skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris

Abstract

High-level secretory expression of wheat (Triticum aestivum) germin/oxalate oxidase was achieved in Pichia pastoris fermentation cultures as an {alpha}-mating factor signal peptide fusion, based on the native wheat cDNA coding sequence. The oxalate oxidase activity of the recombinant enzyme is substantially increased (7-fold) by treatment with sodium periodate, followed by ascorbate reduction. Using these methods, approximately 1 g (4 x 10{sup 4} U) of purified, activated enzyme was obtained following eight days of induction of a high density Pichia fermentation culture, demonstrating suitability for large-scale production of oxalate oxidase for biotechnological applications. Characterization of the recombinant protein shows that it is glycosylated, with N-linked glycan attached at Asn47. For potential biomedical applications, a nonglycosylated (S49A) variant was also prepared which retains essentially full enzyme activity, but exhibits altered protein-protein interactions.

Authors:
 [1];  [1];  [2];  [2];  [2];  [3]
  1. Department of Environmental and Biomolecular Systems, Oregon Health and Sciences University, 20000 N.W. Walker Road, Beaverton, OR 97006-8921 (United States)
  2. IBMP-Institut de Botanique, Strasbourg (France)
  3. Department of Environmental and Biomolecular Systems, Oregon Health and Sciences University, 20000 N.W. Walker Road, Beaverton, OR 97006-8921 (United States). E-mail: jim@ebs.ogi.edu
Publication Date:
OSTI Identifier:
20991343
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 356; Journal Issue: 4; Other Information: DOI: 10.1016/j.bbrc.2007.03.097; PII: S0006-291X(07)00547-5; Copyright (c) 2007 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; BIOTECHNOLOGY; ENZYME ACTIVITY; FERMENTATION; GLYCOPROTEINS; MATING; OXALATES; OXIDASES; PEPTIDES; PERIODATES; SODIUM; WHEAT

Citation Formats

Pan, Heng-Yen, Whittaker, Mei M., Bouveret, Romaric, Berna, Anne, Bernier, Francois, and Whittaker, James W.. Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris. United States: N. p., 2007. Web. doi:10.1016/j.bbrc.2007.03.097.
Pan, Heng-Yen, Whittaker, Mei M., Bouveret, Romaric, Berna, Anne, Bernier, Francois, & Whittaker, James W.. Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris. United States. doi:10.1016/j.bbrc.2007.03.097.
Pan, Heng-Yen, Whittaker, Mei M., Bouveret, Romaric, Berna, Anne, Bernier, Francois, and Whittaker, James W.. Fri . "Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris". United States. doi:10.1016/j.bbrc.2007.03.097.
@article{osti_20991343,
title = {Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris},
author = {Pan, Heng-Yen and Whittaker, Mei M. and Bouveret, Romaric and Berna, Anne and Bernier, Francois and Whittaker, James W.},
abstractNote = {High-level secretory expression of wheat (Triticum aestivum) germin/oxalate oxidase was achieved in Pichia pastoris fermentation cultures as an {alpha}-mating factor signal peptide fusion, based on the native wheat cDNA coding sequence. The oxalate oxidase activity of the recombinant enzyme is substantially increased (7-fold) by treatment with sodium periodate, followed by ascorbate reduction. Using these methods, approximately 1 g (4 x 10{sup 4} U) of purified, activated enzyme was obtained following eight days of induction of a high density Pichia fermentation culture, demonstrating suitability for large-scale production of oxalate oxidase for biotechnological applications. Characterization of the recombinant protein shows that it is glycosylated, with N-linked glycan attached at Asn47. For potential biomedical applications, a nonglycosylated (S49A) variant was also prepared which retains essentially full enzyme activity, but exhibits altered protein-protein interactions.},
doi = {10.1016/j.bbrc.2007.03.097},
journal = {Biochemical and Biophysical Research Communications},
number = 4,
volume = 356,
place = {United States},
year = {Fri May 18 00:00:00 EDT 2007},
month = {Fri May 18 00:00:00 EDT 2007}
}