Plant Sulfite Oxidase as Novel Producer of H2O2

Abstract

Sulfite oxidase (EC 1.8.3.1) from the plant Arabidopsis thaliana is the smallest eukaryotic molybdenum enzyme consisting of a molybdenum cofactor-binding domain but lacking the heme domain that is known from vertebrate sulfite oxidase. While vertebrate sulfite oxidase is a mitochondrial enzyme with cytochrome c as the physiological electron acceptor, plant sulfite oxidase is localized in peroxisomes and does not react with cytochrome c. Here we describe results that identified oxygen as the terminal electron acceptor for plant sulfite oxidase and hydrogen peroxide as the product of this reaction in addition to sulfate. The latter finding might explain the peroxisomal localization of plant sulfite oxidase. 18 O labeling experiments and the use of catalase provided evidence that plant sulfite oxidase combines its catalytic reaction with a subsequent non-enzymatic step where its reaction product hydrogen peroxide oxidizes another molecule of sulfite. In vitro, for each catalytic cycle plant SO will bring about the oxidation of two molecules of sulfite by one molecule of oxygen. In the plant, sulfite oxidase could be responsible for removing sulfite as a toxic metabolite, which might represent a means to protect the cell against excess of sulfite derived from SO 2 gas in the atmosphere (acid rain) or during the decomposition of sulfur-containing amino acids. Finally we present a model for the metabolic interaction between sulfite and catalase in the peroxisome. Sulfite oxidases (SO)3 from vertebrates (published as EC 1.8.3.1) play an essential role in sulfur detoxification by catalyzing the reaction (1), which is the terminal step in the oxidative degradation of cysteine and methionine. Different electron acceptors were reported to interact with vertebrate SO including cytochrome c, ferricyanide, and oxygen (2-4). In mammals, SO is localized in the intermembrane space of mitochondria (5) where electrons derived from sulfite are passed to cytochrome c, the physiological electron acceptor. Vertebrate SO is a homodimeric protein with monomers subdivided into a Moco domain and a heme domain, as verified by the atomic structure of chicken SO (6).Recently we have described plant SO (7) from Arabidopsis thaliana, which is the fourth molybdenum enzyme present in plants in addition to nitrate reductase, xanthine dehydrogenase, and aldehyde oxidase. Cloning and characterization of plant SO was possible by using sequence homologies to the mammalian counterpart. However, in contrast to the animal enzyme plant SO lacks the heme domain, which is evident from the amino acid sequence, its enzymological and spectral properties (7), and the atomic structure (8). Also its subcellular localization differs from that of animals, in plants we showed SO to be localized in peroxisomes (9). SO is wide spread and highly conserved within the plant kingdom; the SO gene is present in higher and lower plants, and the protein encoded seems to be highly conserved because antibodies directed against Arabidopsis SO detect proteins of the correct size...

The Mononuclear Molybdenum Enzymes

“…It has been shown that the molybdenum center of the plant enzyme reacts with O 2 directly, with H 2 O 2 as the ultimate product of the reaction. 313 It has subsequently been demonstrated, however, that O 2 •− is the immediate product of the reaction, and that H 2 O 2 is formed only indirectly by the spontaneous disproportionation of O 2 •−.314 The observed rate constant for the reaction of reduced enzyme with O 2 exhibits a linear dependence on [O 2 ] ( k ox = 5.3 × 10 4 M −1 s −1 at pH 8.0); given that the rate of reduction of the plant enzyme by sulfite exceeds that accessible by stopped-flow, 308 the oxidative half-reaction is clearly rate-limiting for overall turnover with the plant enzyme. Plant sulfite oxidase is localized in the peroxisome, and it is possible that the superoxide it generates is involved in antimicrobial generation of reactive oxygen species, although an appropriate source of sulfite to provide the reducing equivalents necessary for generation of a superoxide burst upon infection has not been identified.…”

Molybdenum metabolism in plants and crosstalk to iron

“…In this process, substrate-derived electrons are transferred to molecular oxygen with formation of superoxide anions (Byrne et al, 2009) and hydrogen peroxides (Haensch et al, 2006). In the physiological context, SO represents a part of an intracellular sulfite homeostasis network required to prevent plant cells from the toxic effects of endogenously arising sulfite (Brychkova et al, 2007; Lang et al, 2007).…”

Molecular Basis for Enzymatic Sulfite Oxidation

“…In bacteria, sulfite oxidation is often linked to energy-generating processes during chemolithotrophic growth on reduced sulfur compounds (2, 3), whereas both plant and vertebrate sulfite oxidases have been shown to detoxify sulfite arising from the degradation of methionine and cysteine and exposure to sulfur dioxide (4,5).…”