Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems

Autores
Mariani, Maria Elisa; Madoery, Ricardo Román; Fidelio, Gerardo Daniel
Año de publicación
2015
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Two secretory phospholipase A2 (sPLA2s) from Glycine max, GmsPLA2-IXA-1 and GmsPLA2-XIB-2, have been purified as recombinant proteins and the activity was evaluated in order to obtain the optimum conditions for catalysis using mixed micelles and lipid monolayers as substrate. Both sPLA2s showed a maximum enzyme activity at pH 7 and a requirement of Ca2þ in the micromolar range. These parameters were similar to those found for animal sPLA2s but a surprising optimum temperature for catalysis at 60 C was observed. The effect of negative interfacial charges on the hydrolysis of organized substrates was evaluated through initial rate measurements using short chain phospholipids with different head groups. The enzymes showed subtle differences in the specificity for phospholipids with different head groups (DLPC, DLPG, DLPE, DLPA) in presence or absence of NaCl. Both recombinant enzymes showed lower activity toward anionic phospholipids and a preference for the zwitterionic ones. The values of the apparent kinetic parameters (Vmax and KM) demonstrated that these enzymes have more affinity for phosphatidylcholine compared with phosphatidylglycerol, in contrast with the results observed for pancreatic sPLA2. A hopping mode of catalysis was proposed for the action of these sPLA2 on mixed phospholipid/triton micelles. On the other hand, Langmuir-monolayers assays indicated an optimum lateral surface pressure for activity in between 13 and 16 mN/m for both recombinant enzymes.
Fil: Mariani, Maria Elisa. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); Argentina
Fil: Madoery, Ricardo Román. Universidad Nacional de Córdoba. Facultad de Ciencias Agropecuarias; Argentina
Fil: Fidelio, Gerardo Daniel. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); Argentina
Materia
Glycine Max Phospholipase A2
Spla2s Substrate Preference
Kinetics
Characterization
Interfacial Catalysis
Soybean Spla2s
Hopping Mode of Lipolysis
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/10622
Acceder
id CONICETDig_e85ad69a5815b8636f6b4770f9947b6f
oai_identifier_str oai:ri.conicet.gov.ar:11336/10622
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systemsMariani, Maria ElisaMadoery, Ricardo RománFidelio, Gerardo DanielGlycine Max Phospholipase A2Spla2s Substrate PreferenceKineticsCharacterizationInterfacial CatalysisSoybean Spla2sHopping Mode of Lipolysishttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Two secretory phospholipase A2 (sPLA2s) from Glycine max, GmsPLA2-IXA-1 and GmsPLA2-XIB-2, have been purified as recombinant proteins and the activity was evaluated in order to obtain the optimum conditions for catalysis using mixed micelles and lipid monolayers as substrate. Both sPLA2s showed a maximum enzyme activity at pH 7 and a requirement of Ca2þ in the micromolar range. These parameters were similar to those found for animal sPLA2s but a surprising optimum temperature for catalysis at 60 C was observed. The effect of negative interfacial charges on the hydrolysis of organized substrates was evaluated through initial rate measurements using short chain phospholipids with different head groups. The enzymes showed subtle differences in the specificity for phospholipids with different head groups (DLPC, DLPG, DLPE, DLPA) in presence or absence of NaCl. Both recombinant enzymes showed lower activity toward anionic phospholipids and a preference for the zwitterionic ones. The values of the apparent kinetic parameters (Vmax and KM) demonstrated that these enzymes have more affinity for phosphatidylcholine compared with phosphatidylglycerol, in contrast with the results observed for pancreatic sPLA2. A hopping mode of catalysis was proposed for the action of these sPLA2 on mixed phospholipid/triton micelles. On the other hand, Langmuir-monolayers assays indicated an optimum lateral surface pressure for activity in between 13 and 16 mN/m for both recombinant enzymes.Fil: Mariani, Maria Elisa. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); ArgentinaFil: Madoery, Ricardo Román. Universidad Nacional de Córdoba. Facultad de Ciencias Agropecuarias; ArgentinaFil: Fidelio, Gerardo Daniel. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); ArgentinaElsevier France-editions Scientifiques Medicales Elsevier2015-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/10622Mariani, Maria Elisa; Madoery, Ricardo Román; Fidelio, Gerardo Daniel; Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems; Elsevier France-editions Scientifiques Medicales Elsevier; Biochimie; 108; 1-2015; 48-580300-9084enginfo:eu-repo/semantics/altIdentifier/doi/10.1016/j.biochi.2014.10.016info:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0300908414003022info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-nd/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:37:16Zoai:ri.conicet.gov.ar:11336/10622instacron: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:34982025-09-29 10:37:16.37CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
title Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
spellingShingle Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
Mariani, Maria Elisa
Glycine Max Phospholipase A2
Spla2s Substrate Preference
Kinetics
Characterization
Interfacial Catalysis
Soybean Spla2s
Hopping Mode of Lipolysis
title_short Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
title_full Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
title_fullStr Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
title_full_unstemmed Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
title_sort Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems
dc.creator.none.fl_str_mv Mariani, Maria Elisa
Madoery, Ricardo Román
Fidelio, Gerardo Daniel
author Mariani, Maria Elisa
author_facet Mariani, Maria Elisa
Madoery, Ricardo Román
Fidelio, Gerardo Daniel
author_role author
author2 Madoery, Ricardo Román
Fidelio, Gerardo Daniel
author2_role author
author
dc.subject.none.fl_str_mv Glycine Max Phospholipase A2
Spla2s Substrate Preference
Kinetics
Characterization
Interfacial Catalysis
Soybean Spla2s
Hopping Mode of Lipolysis
topic Glycine Max Phospholipase A2
Spla2s Substrate Preference
Kinetics
Characterization
Interfacial Catalysis
Soybean Spla2s
Hopping Mode of Lipolysis
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Two secretory phospholipase A2 (sPLA2s) from Glycine max, GmsPLA2-IXA-1 and GmsPLA2-XIB-2, have been purified as recombinant proteins and the activity was evaluated in order to obtain the optimum conditions for catalysis using mixed micelles and lipid monolayers as substrate. Both sPLA2s showed a maximum enzyme activity at pH 7 and a requirement of Ca2þ in the micromolar range. These parameters were similar to those found for animal sPLA2s but a surprising optimum temperature for catalysis at 60 C was observed. The effect of negative interfacial charges on the hydrolysis of organized substrates was evaluated through initial rate measurements using short chain phospholipids with different head groups. The enzymes showed subtle differences in the specificity for phospholipids with different head groups (DLPC, DLPG, DLPE, DLPA) in presence or absence of NaCl. Both recombinant enzymes showed lower activity toward anionic phospholipids and a preference for the zwitterionic ones. The values of the apparent kinetic parameters (Vmax and KM) demonstrated that these enzymes have more affinity for phosphatidylcholine compared with phosphatidylglycerol, in contrast with the results observed for pancreatic sPLA2. A hopping mode of catalysis was proposed for the action of these sPLA2 on mixed phospholipid/triton micelles. On the other hand, Langmuir-monolayers assays indicated an optimum lateral surface pressure for activity in between 13 and 16 mN/m for both recombinant enzymes.
Fil: Mariani, Maria Elisa. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); Argentina
Fil: Madoery, Ricardo Román. Universidad Nacional de Córdoba. Facultad de Ciencias Agropecuarias; Argentina
Fil: Fidelio, Gerardo Daniel. Universidad Nacional de Cordoba. Facultad de Ciencias Quimicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Química Biológica de Córdoba (p); Argentina
description Two secretory phospholipase A2 (sPLA2s) from Glycine max, GmsPLA2-IXA-1 and GmsPLA2-XIB-2, have been purified as recombinant proteins and the activity was evaluated in order to obtain the optimum conditions for catalysis using mixed micelles and lipid monolayers as substrate. Both sPLA2s showed a maximum enzyme activity at pH 7 and a requirement of Ca2þ in the micromolar range. These parameters were similar to those found for animal sPLA2s but a surprising optimum temperature for catalysis at 60 C was observed. The effect of negative interfacial charges on the hydrolysis of organized substrates was evaluated through initial rate measurements using short chain phospholipids with different head groups. The enzymes showed subtle differences in the specificity for phospholipids with different head groups (DLPC, DLPG, DLPE, DLPA) in presence or absence of NaCl. Both recombinant enzymes showed lower activity toward anionic phospholipids and a preference for the zwitterionic ones. The values of the apparent kinetic parameters (Vmax and KM) demonstrated that these enzymes have more affinity for phosphatidylcholine compared with phosphatidylglycerol, in contrast with the results observed for pancreatic sPLA2. A hopping mode of catalysis was proposed for the action of these sPLA2 on mixed phospholipid/triton micelles. On the other hand, Langmuir-monolayers assays indicated an optimum lateral surface pressure for activity in between 13 and 16 mN/m for both recombinant enzymes.
publishDate 2015
dc.date.none.fl_str_mv 2015-01
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/11336/10622
Mariani, Maria Elisa; Madoery, Ricardo Román; Fidelio, Gerardo Daniel; Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems; Elsevier France-editions Scientifiques Medicales Elsevier; Biochimie; 108; 1-2015; 48-58
0300-9084
url http://hdl.handle.net/11336/10622
identifier_str_mv Mariani, Maria Elisa; Madoery, Ricardo Román; Fidelio, Gerardo Daniel; Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems; Elsevier France-editions Scientifiques Medicales Elsevier; Biochimie; 108; 1-2015; 48-58
0300-9084
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1016/j.biochi.2014.10.016
info:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0300908414003022
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier France-editions Scientifiques Medicales Elsevier
publisher.none.fl_str_mv Elsevier France-editions Scientifiques Medicales Elsevier
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 13.070432

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