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

Paper ID Volume ID Publish Year Pages File Format Full-Text
35600 45098 2010 6 PDF Available
Title
Horseradish peroxidase production from Spodoptera frugiperda larvae: A simple and inexpensive method
Abstract

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.

Keywords
Horseradish peroxidase; Spodoptera frugiperda; Expression; Insect larvae; Purification
First Page Preview
Horseradish peroxidase production from Spodoptera frugiperda larvae: A simple and inexpensive method
Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 45, Issue 6, June 2010, Pages 835–840
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering