Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)

Autores
Trejo, Sebastián Alejandro; López, Laura María Isabel; Cimino, Cecilia Verónica; Caffini, Néstor Oscar; Natalucci, Claudia Luisa
Año de publicación
2001
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Asclepias fruticosa L. is a small shrub containing latex with proteolytic activity. The crude extract (latex diluted 1:250 and ultracentrifuged) contained 276 μg of protein/mL and the proteolytic activity reached 1.2 caseinolytic U/mL. This enzyme preparation was very stable even after 2 hours at 45°C, but was quickly inactivated after 5 minutes at 80°C. Chromatographic purification was achieved by FPLC using a cation exchanger (SP-Sepharose FF). Thus, a unique proteolitically active fraction could be isolated, being homogeneous by bidimensional electrophoresis and mass spectrometry (Mr = 23,652). The optimum pH range was achieved at 8.5–10.5. The enzyme activity was completely inhibited by specific cysteine peptidases inhibitors. Isoelectric focusing followed by zymogram showed the enzyme had a pI greater than 9.3. The N-terminus sequence (LPDSVDWREKGVVFPIRNQGK) shows a great deal of similarity to those of the other cysteine endopeptidases isolated from latices of Asclepiadaceae even when a high degree of homology could be observed with other plant cysteine endopeptidases.
Centro de Investigación de Proteínas Vegetales
Materia
Biología
Asclepias fruticosa
Asclepiadaceae
Latex
Milkweed
Plant endopeptidases
Nivel de accesibilidad
acceso abierto
Condiciones de uso
http://creativecommons.org/licenses/by-nc-sa/4.0/
Repositorio
SEDICI (UNLP)
Institución
Universidad Nacional de La Plata
OAI Identificador
oai:sedici.unlp.edu.ar:10915/132505
Acceder
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oai_identifier_str oai:sedici.unlp.edu.ar:10915/132505
network_acronym_str SEDICI
repository_id_str 1329
network_name_str SEDICI (UNLP)
spelling Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)Trejo, Sebastián AlejandroLópez, Laura María IsabelCimino, Cecilia VerónicaCaffini, Néstor OscarNatalucci, Claudia LuisaBiologíaAsclepias fruticosaAsclepiadaceaeLatexMilkweedPlant endopeptidasesAsclepias fruticosa L. is a small shrub containing latex with proteolytic activity. The crude extract (latex diluted 1:250 and ultracentrifuged) contained 276 μg of protein/mL and the proteolytic activity reached 1.2 caseinolytic U/mL. This enzyme preparation was very stable even after 2 hours at 45°C, but was quickly inactivated after 5 minutes at 80°C. Chromatographic purification was achieved by FPLC using a cation exchanger (SP-Sepharose FF). Thus, a unique proteolitically active fraction could be isolated, being homogeneous by bidimensional electrophoresis and mass spectrometry (Mr = 23,652). The optimum pH range was achieved at 8.5–10.5. The enzyme activity was completely inhibited by specific cysteine peptidases inhibitors. Isoelectric focusing followed by zymogram showed the enzyme had a pI greater than 9.3. The N-terminus sequence (LPDSVDWREKGVVFPIRNQGK) shows a great deal of similarity to those of the other cysteine endopeptidases isolated from latices of Asclepiadaceae even when a high degree of homology could be observed with other plant cysteine endopeptidases.Centro de Investigación de Proteínas Vegetales2001info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionArticulohttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdf469-477http://sedici.unlp.edu.ar/handle/10915/132505enginfo:eu-repo/semantics/altIdentifier/issn/0277-8033info:eu-repo/semantics/altIdentifier/issn/1573-4943info:eu-repo/semantics/altIdentifier/doi/10.1023/a:1012502412612info:eu-repo/semantics/altIdentifier/pmid/11760121info:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-sa/4.0/Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)reponame:SEDICI (UNLP)instname:Universidad Nacional de La Platainstacron:UNLP2025-11-05T13:11:31Zoai:sedici.unlp.edu.ar:10915/132505Institucionalhttp://sedici.unlp.edu.ar/Universidad públicaNo correspondehttp://sedici.unlp.edu.ar/oai/snrdalira@sedici.unlp.edu.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:13292025-11-05 13:11:32.194SEDICI (UNLP) - Universidad Nacional de La Platafalse
dc.title.none.fl_str_mv Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
title Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
spellingShingle Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
Trejo, Sebastián Alejandro
Biología
Asclepias fruticosa
Asclepiadaceae
Latex
Milkweed
Plant endopeptidases
title_short Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
title_full Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
title_fullStr Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
title_full_unstemmed Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
title_sort Purification and Characterization of a New Plant Endopeptidase Isolated from Latex of Asclepias fruticosa L. (Asclepiadaceae)
dc.creator.none.fl_str_mv Trejo, Sebastián Alejandro
López, Laura María Isabel
Cimino, Cecilia Verónica
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
author Trejo, Sebastián Alejandro
author_facet Trejo, Sebastián Alejandro
López, Laura María Isabel
Cimino, Cecilia Verónica
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
author_role author
author2 López, Laura María Isabel
Cimino, Cecilia Verónica
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
author2_role author
author
author
author
dc.subject.none.fl_str_mv Biología
Asclepias fruticosa
Asclepiadaceae
Latex
Milkweed
Plant endopeptidases
topic Biología
Asclepias fruticosa
Asclepiadaceae
Latex
Milkweed
Plant endopeptidases
dc.description.none.fl_txt_mv Asclepias fruticosa L. is a small shrub containing latex with proteolytic activity. The crude extract (latex diluted 1:250 and ultracentrifuged) contained 276 μg of protein/mL and the proteolytic activity reached 1.2 caseinolytic U/mL. This enzyme preparation was very stable even after 2 hours at 45°C, but was quickly inactivated after 5 minutes at 80°C. Chromatographic purification was achieved by FPLC using a cation exchanger (SP-Sepharose FF). Thus, a unique proteolitically active fraction could be isolated, being homogeneous by bidimensional electrophoresis and mass spectrometry (Mr = 23,652). The optimum pH range was achieved at 8.5–10.5. The enzyme activity was completely inhibited by specific cysteine peptidases inhibitors. Isoelectric focusing followed by zymogram showed the enzyme had a pI greater than 9.3. The N-terminus sequence (LPDSVDWREKGVVFPIRNQGK) shows a great deal of similarity to those of the other cysteine endopeptidases isolated from latices of Asclepiadaceae even when a high degree of homology could be observed with other plant cysteine endopeptidases.
Centro de Investigación de Proteínas Vegetales
description Asclepias fruticosa L. is a small shrub containing latex with proteolytic activity. The crude extract (latex diluted 1:250 and ultracentrifuged) contained 276 μg of protein/mL and the proteolytic activity reached 1.2 caseinolytic U/mL. This enzyme preparation was very stable even after 2 hours at 45°C, but was quickly inactivated after 5 minutes at 80°C. Chromatographic purification was achieved by FPLC using a cation exchanger (SP-Sepharose FF). Thus, a unique proteolitically active fraction could be isolated, being homogeneous by bidimensional electrophoresis and mass spectrometry (Mr = 23,652). The optimum pH range was achieved at 8.5–10.5. The enzyme activity was completely inhibited by specific cysteine peptidases inhibitors. Isoelectric focusing followed by zymogram showed the enzyme had a pI greater than 9.3. The N-terminus sequence (LPDSVDWREKGVVFPIRNQGK) shows a great deal of similarity to those of the other cysteine endopeptidases isolated from latices of Asclepiadaceae even when a high degree of homology could be observed with other plant cysteine endopeptidases.
publishDate 2001
dc.date.none.fl_str_mv 2001
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
Articulo
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://sedici.unlp.edu.ar/handle/10915/132505
url http://sedici.unlp.edu.ar/handle/10915/132505
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/issn/0277-8033
info:eu-repo/semantics/altIdentifier/issn/1573-4943
info:eu-repo/semantics/altIdentifier/doi/10.1023/a:1012502412612
info:eu-repo/semantics/altIdentifier/pmid/11760121
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
http://creativecommons.org/licenses/by-nc-sa/4.0/
Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
eu_rights_str_mv openAccess
rights_invalid_str_mv http://creativecommons.org/licenses/by-nc-sa/4.0/
Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
dc.format.none.fl_str_mv application/pdf
469-477
dc.source.none.fl_str_mv reponame:SEDICI (UNLP)
instname:Universidad Nacional de La Plata
instacron:UNLP
reponame_str SEDICI (UNLP)
collection SEDICI (UNLP)
instname_str Universidad Nacional de La Plata
instacron_str UNLP
institution UNLP
repository.name.fl_str_mv SEDICI (UNLP) - Universidad Nacional de La Plata
repository.mail.fl_str_mv alira@sedici.unlp.edu.ar
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score 13.082534

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