Purification and some properties of rat liver cysteine oxidase (cysteine dioxygenase)

Sakakibara, Shigeki; Yamagata, Kenji; Hosokawa, Yu; Kohashi, Noriyuki; Ueda, Iwao; Sakamoto, Yukiya

doi:10.1016/0005-2744(76)90138-8

Cited by 43 publications

(25 citation statements)

References 7 publications

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“…Reconstitution of the rat apoenzyme with various transition metals confirmed that iron was required for activity, in accord with the earlier conclusions (1). Moreover, the recombinant rat enzyme was active without a second interacting factor, despite previous reports suggesting that additional components were required (12,13).…”

mentioning

confidence: 47%

Structure and mechanism of mouse cysteine dioxygenase

McCoy

Bailey

Bitto

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

191

238

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Cysteine dioxygenase (CDO) catalyzes the oxidation of L-cysteine to cysteine sulfinic acid. Deficiencies in this enzyme have been linked to autoimmune diseases and neurological disorders. The x-ray crystal structure of CDO from Mus musculus was solved to a nominal resolution of 1.75 Å. The sequence is 91% identical to that of a human homolog. The structure reveals that CDO adopts the typical ␤-barrel fold of the cupin superfamily. The NE2 atoms of His-86, -88, and -140 provide the metal binding site. The structure further revealed a covalent linkage between the side chains of Cys-93 and Tyr-157, the cysteine of which is conserved only in eukaryotic proteins. Metal analysis showed that the recombinant enzyme contained a mixture of iron, nickel, and zinc, with increased iron content associated with increased catalytic activity. Details of the predicted active site are used to present and discuss a plausible mechanism of action for the enzyme.cupin ͉ cysteine metabolism ͉ O2-activation M ouse cysteine dioxygenase (CDO) catalyzes the initial step in the biochemical pathway used for oxidation of cysteine to sulfate (1), namely the oxidation of L-cysteine to cysteine sulfinic acid as shown in Fig. 1. The enzyme activity has important medical implications because elevated cysteine levels have been associated with Parkinson's and Alzheimer's diseases (2). High cysteine-to-sulfate ratios have been observed in patients suffering from systemic lupus erthematosus and rheumatoid arthritis (3, 4). Moreover, the Hallervorden-Spatz syndrome, a neurological disorder associated with iron accumulation, has been linked to a decline in CDO activity (5).CDO displays significant sequence identity with some members of the cupin superfamily (6), which have a conserved ␤-barrel fold and share two conserved sequence motifs: G(X) 5 HXH(X) 3,4 E(X) 6 G and G(X) 5 PXG(X) 2 H(X) 3 N (6-8).The two His and Glu residues from the first motif and the His from the second motif coordinate the metal ion in germin, the superfamily archetype (9). The Mus musculus CDO sequence contains the first motif with the exception of the glutamate, which is replaced by cysteine. This substitution is conserved in other eukaryotic CDOs. The second motif is less conserved, and only the His and Asn residues are present in the mouse CDO.CDO does not require an external reductant ( Fig. 1) and incorporates both oxygen atoms from O 2 (10), which justifies the dioxygenase classification, but relatively little else is known about the reaction mechanism. The recombinant enzyme from Rattus norvegicus has been purified and characterized by steadystate kinetics (11); the mouse enzyme investigated here has an identical sequence. Reconstitution of the rat apoenzyme with various transition metals confirmed that iron was required for activity, in accord with the earlier conclusions (1). Moreover, the recombinant rat enzyme was active without a second interacting factor, despite previous reports suggesting that additional components were required (12, 13).Here, we describe the x-ra...

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mentioning

confidence: 47%

Structure and mechanism of mouse cysteine dioxygenase

McCoy

Bailey

Bitto

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

191

238

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“…The possibility that an active site sulfhydryl group might be oxidized to a disulfide, but could be reduced by cysteamine, was an attractive possibility given the literature on the activity of the enzyme with preincubation with cysteine (18). However, CDO treated with the sulfhydryl-binding agent iodoacetamide had no effect on the enzyme activity.…”

Section: Resultsmentioning

confidence: 99%

“…CDO is a metalloenzyme that requires ferrous ion to be active (18). We also investigated the inhibition of recombinant CDO using metalbinding anions.…”

Section: Resultsmentioning

confidence: 99%

Probes of the Catalytic Site of Cysteine Dioxygenase

Chai

Bruyere

Maroney

2006

Journal of Biological Chemistry

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The first major step of cysteine catabolism, the oxidation of cysteine to cysteine sulfinic acid, is catalyzed by cysteine dioxygenase (CDO). In the present work, we utilize recombinant rat liver CDO and cysteine derivatives to elucidate structural parameters involved in substrate recognition and x-ray absorption spectroscopy to probe the interaction of the active site iron center with cysteine. Kinetic studies using cysteine structural analogs show that most are inhibitors and that a terminal functional group bearing a negative charge (e.g. a carboxylate) is required for binding. The substrate-binding site has no stringent restrictions with respect to the size of the amino acid. Lack of the amino or carboxyl groups at the ␣-carbon does not prevent the molecules from interacting with the active site. In fact, cysteamine is shown to be a potent activator of the enzyme without being a substrate. CDO was also rendered inactive upon complexation with the metal-binding inhibitors azide and cyanide. Unlike many non-heme iron dioxygenases that employ ␣-keto acids as cofactors, CDO was shown to be the only dioxygenase known to be inhibited by ␣-ketoglutarate.The first major step in cysteine catabolism involves its conversion to cysteine sulfinic acid by cysteine dioxygenase (CDO), 2 which is a nonheme iron-containing dioxygenase present in mammalian tissues. CDO plays an important role in the formation of essential metabolites such as taurine and sulfate. Because cysteine is toxic at high levels (1), CDO assists in maintaining low intracellular cysteine levels without compromising its availability for incorporation into proteins and synthesis of major metabolites.Since the early studies on CDO by Sörbo and Ewetz (2), there have been great advances in understanding the regulation and biochemical properties of this enzyme. However, there is little information regarding the active site of CDO and its specificity for the only known substrate, L-cysteine (3).Herein we report the results of probing the active site requirements of CDO with cysteine structural analogs and the substrate binding mode with x-ray absorption spectroscopy. In addition to exploring the features required for enzyme inhibition, using substrate derivatives has attracted attention in the development of potent and target-specific drugs (4, 5). Because CDO is at a crucial branching point of cysteine catabolism, and several diseases have been linked to this metalloenzyme (6 -8), information about substrate recognition also provides insights for drug design. Many of the cysteine analogs utilized in our current work are metabolites that are present in mammalian cells, and their interaction with CDO could aid in unraveling thiol homeostasis and regulation. MATERIALS AND METHODSChemicals-Cysteine, cysteine sulfinic acid, and heptafluorobutyric acid were obtained from Aldrich Chemical Co. Sodium phosphate, sodium chloride, and ferrous sulfate were purchased from Fisher Scientific. Yeast extract, Tryptone, phenylmethylsulfonyl fluoride, ampicillin, chloramphenicol...

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“…The rat (296), human (232), and Caenorhabditis elegans genes have been well characterized, with the closest bacterial relatives of these eukaryotic sequences being those from B. subtilis (gi2635598), Streptomyces coelicolor (gi2687337), and Mycobacterium tuberculosis (gi2896702). This enzyme is known to be monomeric, with one atom of iron per molecule (312); its activity is strongly reduced by chelators of Cu ϩ and Fe 2ϩ (247).…”

Section: Dioxygenasesmentioning

confidence: 99%

Microbial Relatives of the Seed Storage Proteins of Higher Plants: Conservation of Structure and Diversification of Function during Evolution of the Cupin Superfamily

Dunwell

Khuri

Gane

2000

Microbiol Mol Biol Rev

313

297

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SUMMARY This review summarizes the recent discovery of the cupin superfamily (from the Latin term “cupa,” a small barrel) of functionally diverse proteins that initially were limited to several higher plant proteins such as seed storage proteins, germin (an oxalate oxidase), germin-like proteins, and auxin-binding protein. Knowledge of the three-dimensional structure of two vicilins, seed proteins with a characteristic β-barrel core, led to the identification of a small number of conserved residues and thence to the discovery of several microbial proteins which share these key amino acids. In particular, there is a highly conserved pattern of two histidine-containing motifs with a varied intermotif spacing. This cupin signature is found as a central component of many microbial proteins including certain types of phosphomannose isomerase, polyketide synthase, epimerase, and dioxygenase. In addition, the signature has been identified within the N-terminal effector domain in a subgroup of bacterial AraC transcription factors. As well as these single-domain cupins, this survey has identified other classes of two-domain bicupins including bacterial gentisate 1,2-dioxygenases and 1-hydroxy-2-naphthoate dioxygenases, fungal oxalate decarboxylases, and legume sucrose-binding proteins. Cupin evolution is discussed from the perspective of the structure-function relationships, using data from the genomes of several prokaryotes, especially Bacillus subtilis. Many of these functions involve aspects of sugar metabolism and cell wall synthesis and are concerned with responses to abiotic stress such as heat, desiccation, or starvation. Particular emphasis is also given to the oxalate-degrading enzymes from microbes, their biological significance, and their value in a range of medical and other applications.

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Purification and some properties of rat liver cysteine oxidase (cysteine dioxygenase)

Cited by 43 publications

References 7 publications

Structure and mechanism of mouse cysteine dioxygenase

Structure and mechanism of mouse cysteine dioxygenase

Probes of the Catalytic Site of Cysteine Dioxygenase

Microbial Relatives of the Seed Storage Proteins of Higher Plants: Conservation of Structure and Diversification of Function during Evolution of the Cupin Superfamily

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