Health, Safety & Environment

Sulfite oxidase

Transient catalytic voltammetry of sulfite oxidase reveals rate limiting conformational changes

Sulfite oxidases are metalloenzymes that' oxidize sulfite to sulfate at a Molybdenum active site. In vertebrate sulfite oxidases, the,electrons generated at the Mo center are transferred to an external,electron acceptor via-a heme domain, which can adopt two conformations: 2-closed conforniation, suitable for internal electron transfer, and an "open" conformation suitable-for intermolecular electron transfer. This conformational change is an integral part of-the catalytic cycle. Sulfite oxidases have been wired to. electrode snrfaces, but their immobilization leads to a significant decrease in their catalytic activity, raising the question of the occurrence of the conformational change when the enzyine is on an electrode. We recorded and quantitatively modeled for the first time the transient response of the catalytic cycle of human sulfite oxidase immobilized on an electrode. We show that conformational changes still occur on the electrode, but at a' ower rate thandn solution, which iS the reason for the decrease in activity of sulfite oiddases upon immobilization.

Zeng, T., Leimkuhler, S., Wollenberger, U., and Fourmond, V.,Transient Catalytic Voltammetry of Sulfite Oxidase Reveals Rate Limiting Conformational Changes, Journal of the American Chemical Society, 2017, 139, 11559-11567.

Sulfite oxidase activity of cytochrome c: role of hydrogen peroxide

In humans, sulfite is generated endogenously by the metabolism of sulfur containing amino acids such as methionine and cysteine.

Sulfite is also formed from exposure to sulfur dioxide, one of the major environmental pollutants.

Sulfite is used as an antioxidant and preservative in dried fruits, vegetables, and beverages such as wine.

Sulfite is also used as a stabilizer in many drugs.

Sulfite toxicity has been associated with allergic reactions characterized by sulfite sensitivity, asthma, and anaphylactic shock. Sulfite is also toxic to neurons and cardiovascular cells. Recent studies suggest that the cytotoxicity of sulfite is mediated by free radicals; however, molecular mechanisms involved in sulfite toxicity are not fully understood. Cytochrome c (cyt c) is known to participate in mitochondrial respiration and has antioxidant and peroxidase activities.

Studies were performed to understand the related mechanism of oxidation of sulfite and radical generation by ferric cytochrome c (Fe3+ cyt c) in the absence and presence of H2O2. Electron paramagnetic resonance (EPR) spin trapping studies using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) were performed with sulfite, Fe3+ cyt c, and H2O2. An EPR spectrum corresponding to the sulfite radical adducts of DMPO (DMPO-SO3 -) was obtained. The amount of DMPO-SO3 - formed from the oxidation of sulfite by the Fe3+ cyt c increased with sulfite concentration. In addition, the amount of DMPO-SO3 - formed by the peroxidase activity of Fe3+ cyt c also increased with sulfite and H2O2 concentration.

From these results, we propose a mechanism in which the Fe3+ cyt c and its peroxidase activity oxidizes sulfite to sulfite radical.

Our results suggest that Fe3+ cyt c could have a novel role in the deleterious effects of sulfite in biological systems due to increased production of sulfite radical.

It also shows that the increased production of sulfite radical may be responsible for neurotoxicity and some of the injuries which occur to humans born with molybdenum cofactor and sulfite oxidase deficiencies.

Velayutham, M., Hemann, C. F., Cardounel, A. J., and Zweier, J. L.,Sulfite Oxidase Activity of Cytochrome c: Role of Hydrogen Peroxide, Biochemistry and biophysics reports, 2016, 5, 96-104.

Enzyme xanthine oxidoreductase

Xanthine oxidoreductase (XOR), which is widely distributed from humans to bacteria, has a key role in purine catabolism, catalyzing two steps of sequential hydroxylation from hypoxanthine to xanthine and from xanthine to urate at its molybdenum cofactor (Moco). Human XOR is considered to be a target of drugs not only for therapy of hyperuricemia and gout, but also potentially for a wide variety of other diseases. In this review, we focus on studies of XOR inhibitors  and their implications for understanding the chemical nature and reaction mechanism of the Moco active site of XOR. We also discuss further experimental or clinical studies that would be helpful to clarify remaining issues.

Nishino T, Okamoto K. Mechanistic insights into xanthine oxidoreductase from development studies of candidate drugs to treat hyperuricemia and gout. J Biol Inorg Chem. 2015 Mar; 20(2):195-207. doi: 10.1007/s00775-014-1210-x. Epub 2014 Dec 12.

Enzyme sulfite oxidase

Sulfite-oxidizing enzymes (SOEs) are molybdenum enzymes that exist in almost all  forms of life where they carry out important functions in protecting cells and organisms against sulfite-induced damage. Due to their nearly ubiquitous presence in living cells, these enzymes can be assumed to be evolutionarily ancient, and this is reflected in the fact that the basic domain architecture and fold structure of all sulfite-oxidizing enzymes studied so far are similar. The Mo centers of all SOEs have five-coordinate square pyramidal coordination geometry,  which incorporates a pyranopterin dithiolene cofactor. However, significant differences exist in the quaternary structure of the enzymes, as well as in the kinetic properties and the nature of the electron acceptors used. In addition, some SOEs also contain an integral heme group that participates in the overall catalytic cycle. Catalytic turnover involves the paramagnetic Mo(V) oxidation state, and EPR spectroscopy, especially high-resolution pulsed EPR spectroscopy,  provides detailed information about the molecular and electronic structure of the Mo center and the Mo-based sulfite oxidation reaction.

Kappler U, Enemark JH. Sulfite-oxidizing enzymes J Biol Inorg Chem. 2015 Mar; 20(2):253-64. doi: 10.1007/s00775-014-1197-3. Epub 2014 Sep 27

See also below: Molybdenum cofactor deficiency in humans: neurological consequences of sulfite oxidase deficiency.

Intramolecular electron transfer in sulfite-oxidizing enzymes: probing the role of aromatic amino acids

Sulfite oxidase (SO) is a molybdoheme enzyme that is important in sulfur catabolism, and mutations in the active site region are known to cause SO deficiency disorder in humans.

This investigation probes the effects that mutating aromatic residues (Y273, W338, and H337) in the molybdenum-containing domain of human SO have on both the intramolecular electron transfer (IET) rate between the molybdenum and iron centers using laser flash photolysis and on catalytic turnover via steady-state kinetic analysis.

The W338 and H337 mutants show large decreases in their IET rate constants (k (ET)) relative to the wild-type values, suggesting the importance of these residues for rapid IET. In contrast, these mutants are catalytically competent and exhibit higher k (cat) values than their corresponding k (ET), implying that these two processes involve different conformational states of the protein.

Redox potential investigations using spectroelectrochemistry revealed that these aromatic residues close to the molybdenum center affect the potential of the presumably distant heme center in the resting state (as shown by the crystal structure of chicken SO), suggesting that the heme may be interacting with these residues during IET and/or catalytic turnover.

These combined results suggest that in solution human SO may adopt different conformations for IET and for catalysis in the presence of the substrate. For IET the H337/W338 surface residues may serve as an alternative-docking site for the heme domain.

The similarities between the mutant and wild-type EPR spectra indicate that the active site geometry around the Mo(V) center is not changed by the mutations studied here.

Rajapakshe, A., Meyers, K. T., Berry, R. E., Tollin, G., and Enemark, J. H., Intramolecular electron transfer in sulfite-oxidizing enzymes: probing the role of aromatic amino acids, Journal of Biological Inorganic Chemistry, 2012, 17, 345-352.

Identity of the exchangeable sulfur-containing ligand at the Mo(V) center of R160Q human sulfite oxidase

In our previous study of the fatal R160Q mutant of human sulfite oxidase (hSO) at low pH (Astashkin et al. J. Am. Chem. Soc. 2008, 130, 8471-8480), a new Mo(V) species, denoted "species 1", was observed at low pH values. Species 1 was ascribed to a six-coordinate Mo(V) center with an exchangeable terminal oxo ligand and an equatorial sulfate group on the basis of pulsed EPR spectroscopy and S-33 and O-17 labeling. Here we report new results for species 1 of R160Q based on substitution of the sulfur-containing ligand by a phosphate group, pulsed EPR spectroscopy in K-a- and W-bands, and extensive density functional theory (DFT) calculations applied to large, more realistic molecular models of the enzyme active site.

The combined results unambiguously show that species 1 has an equatorial sulfite as the only exchangeable ligand. The two types of O-17 signals that are observed arise from the coordinated and remote oxygen atoms of the sulfite ligand. A typical five-coordinate Mo(V) site is compatible with the observed and calculated EPR parameters.

Klein, E. L., Raitsimring, A. M., Astashkin, A. V., Rajapakshe, A., Johnson-Winters, K., Arnold, A. R., Potapov, A., Goldfarb, D., and Enemark, J. H., Identity of the Exchangeable Sulfur-Containing Ligand at the Mo(V) Center of R160Q Human Sulfite Oxidase, Inorganic Chemistry, 2012, 51, 1408-1418.

Sulfite is the main intermediate in the oxidation of sulfur compounds to sulfate, the major product of most dissimilatory sulfur-oxidizing prokaryotes. Two pathways of sulfite oxidation are known: (1) direct oxidation to sulfate catalyzed by a sulfite: acceptor oxidoreductase, which is thought to be a molybdenum-containing enzyme; (2) indirect oxidation under the involvement of the enzymes adenylylsulfate (APS) reductase and ATP sulfurylase and/or adenylylsulfate phosphate adenylyltransferase with APS as an intermediate. Direct oxidation appears to have a wider distribution. In many pro- and also eukaryotes sulfite is formed as a degradative product from molecules containing sulfur as a heteroatom. In these organisms detoxification of sulfite is generally achieved by direct oxidation to sulfate.

Kappler, U. and Dahl, C., Enzymology and molecular biology of prokaryotic sulfite oxidation, Fems Microbiology Letters, 2001, 203, 1-9.

Sulfite oxidase which was deficient in molybdopterin was reconstituted in vitro with the molybdenum cofactor (Moco) synthesized de novo from precursor and molybdate. In vitro reconstitution of the purified apoprotein was achieved using an incubation mixture containing purified precursor, purified molybdopterin synthase, and sodium molybdate.

Leimkuhler, S. and Rajagopalan, K. V., In vitro incorporation of nascent molybdenum cofactor into human sulfite oxidase, Journal of Biological Chemistry, 2001, 276, 1837-1844.

The electronic (charge-transfer) spectrum of the enzyme sulfite oxidase has been probed by temperature-dependent magnetic circular dichroism (MCD) spectroscopy. The enzyme was poised in the catalytically relevant [Mo(V):Fe(II)] state by anaerobic reduction with sulfite in the absence of cytochrome c. A feature at 22 250 cm-1 in the MCD is assigned as the cysteine S(sigma)-->Mo d(xy) charge transfer. The primary role of the co-ordinated cysteine is to decrease the effective nuclear charge on Mo by charge donation to the metal, statically poising the active site at more negative reduction potentials during electron transfer regeneration.

Helton, M.E., Pacheco, A., McMaster, J., Enemark, J. H., and Kirk, M. L., An MCD spectroscopic study of the molybdenum active site in sulfite oxidase: insight into the role of co-ordinated cysteine, Journal of Inorganic Biochemistry, 2000, 80, 227-233.

Molybdenum is an essential constituent of the enzyme hepatic sulfite oxidase. The reduced enzyme, like xanthine oxidase, gives an electron paramagnetic resonance signal of molybdenum(V).

Sheep and cows, develop adverse reactions to feed containing 2-30 ppm molybdenum; horses and pigs tolerate feed with concentrations > 1000 ppm molybdenum [Smyth, 1956].

Cohen, H. J., Fridovich, I. and Rajagopalan, K. V., J. Biol. Chem., 1971, 246 , 374.
Smyth, H.E., Hygienic standard for daily inhalation. Ind Hyg Q, 1956, 17 ,129-185.

Molybdenum K-edge X-ray absorption studies of the oxidized and reduced active sites of the sulfite dehydrogenase from Starkeya novella showed that the molybdenum atom of the oxidized enzyme is bound by two Mo=O ligands at 1.73 angstrom and three thiolate Mo-S ligands at 2.42 angstrom, whereas the reduced enzyme has one oxo at 1.74 angstrom, one long oxygen at 2.19 angstrom (characteristic of Mo-OH2), and three Mo-S ligands at 2.40 angstrom.

Doonan, C. J., Kappler, U., and George, G. N., Structure of the active site of sulfite dehydrogenase from Starkeya novella, Inorganic Chemistry, 2006, 45, 7488-7492.
Enemark, J. H., Astashkin, A. V., and Raitsimring, A. M., Investigation of the coordination structures of the molybdenum(v) sites of sulfite oxidizing enzymes by pulsed EPR spectroscopy, Dalton Transactions, 2006, 3501-3514.
Hemann, C., Hood, B. L., Fulton, M., Hansch, R., Schwarz, G., Mendel, R. R., Kirk, M. L., and Hille, R., Spectroscopic and kinetic studies of Arabidopsis thaliana sulfite oxidase: Nature of the redox-active orbital and electronic structure contributions to catalysis, Journal of the American Chemical Society, 2005, 127, 16567-16577.

The recent developments in our understanding of sulfite oxidizing enzyme mechanisms that are driven by a combination of molecular biology, rapid kinetics, pulsed electron paramagnetic resonance (EPR), and computational techniques are the subject of this review.

Feng, C. J., Tollin, G., and Enernark, J. H., Sulfite oxidizing enzymes, Biochimica et Biophysica Acta-Proteins and Proteomics, 2007, 1774, 527-539.

Cofactor-dependent maturation of mammalian sulfite oxidase links two mitochondrial import pathways

Sulfite oxidase (SO) catalyses the metabolic detoxification of sulfite to sulfate within the intermembrane space of mitochondria. The enzyme follows a complex maturation pathway, including mitochondrial transport and processing, integration of two prosthetic groups, molybdenum cofactor (Moco) and heme, as well as homodimerisation. We have identified the sequential and cofactor-dependent maturation steps of SO. The N-terminal bipartite targeting signal of SO was required but not sufficient for mitochondrial localization. In the absence of Moco, most of the SO, although processed by the inner membrane peptidase of mitochondria, was found in the cytosol. Moco binding was required to induce mitochondrial trapping and retention, thus ensuring unidirectional translocation of SO. In the absence of the N-terminal targeting sequence, SO assembled in the cytosol, suggesting an important function for the leader sequence in preventing premature cofactor binding. In vivo, heme binding and dimerisation did not occur in the absence of Moco and only occurred after Moco integration. In conclusion, the identified molecular hierarchy of SO maturation represents a novel link between the canonical presequence pathway and folding-trap mechanisms of mitochondrial import.

Klein, J. M. and Schwarz, G., Cofactor-dependent maturation of mammalian sulfite oxidase links two mitochondrial import pathways, Journal of Cell Science, 2012, 125, 4876-4885.

Applications of pulsed EPR spectroscopy to structural studies of sulfite oxidizing enzymes

Sulfite oxidizing enzymes (SOEs), including sulfite oxidase (SO) and bacterial sulfite dehydrogenase (SDH), catalyze the oxidation of sulfite (SO32-) to sulfate (SO42-). The active sites of SO and SDH are nearly identical, each having a 5-coordinate, pseudo-square-pyramidal Mo with an axial oxo ligand and three equatorial sulfur donor atoms. One sulfur is from a conserved Cys residue and two are from a pyranopterindithiolene (molybdopterin, MPT) cofactor. The identity of the remaining equatorial ligand, which is solvent-exposed, varies during the catalytic cycle. Numerous in vitro studies, particularly those involving electron paramagnetic resonance (EPR) spectroscopy of the Mo(V) states of SOEs, have shown that the identity and orientation of this exchangeable equatorial ligand depends on the buffer pH, the presence and concentration of certain anions in the buffer, as well as specific point mutations in the protein. Until very recently, however, EPR has not been a practical technique for directly probing specific structures in which the solvent-exposed, exchangeable ligand is an O, OH-, H2O, SO32-, or SO42- group, because the primary O and S isotopes (O-16 and S-32) are magnetically silent (I = 0). This review focuses on the recent advances in the use of isotopic labeling, variable-frequency high resolution pulsed EPR spectroscopy, synthetic model compounds, and DFT calculations to elucidate the roles of various anions. point mutations, and steric factors in the formation, stabilization, and transformation of SOE active site structures. (C) 2012 Elsevier B.V. All rights reserved

Klein, E. L., Astashkin, A. V., Raitsimring, A. M., and Enemark, J. H., Applications of pulsed EPR spectroscopy to structural studies of sulfite oxidizing enzymes, Coordination Chemistry Reviews, 2013, 257, 110-118.

Chloride and sulfite oxidase

Chloro ligands in model oxomolybdenum(V) chloro complexes in an electron spin echo envelope modulation (ESEEM) spectroscopy study had greater ESEEM amplitude than chloride near the oxomolybdenum active site in the high chloride, low-pH form of sulfite oxidase so ruling out equatorial coordination of chloride in the enzyme.

Astashkin, A. V., Klein, E. L., and Enemark, J. H., Toward modeling the high chloride, low pH form of sulfite oxidase: K-a-band ESEEM of equatorial chloro ligands in oxomolybdenum(V) complexes, Journal of Inorganic Biochemistry, 2007, 101, 1623-1629.

Sulfite oxidase coordinated sulfate

Astashkin, A. V., Johnson-Winters, K., Klein, E. L., Byrne, R. S., Hille, R., Raitsimring, A. M., and Enemark, J. H., Direct demonstration of the presence of coordinated sulfate in the reaction pathway of Arabidopsis thaliana sulfite oxidase using S-33 Labeling and ESEEM Spectroscopy, Journal of the American Chemical Society, 2007, 129, 14800-14810

Sulfite oxidase vasorelaxation

The effect of dietary sulphite supplementation on vascular responsiveness in sulphite oxidase (SO)-deficient rats was studied. Increased production of reactive oxygen species and the resultant increment in L-arginine/nitric oxide consumption may play a role in the reduced endothelium-dependent vasorelaxation in sulphite-treated SO-deficient rats.

Nacitarban, C., Kucukatay, V., Sadan, G., Ozturkl, O. H., and Agao, A., Effects of sulphite supplementation on vascular responsiveness in sulphite oxidase-deficient rats, Clinical and Experimental Pharmacology and Physiology, 2008, 35, 268-272.

Mo(V) center of the Y343F mutant of human sulfite oxidase by variable frequency pulsed EPR spectroscopy

Raitsimring, A.M., Astashkin, A. V., Feng, C., Wilson, H. L., Rajagopalan, K. V., and Enemark, J. H., Studies of the Mo(V) center of the Y343F mutant of human sulfite oxidase by variable frequency pulsed EPR spectroscopy, Inorganica Chimica Acta, 2008, 361, 941-946.

Sulfite dehydrogenases (SDHs) catalyze the oxidation and detoxification of sulfite to sulfate, a reaction critical to all forms of life. Sulfite-oxidizing enzymes contain three conserved active site amino acids (Arg-55, His-57, and Tyr-236) that are crucial for catalytic competency. Here we have studied the kinetic and structural effects of two novel and one previously reported substitution (R55M, H57A, Y236F) in these residues on SDH catalysis. Both Arg-55 and His-57 were found to have key roles in substrate binding. An R55M substitution increased K-m(sulfite)(app) by 2-3 orders of magnitude, whereas His-57 was required for maintaining a high substrate affinity at low pH when the imidazole ring is fully protonated. This effect may be mediated by interactions of His-57 with Arg-55 that stabilize the position of the Arg-55 side chain or, alternatively, may reflect changes in the protonation state of sulfite. Unlike what is seen for SDHWT and SDHY236F, the catalytic turnover rates of SDHR55M and SDHH57A are relatively insensitive to pH (similar to 60 and 200 s-1, respectively). On the structural level, striking kinetic effects appeared to correlate with disorder (in SDHH57A and SDHY236F) or absence of Arg-55 (SDHR55M), suggesting that Arg-55 and the hydrogen bonding interactions it engages in are crucial for substrate binding and catalysis. The structure of SDHR55M has sulfate bound at the active site, a fact that coincides with a significant increase in the inhibitory effect of sulfate in SDHR55M. Thus, Arg-55 also appears to be involved in enabling discrimination between the substrate and product in SDH.

Bailey, S., Rapson, T., Johnson-Winters, K., Astashkin, A. V., Enemark, J. H., and Kappler, U., Molecular Basis for Enzymatic Sulfite Oxidation HOW THREE CONSERVED ACTIVE SITE RESIDUES SHAPE ENZYME ACTIVITY, Journal of Biological Chemistry, 2009, 284, 2053-2063.

Sulfite oxidase in plants

The occurrence of sulfite oxidase in plants has been established by identification of a cDNA from Arabidopsis thaliana encoding a functional sulfite oxidase. The aim was to identify herbaceous and woody plants (Azardirachta indica L., Cassia fistula L., Saraca indica L., Spinacea oleracea L., and S Syzyzium cumini L.) with sulfite oxidase activity and to characterize some of its immuno-biochemical properties. The Syzyzium cumini was chosen to characterize sulfite oxidase as it showed maximum enzyme activity in the crude extract as compared to other plants. Absorption spectra of sulfite oxidase revealed two peaks at 235 and 277 nm, but no distinct peak in the visible region. Crude extracts of the plants were studied for immuno-biochemical studies. Despite protein structure-functional similarities between plant and animal sulfite oxidase, no cross-reactivity could be established between the two sources of sulfite oxidase. These data suggested that plants sulfite oxidase, however, differed with regards to their immuno-biochemical properties.

Ahmad, A. and Ahmad, S., Screening and partial immunochemical characterization of sulfite oxidase from plant source, Indian Journal of Experimental Biology, 2010, 48, 83-86.

Sulfite oxidase mechanism molecular dynamics simulations to understand the large-scale domain motions of the enzyme

Molecular dynamics simulations were undertaken to understand the large-scale domain motions of the enzyme. Motion of the N-terminal domain into an orientation similar to that postulated for rapid electron transfer was observed. Simulations also probe the dynamics of the active site and surrounding residues, adding a further level of structural and thermodynamic detail in understanding sulfite oxidase function.

Pushie, M. J. and George, G. N., Active-Site Dynamics and Large-Scale Domain Motions of Sulfite Oxidase: A Molecular Dynamics Study, Journal of Physical Chemistry B, 2010, 114, 3266-3275

Essential molybdenum  -  Molybdenum-dependent sulfite-oxidizing enzymes

Sulfite-oxidizing enzymes (SOEs) are molybdenum-dependent and are found in vertebrates, plants and bacteria. They catalyze the oxidation of sulfite to sulfate :

<SO32− + H2O = SO4 2 − + 2H+ + 2e

The oxygen atom that is incorporated into sulfite comes from water (rather than from dioxygen). In the fully oxidized resting state of the enzymes, the catalytic site is a nearly square-pyramidal dioxo-molybdenum centre that has three equatorial sulfur ligands (one from the conserved cysteinyl side chain and two from the molybdopterin (MPT) cofactor) and two oxo ligands—one equatorial and one axial.The mechanism for SOEs involves attack of sulfite on the electrophilic equatorial oxo ligand that is exposed to solvent, followed by hydrolysis of sulfate and two sequential one-electron oxidations to return the enzyme to the fully oxidized resting state:

<MoVI=O + SO32 − à< MoIV-OSO3, + H2O à< MoV-OH, − SO42 − à<MoVI=O, − H+, − e

In chloride-depleted samples at low pH a blocked form of the enzyme is obtained having molybdenum(V) binding sulfite.

Enemark, J. H., Raitsimring, A. M., Astashkin, A. V., and Klein, E. L., Implications for the mechanism of sulfite oxidizing enzymes from pulsed EPR spectroscopy and DFT calculations for "difficult'' nuclei, Faraday Discussions, 2011, 148, 249-267.

Sulfite oxidase variants having nitrate reductase activity

Eukaryotic sulfite oxidase is a dimeric protein that contains molybdenum cofactor and catalyzes the metabolically essential conversion of sulfite to sulfate as the terminal step in the metabolism of cysteine and methionine.

Nitrate reductase is an evolutionarily related molybdoprotein in lower organisms that is essential for growth on nitrate.

Human and chicken sulfite oxidase variants in which the active site has been modified to alter substrate specificity and activity from sulfite oxidation to nitrate reduction are described.

The crystal structures of the Mo domains of the double and triple mutants were determined to and 2.1 angstrom resolution.

Qiu, J.A., Wilson, H.L., Rajagopalan, K. V. BIOCHEMISTRY, 2012, 51,1134-1147. Structure-Based Alteration of Substrate Specificity and Catalytic Activity of Sulfite Oxidase from Sulfite Oxidation to Nitrate Reduction

Sulfite oxidase
Tungsten inhibition
Alteration of drug metabolizing enzymes in sulphite oxidase deficiency

The aim of this study was to investigate the possible effects of sulphite oxidase (SOX, E.C. 1.8.3.1) deficiency on xenobiotic metabolism. For this purpose, SOX deficiency was produced in rats by the administration of a low molybdenum diet with concurrent addition of 200 ppm tungsten to their drinking water. First, hepatic SOX activity in deficient groups was measured to confirm SOX deficiency. Then, aminopyrine N-demethylase, aniline 4-hydroxylase, aromatase, caffeine N-demethylase, cytochrome b5 reductase, erythromycin N-demethylase, ethoxyresorufin O-deethylase, glutathione S-transferase, N-nitrosodimethylamine N-demethylase and penthoxyresorufin O-deethylase activities were determined to follow changes in the activity of drug metabolizing enzymes in SOX-deficient rats. Our results clearly demonstrated that SOX deficiency significantly elevated A4H, caffeine N-demethylase, erythromycin N-demethylase and N-nitrosodimethylamine N-demethylase activities while decreasing ethoxyresorufin O-deethylase and aromatase activities. These alterations in drug metabolizing enzymes can contribute to the varying susceptibility and response of sulphite-sensitive individuals to different drugs and/or therapeutics used for treatments.

Tutuncu, Begum, Kucukatay, Vural, Arslan, Sevki, Sahin, Barbaros, Semiz, Asli, and Sen, Alaattin, Alteration of drug metabolizing enzymes in sulphite oxidase deficiency, Journal of Clinical Biochemistry and Nutrition, 2012, 51, 50-54.

Catalytic Voltammetry of the Molybdoenzyme Sulfite Dehydrogenase from Sinorhizobium meliloti

Sulfite dehydrogenase from the soil bacterium Sinorhizobium meliloti (SorT) is a periplasmic, homodimeric molybdoenzyme with a molecular mass of 78 kDa. It differs from most other well studied sulfite oxidizing enzymes, as it bears no heme cofactor. SorT does not readily reduce ferrous horse heart cytochrome c which is the preferred electron acceptor for vertebrate sulfite oxidases.

In the present study, ferrocene methanol (FM) (in its oxidized ferrocenium form) was utilized as an artificial electron acceptor for the catalytic SorT sulfite oxidation reaction. Cyclic voltammetry of FM was used to generate the active form of the mediator at the electrode surface. The FM-mediated catalytic sulfite oxidation by SorT was investigated by two different voltammetric methods, namely, (i) SorT freely diffusing in solution and (ii) SorT confined to a thin layer at the electrode surface by a semipermeable dialysis membrane. A single set of rate and equilibrium constants was used to simulate the catalytic voltammograms performed under different sweep rates and with various concentrations of sulfite and FM which provides new insights into the kinetics of the SorT catalytic mechanism. Further, we were able to model the role of the dialysis membrane in the kinetics of the overall catalytic system.

Kalimuthu, P., Kappler, U., and Bernhardt, P. V., Catalytic Voltammetry of the Molybdoenzyme Sulfite Dehydrogenase from Sinorhizobium meliloti, Journal of Physical Chemistry B, 2014, 118, 7091-7099.

Molybdenum Trioxide Nanoparticles with Intrinsic Sulfite Oxidase Activity

Sulfite oxidase is a mitochondria-located molybdenum-containing enzyme catalyzing the oxidation of sulfite to sulfate in the amino acid and lipid metabolism. Therefore, it plays a major role in detoxification processes, where defects in the enzyme cause a severe infant disease leading to early death with no efficient or cost-effective therapy in sight.

Here we report that molybdenum trioxide (MoO3) nanoparticles display an intrinsic biomimetic sulfite oxidase activity under physiological conditions, and, functionalized with a customized bifunctional ligand containing dopamine as anchor group and triphenylphosphonium ion as targeting agent, they selectively target the mitochondria while being highly dispersible in aqueous solutions.

Chemically induced sulfite oxidase knockdown cells treated with MoO3 nanoparticles recovered their sulfite oxidase activity in vitro, which makes MoO3 nanoparticles a potential therapeutic for sulfite oxidase deficiency and opens new avenues for cost-effective therapies for gene-induced deficiencies.

Ragg, R., Natalio, F., Tahir, M. N., Janssen, H., Kashyap, A., Strand, D., Strand, S., and Tremel, W., Molybdenum Trioxide Nanoparticles with Intrinsic Sulfite Oxidase Activity, Acs Nano, 2014, 8, 5182-5189.