Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates

Autores
Sanchez, Julieta Maria; Perillo, Maria Angelica
Año de publicación
2002
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
We investigated if in complex environments, like those where β-galactosidase activity is usually assayed, the kinetics of hydrolysis against soluble substrates could be modulated through enzyme-surface interactions. Kinetic parameters were determined using ortho-nitrophenyl-β-D-galactopiranoside (ONPG) as substrate, in the absence or presence of multilamellar vesicles (MLVs) of pure phosphatidyl cholines (PCs) or PCs:cholesterol mixtures, by visible spectroscopy. Light scattering was carefully corrected by three different methods obtaining similar results. The spectroscopic behavior of the reaction product in the presence of liposomes was also taken into account in order to avoid overestimating the reaction rate calculated from absorbance data. At low [PC] (<0.0024 mM) KM and Vmax decreased compared with the control in the absence of lipids. At high [PC] (1.2 mM), enzyme interaction with highly packed bilayers of dpPC induced an increment in both kinetic parameters. Both kinetic parameters decreased upon the interaction with low packed bilayers (soybean PC) at very low concentration (24 μM) but at higher concentration (1.2 mM) only an increment in Vmax was observed. The dpPC MLVs samples used were four times bigger than those of PCsoybean (approximately 1 μm mean diameter) as measured by quasi elastic light scattering. The increments in Vmax were due to a modulation of the kinetics of the enzymatic reaction and not to non-enzymatic hydrolysis of ONPG at the vesicle-water interface. Enzyme-membrane interaction was confirmed using monomolecular-layers at the air-water-interface. Interestingly, β-galactosidase showed a higher tendency to be localized at a lipid-water interface compared with the free air-water interface; membrane penetration was favored in lower packed membranes. Differences in surface curvature, and thus in surface molecular packing and hydration, might account for the effects observed as the main modulating factor. Our results suggest that β-galactosidase activity was differentially modulated according to the enzyme possibility to penetrate or just be adsorbed to a dimensionality restricted space. © 2002 Elsevier Science B.V. All rights reserved.
Fil: Sanchez, Julieta Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina
Fil: Perillo, Maria Angelica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina
Materia
&Beta;-Galactosidase
Activity Modulation
Adsorption
Enzyme-Membrane Interaction
Monomolecular Layers
Penetration
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:11336/66452

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network_name_str CONICET Digital (CONICET)
spelling Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substratesSanchez, Julieta MariaPerillo, Maria Angelica&Beta;-GalactosidaseActivity ModulationAdsorptionEnzyme-Membrane InteractionMonomolecular LayersPenetrationhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1We investigated if in complex environments, like those where β-galactosidase activity is usually assayed, the kinetics of hydrolysis against soluble substrates could be modulated through enzyme-surface interactions. Kinetic parameters were determined using ortho-nitrophenyl-β-D-galactopiranoside (ONPG) as substrate, in the absence or presence of multilamellar vesicles (MLVs) of pure phosphatidyl cholines (PCs) or PCs:cholesterol mixtures, by visible spectroscopy. Light scattering was carefully corrected by three different methods obtaining similar results. The spectroscopic behavior of the reaction product in the presence of liposomes was also taken into account in order to avoid overestimating the reaction rate calculated from absorbance data. At low [PC] (<0.0024 mM) KM and Vmax decreased compared with the control in the absence of lipids. At high [PC] (1.2 mM), enzyme interaction with highly packed bilayers of dpPC induced an increment in both kinetic parameters. Both kinetic parameters decreased upon the interaction with low packed bilayers (soybean PC) at very low concentration (24 μM) but at higher concentration (1.2 mM) only an increment in Vmax was observed. The dpPC MLVs samples used were four times bigger than those of PCsoybean (approximately 1 μm mean diameter) as measured by quasi elastic light scattering. The increments in Vmax were due to a modulation of the kinetics of the enzymatic reaction and not to non-enzymatic hydrolysis of ONPG at the vesicle-water interface. Enzyme-membrane interaction was confirmed using monomolecular-layers at the air-water-interface. Interestingly, β-galactosidase showed a higher tendency to be localized at a lipid-water interface compared with the free air-water interface; membrane penetration was favored in lower packed membranes. Differences in surface curvature, and thus in surface molecular packing and hydration, might account for the effects observed as the main modulating factor. Our results suggest that β-galactosidase activity was differentially modulated according to the enzyme possibility to penetrate or just be adsorbed to a dimensionality restricted space. © 2002 Elsevier Science B.V. All rights reserved.Fil: Sanchez, Julieta Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Perillo, Maria Angelica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaElsevier Science2002-01info: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/11336/66452Sanchez, Julieta Maria; Perillo, Maria Angelica; Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates; Elsevier Science; Colloids and Surfaces B: Biointerfaces; 24; 1; 1-2002; 21-310927-7765CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1016/S0927-7765(01)00216-8info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0927776501002168info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:12:58Zoai:ri.conicet.gov.ar:11336/66452instacron: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 10:12:59.027CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
title Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
spellingShingle Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
Sanchez, Julieta Maria
&Beta;-Galactosidase
Activity Modulation
Adsorption
Enzyme-Membrane Interaction
Monomolecular Layers
Penetration
title_short Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
title_full Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
title_fullStr Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
title_full_unstemmed Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
title_sort Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates
dc.creator.none.fl_str_mv Sanchez, Julieta Maria
Perillo, Maria Angelica
author Sanchez, Julieta Maria
author_facet Sanchez, Julieta Maria
Perillo, Maria Angelica
author_role author
author2 Perillo, Maria Angelica
author2_role author
dc.subject.none.fl_str_mv &Beta;-Galactosidase
Activity Modulation
Adsorption
Enzyme-Membrane Interaction
Monomolecular Layers
Penetration
topic &Beta;-Galactosidase
Activity Modulation
Adsorption
Enzyme-Membrane Interaction
Monomolecular Layers
Penetration
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv We investigated if in complex environments, like those where β-galactosidase activity is usually assayed, the kinetics of hydrolysis against soluble substrates could be modulated through enzyme-surface interactions. Kinetic parameters were determined using ortho-nitrophenyl-β-D-galactopiranoside (ONPG) as substrate, in the absence or presence of multilamellar vesicles (MLVs) of pure phosphatidyl cholines (PCs) or PCs:cholesterol mixtures, by visible spectroscopy. Light scattering was carefully corrected by three different methods obtaining similar results. The spectroscopic behavior of the reaction product in the presence of liposomes was also taken into account in order to avoid overestimating the reaction rate calculated from absorbance data. At low [PC] (<0.0024 mM) KM and Vmax decreased compared with the control in the absence of lipids. At high [PC] (1.2 mM), enzyme interaction with highly packed bilayers of dpPC induced an increment in both kinetic parameters. Both kinetic parameters decreased upon the interaction with low packed bilayers (soybean PC) at very low concentration (24 μM) but at higher concentration (1.2 mM) only an increment in Vmax was observed. The dpPC MLVs samples used were four times bigger than those of PCsoybean (approximately 1 μm mean diameter) as measured by quasi elastic light scattering. The increments in Vmax were due to a modulation of the kinetics of the enzymatic reaction and not to non-enzymatic hydrolysis of ONPG at the vesicle-water interface. Enzyme-membrane interaction was confirmed using monomolecular-layers at the air-water-interface. Interestingly, β-galactosidase showed a higher tendency to be localized at a lipid-water interface compared with the free air-water interface; membrane penetration was favored in lower packed membranes. Differences in surface curvature, and thus in surface molecular packing and hydration, might account for the effects observed as the main modulating factor. Our results suggest that β-galactosidase activity was differentially modulated according to the enzyme possibility to penetrate or just be adsorbed to a dimensionality restricted space. © 2002 Elsevier Science B.V. All rights reserved.
Fil: Sanchez, Julieta Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina
Fil: Perillo, Maria Angelica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; Argentina
description We investigated if in complex environments, like those where β-galactosidase activity is usually assayed, the kinetics of hydrolysis against soluble substrates could be modulated through enzyme-surface interactions. Kinetic parameters were determined using ortho-nitrophenyl-β-D-galactopiranoside (ONPG) as substrate, in the absence or presence of multilamellar vesicles (MLVs) of pure phosphatidyl cholines (PCs) or PCs:cholesterol mixtures, by visible spectroscopy. Light scattering was carefully corrected by three different methods obtaining similar results. The spectroscopic behavior of the reaction product in the presence of liposomes was also taken into account in order to avoid overestimating the reaction rate calculated from absorbance data. At low [PC] (<0.0024 mM) KM and Vmax decreased compared with the control in the absence of lipids. At high [PC] (1.2 mM), enzyme interaction with highly packed bilayers of dpPC induced an increment in both kinetic parameters. Both kinetic parameters decreased upon the interaction with low packed bilayers (soybean PC) at very low concentration (24 μM) but at higher concentration (1.2 mM) only an increment in Vmax was observed. The dpPC MLVs samples used were four times bigger than those of PCsoybean (approximately 1 μm mean diameter) as measured by quasi elastic light scattering. The increments in Vmax were due to a modulation of the kinetics of the enzymatic reaction and not to non-enzymatic hydrolysis of ONPG at the vesicle-water interface. Enzyme-membrane interaction was confirmed using monomolecular-layers at the air-water-interface. Interestingly, β-galactosidase showed a higher tendency to be localized at a lipid-water interface compared with the free air-water interface; membrane penetration was favored in lower packed membranes. Differences in surface curvature, and thus in surface molecular packing and hydration, might account for the effects observed as the main modulating factor. Our results suggest that β-galactosidase activity was differentially modulated according to the enzyme possibility to penetrate or just be adsorbed to a dimensionality restricted space. © 2002 Elsevier Science B.V. All rights reserved.
publishDate 2002
dc.date.none.fl_str_mv 2002-01
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/11336/66452
Sanchez, Julieta Maria; Perillo, Maria Angelica; Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates; Elsevier Science; Colloids and Surfaces B: Biointerfaces; 24; 1; 1-2002; 21-31
0927-7765
CONICET Digital
CONICET
url http://hdl.handle.net/11336/66452
identifier_str_mv Sanchez, Julieta Maria; Perillo, Maria Angelica; Membrane adsorption or penetration differentially modulates β-galactosidase activity against soluble substrates; Elsevier Science; Colloids and Surfaces B: Biointerfaces; 24; 1; 1-2002; 21-31
0927-7765
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1016/S0927-7765(01)00216-8
info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0927776501002168
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
dc.publisher.none.fl_str_mv Elsevier Science
publisher.none.fl_str_mv Elsevier Science
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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