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Artículo

Horseradish peroxidase production from Spodoptera frugiperda larvae: A simple and inexpensive method

Targovnik, Alexandra MarisaIcon ; Romero, Lucía VirginiaIcon ; Wolman, Federico JavierIcon ; Cascone, OsvaldoIcon ; Miranda, Maria VictoriaIcon
Fecha de publicación: 06/2010
Editorial: Elsevier
Revista: Process Biochemistry
ISSN: 0032-9592
e-ISSN: 1359-5113
Idioma: Inglés
Tipo de recurso: Artículo publicado
Clasificación temática:
Alimentos y Bebidas

Resumen

Horseradish peroxidase is used in many biotechnological fields including diagnostics, biocatalysts and biosensors. Horseradish peroxidase isozyme C (HRPC) was extracellularly expressed in Spodoptera frugiperda Sf9 cell culture and in intact larvae. At day 6 post-infection, the concentration of active HRPC in suspension cultures was 3.0 ± 0.1 μg per 1 × 106 cells or 3.0 ± 0.1 mg l-1 with a multiplicity of infection of 1 in the presence of 7.2 μM hemin. Similar yields were obtained in monolayer cultures. In larvae, the HRPC expression level was 137 ± 17 mg HRPC kg-1 larvae at day 6 post-infection with a single larvae thus producing approximately 41 μg HRPC. The whole larval extract was separated by ion exchange chromatography and HRPC was purified in a single step with a yield of 75% and a purification factor of 117. The molecular weight of recombinant HRPC was 44,016 Da, and its glycosylation pattern agreed with that expected for invertebrates. The Km and Vmax were 12.1 ± 1.7 mM and 2673 ± 113 U mg-1, respectively, similar to those of HRP purified from Armoracia rusticana roots. The method described in this study, based on overexpression of HRPC in S. frugiperda larvae, is a simple and inexpensive way to obtain high levels of active enzyme for research and other biotechnological applications. © 2010 Elsevier Ltd. All rights reserved.
Palabras clave: Expression , Horseradish Peroxidase , Insect Larvae , Purification , Spodoptera Frugiperda
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info:eu-repo/semantics/openAccess Excepto donde se diga explícitamente, este item se publica bajo la siguiente descripción: Creative Commons Attribution-NonCommercial-ShareAlike 2.5 Unported (CC BY-NC-SA 2.5)
Identificadores
URI: http://hdl.handle.net/11336/76744
URL: http://www.sciencedirect.com/science/article/pii/S1359511310000413
DOI: https://doi.org/10.1016/j.procbio.2010.02.003
Colecciones
Articulos(IBYME)
Articulos de INST.DE BIOLOGIA Y MEDICINA EXPERIMENTAL (I)
Articulos(OCA HOUSSAY)
Articulos de OFICINA DE COORDINACION ADMINISTRATIVA HOUSSAY
Citación
Targovnik, Alexandra Marisa; Romero, Lucía Virginia; Wolman, Federico Javier; Cascone, Osvaldo; Miranda, Maria Victoria; Horseradish peroxidase production from Spodoptera frugiperda larvae: A simple and inexpensive method; Elsevier; Process Biochemistry; 45; 6; 6-2010; 835-840
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