator.When evaluating the role of ATP and Cu + in the reaction cycle of LpCopA weobserved that ATPase activity increases as Cu + concentration increases with afunctional dependence that can be described by a sum of two hyperboles. Onthe other hand, the increment on ATP concentration in the reaction mediaproduces an increment of ATPase activity that can be described by a hyperbolaplus a constant value. Based on that, and the [Cu + ] and [ATP] dependencies ofthe best fitting parameters of the functions pointed above, we propose aminimal reaction scheme for LpCopA catalytic mechanism that contemplatestwo enzyme conformations with different affinities for ATP, enzymephosphorylation and binding of at least two Cu + ions with different affinities. Thismodel is compatible with the structural information available and the maincharacteristics of the reaction cycle models for the most characterized P-TypeATPases.
publishDate 2021
dc.date.none.fl_str_mv 2021
dc.type.none.fl_str_mv info:eu-repo/semantics/publishedVersion
info:eu-repo/semantics/conferenceObject
Congreso
Book
http://purl.org/coar/resource_type/c_5794
info:ar-repo/semantics/documentoDeConferencia
status_str publishedVersion
format conferenceObject
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/173666
Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP ); 20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences Congress; Brasil; 2021; 128-128
CONICET Digital
CONICET
url http://hdl.handle.net/11336/173666
identifier_str_mv Steady state kinetic analysis of Legionella pneumophila Cu+ transport ATPase: The activation by Cu+ and ATP ); 20th Internaitional Congress of the INternational Union for Pure Applied Biophysics; 50th Annual Meeting of the Brazilian Society for Biochemistry and Molecular Biology; 45th Congress of Brazilian Biophysics Society anda 13th Brazilian Society on Nuclear Biosciences Congress; Brasil; 2021; 128-128
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://easyapp.ekmf.com.br/sg/uploads/eventos/evento8/documentos/abstract_book.pdf
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
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rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
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dc.coverage.none.fl_str_mv Internacional
dc.publisher.none.fl_str_mv Sociedade Brasileira de Bioquímica e Biología Molecular
publisher.none.fl_str_mv Sociedade Brasileira de Bioquímica e Biología Molecular
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
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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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