Purification and characterization of oxalate oxidase from wheat seedlings

Abstract

Oxalate oxidase (OxO, EC 1.2.3.4.) was purified to homogeneity from wheat (Triticum aestivum) seedlings by sequential thermal treatment, ultrafiltration, Sephadex G-100 gel filtration and affinity chromatography with concanavalin A. The enzyme was purified 66.11-fold with a recovery of 21.97%. It showed a subunit molecular mass of 32.6 kDa on SDS-PAGE and a native molecular mass of 170 kDa on Sephadex G-150 filtration, suggesting that it is a pentamer. The wheat OxO had a maximum activity at pH 3.5. Its K m for oxalate was 0.21 mM. Chemical modification revealed that cysteine, lysine and carboxylate residues were essential for OxO activity, whereas arginine, serine, threonine and tryptophane residues were not essential.

Increased tolerance to oxidative stress in transgenic tobacco expressing a wheat oxalate oxidase gene via induction of antioxidant enzymes is mediated by H2O2

“…Many plants accumulate significant soluble salts of oxalate in addition to insoluble crystalline calcium oxalate (Kinzel 1989, Libert and Franceschi 1987,). The K m of wheat OxO for oxalic acid is 0.21 m M (Hu and Guo 2009). High levels of oxalic acid (1–1.1 mg g −1 FW) exist in both the wild‐type and the transgenic plants, suggesting that oxalic acid is available for OxO as substrate.…”

“…This protein is found both in the cytoplasm and in the apoplast of wheat and transgenic tobacco transformed with the wheat gf‐2.8 (Berna and Bernier 1997, Lane et al 1993). Lysine, cysteine and carboxylated residues are essential for wheat OxO activity (Hu and Guo 2009). OxO is developmentally regulated and expressed in germinating or maturing seeds (Dumas et al 1995, Lane 2000).…”

Comparative structural modeling and docking studies of oxalate oxidase: Possible implication in enzyme supplementation therapy for urolithiasis

“…Germin family proteins have been widely studied in crop plants due to their diverse role against invading pathogens producing oxalate during pathogenesis. It also regulates release of Ca ++ and H 2 O 2 as two important signal molecules in plants [9]. Oxalate degrading capability of present enzyme is a matter of interest for present study.…”

“…Absence of the three dimensional structures for OxOx of T. aestivum, A. thaliana, S. sclerotiarum, the isoforms C. subvermispora 336 and, C. subvermispora 422 prompted us to construct their homology models. OxOx of T. aestivum is most stable under heat, acidic and proteolytic conditions [9], A. thaliana is species of choice as a model for plant tissue culture experiments that can be easily cultured in lab while microorganisms are more convenient and cheap source of enzyme, therefore bacterial strain S. sclerotiarum found to have OxOx were selected for comparative purpose.…”

Oxalic acid and oxalate oxidase enzyme in Costus pictus D. Don

“…Purification of oxalate oxidase employing heat treatment and ammonium sulphate precipitation was done in barley seedlings (Chiriboga 1966;Kotsira and Clonis 1998;Requena and Bornemann 1999), sorghum (Pundir 1991a) and wheat (Hu and Guo 2009). Purification of oxalate oxidase employing ammonium sulphate precipitation with out heat treatment was done in Amaranthus (Mongkolisirikieat and Srisawan 1987;Goyal et al 1999), maize (Vuletic andŠ ukalovic 2000), barley (Thakur et al 2000) and sorghum leaves (Singh et al 2006).…”