Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex

Asclepain f is a papain-like protease previously isolated and characterized from latex of <i>Asclepias fruticosa</i>. This enzyme is a member of the C1 family of cysteine proteases that are synthesized as preproenzymes. The enzyme belongs to the alpha + beta class of proteins, with two d...

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Detalles Bibliográficos
Autores principales: Trejo, Sebastián Alejandro, López, Laura María Isabel, Caffini, Néstor Oscar, Natalucci, Claudia Luisa, Canals, Francesc, Avilés, Francesc X.
Formato: Articulo
Lenguaje:Inglés
Publicado: 2009
Materias:
Ciencias Exactas
Biología
Asclepias fruticosa
Gomphocarpus fruticosus subsp. fruticosus
Plant latex
Cysteine endopeptidase
Cloning
Overexpression in Pichia pastoris
Acceso en línea:http://sedici.unlp.edu.ar/handle/10915/142367
Aporte de:
SEDICI (UNLP) de Universidad Nacional de La Plata
id I19-R120-10915-142367
record_format dspace
institution Universidad Nacional de La Plata
institution_str I-19
repository_str R-120
collection SEDICI (UNLP)
language Inglés
topic Ciencias Exactas
Biología
Asclepias fruticosa
Gomphocarpus fruticosus subsp. fruticosus
Plant latex
Cysteine endopeptidase
Cloning
Overexpression in Pichia pastoris
spellingShingle Ciencias Exactas
Biología
Asclepias fruticosa
Gomphocarpus fruticosus subsp. fruticosus
Plant latex
Cysteine endopeptidase
Cloning
Overexpression in Pichia pastoris
Trejo, Sebastián Alejandro
López, Laura María Isabel
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
Canals, Francesc
Avilés, Francesc X.
Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
topic_facet Ciencias Exactas
Biología
Asclepias fruticosa
Gomphocarpus fruticosus subsp. fruticosus
Plant latex
Cysteine endopeptidase
Cloning
Overexpression in Pichia pastoris
description Asclepain f is a papain-like protease previously isolated and characterized from latex of <i>Asclepias fruticosa</i>. This enzyme is a member of the C1 family of cysteine proteases that are synthesized as preproenzymes. The enzyme belongs to the alpha + beta class of proteins, with two disulfide bridges (Cys22-Cys63 and Cys56-Cys95) in the alpha domain, and another one (Cys150-Cys201) in the beta domain, as was determined by molecular modeling. A full-length 1,152 bp cDNA was cloned by RT-RACE-PCR from latex mRNA. The sequence was predicted as an open reading frame of 340 amino acid residues, of which 16 residues belong to the signal peptide, 113 to the propeptide and 211 to the mature enzyme. The full-length cDNA was ligated to pPICZα vector and expressed in <i>Pichia pastoris</i>. Recombinant asclepain f showed endopeptidase activity on pGlu-Phe-Leu-p-nitroanilide and was identified by PMF-MALDI-TOF MS. Asclepain f is the first peptidase cloned and expressed from mRNA isolated from plant latex, confirming the presence of the preprocysteine peptidase in the latex.
format Articulo
Articulo
author Trejo, Sebastián Alejandro
López, Laura María Isabel
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
Canals, Francesc
Avilés, Francesc X.
author_facet Trejo, Sebastián Alejandro
López, Laura María Isabel
Caffini, Néstor Oscar
Natalucci, Claudia Luisa
Canals, Francesc
Avilés, Francesc X.
author_sort Trejo, Sebastián Alejandro
title Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
title_short Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
title_full Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
title_fullStr Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
title_full_unstemmed Sequencing and characterization of asclepain f: the first cysteine peptidase cDNA cloned and expressed from <i>Asclepias fruticosa</i> latex
title_sort sequencing and characterization of asclepain f: the first cysteine peptidase cdna cloned and expressed from <i>asclepias fruticosa</i> latex
publishDate 2009
url http://sedici.unlp.edu.ar/handle/10915/142367
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