Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.17.1.4 (xanthine dehydrogenase)
1,236 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The xanthine oxidase class of molybdenum enzyzmes requires a terminal sulfur ligand at the active site. It has been proposed that a special sulfurase catalyzes the insertion of this ligand thereby activating the enzymes. Previous analyses of mutants in plants indicated that the genetic locus aba3 is involved in this step leading to activation of the molybdenum enzymes aldehyde oxidase and xanthine dehydrogenase. Here we report the cloning of the aba3 gene from Arabidopsis thaliana and the biochemical characterization of the purified protein. ABA3 is a two-domain protein with a N-terminal NifS-like sulfurase domain and a C-terminal domain that might be involved in recognizing the target enzymes. Molecular analysis of three aba3 mutants identified mutations in both domains. ABA3 contains highly conserved binding motifs for pyridoxal phosphate and for a persulfide. The purified recombinant protein possesses a cysteine desulfurase activity, is yellow in color, and shows a NifS-like change in absorbance in the presence of L-cysteine. Pretreatment of ABA3 with a thiol-specific alkylating reagent inhibited its desulfurase activity. These data indicate a transsulfuration reaction similar to bacterial NifS. In a fully defined in vitro system, the purified protein was able to activate aldehyde oxidase by using L-cysteine as sulfur donor. Finally, we show that the expression of the aba3 gene is inducible by drought-stress.
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PMID:ABA3 is a molybdenum cofactor sulfurase required for activation of aldehyde oxidase and xanthine dehydrogenase in Arabidopsis thaliana. 1155 8

The isc and suf operons in Escherichia coli represent alternative genetic systems optimized to mediate the essential metabolic process of iron-sulfur cluster (Fe-S) assembly under basal or oxidative-stress conditions, respectively. Some of the proteins in these two operons share strong sequence homology, e.g. the cysteine desulfurases IscS and SufS, and presumably play the same role in the oxygen-sensitive assembly process. However, other proteins in these operons share no significant homology and occur in a mutually exclusive manner in Fe-S assembly operons in other organisms (e.g. IscU and SufE). These latter proteins presumably play distinct roles adapted to the different assembly mechanisms used by the two systems. IscU has three invariant cysteine residues that function as a template for Fe-S assembly while accepting a sulfur atom from IscS. SufE, in contrast, does not function as an Fe-S assembly template but has been suggested to function as a shuttle protein that uses a persulfide linkage to a single invariant cysteine residue to transfer a sulfur atom from SufS to an alternative Fe-S assembly template. Here, we present and analyze the 2.0A crystal structure of E.coli SufE. The structure shows that the persulfide-forming cysteine occurs at the tip of a loop with elevated B-factors, where its side-chain is buried from solvent exposure in a hydrophobic cavity located beneath a highly conserved surface. Despite the lack of sequence homology, the core of SufE shows strong structural similarity to IscU, and the sulfur-acceptor site in SufE coincides with the location of the cysteine residues mediating Fe-S cluster assembly in IscU. Thus, a conserved core structure is implicated in mediating the interactions of both SufE and IscU with the mutually homologous cysteine desulfurase enzymes present in their respective operons. A similar core structure is observed in a domain found in a variety of Fe-S cluster containing flavoenzymes including xanthine dehydrogenase, where it also mediates interdomain interactions. Therefore, the core fold of SufE/IscU has been adapted to mediate interdomain interactions in diverse redox protein systems in the course of evolution.
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PMID:The SufE sulfur-acceptor protein contains a conserved core structure that mediates interdomain interactions in a variety of redox protein complexes. 1552 4

The molybdenum cofactor sulfurase ABA3 from Arabidopsis thaliana specifically regulates the activity of the molybdenum enzymes aldehyde oxidase and xanthine dehydrogenase by converting their molybdenum cofactor from the desulfo-form into the sulfo-form. ABA3 is a two-domain protein with an NH2-terminal domain sharing significant similarities to NifS proteins that catalyze the decomposition of l-cysteine to l-alanine and elemental sulfur for iron-sulfur cluster synthesis. Although different in its physiological function, the mechanism of ABA3 for sulfur mobilization was found to be similar to NifS proteins. The protein binds a pyridoxal phosphate cofactor and a substrate-derived persulfide intermediate, and site-directed mutagenesis of strictly conserved binding sites for the cofactor and the persulfide demonstrated that they are essential for molybdenum cofactor sulfurase activity. In vitro, the NifS-like domain of ABA3 activates aldehyde oxidase and xanthine dehydrogenase in the absence of the C-terminal domain, but in vivo, the C-terminal domain is required for proper activation of both target enzymes. In addition to its cysteine desulfurase activity, ABA3-NifS also exhibits selenocysteine lyase activity. Although l-selenocysteine is unlikely to be a natural substrate for ABA3, it is decomposed more efficiently than l-cysteine. Besides mitochondrial AtNFS1 and plastidial AtNFS2, which are both proposed to be involved in iron-sulfur cluster formation, ABA3 is proposed to be a third and cytosolic NifS-like cysteine desulfurase in A. thaliana. However, the sulfur transferase activity of ABA3 is used for post-translational activation of molybdenum enzymes rather than for iron-sulfur cluster assembly.
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PMID:Characterization of the NifS-like domain of ABA3 from Arabidopsis thaliana provides insight into the mechanism of molybdenum cofactor sulfuration. 1556 8

Classical xanthinuria type II is an autosomal recessive disorder characterized by deficiency of xanthine dehydrogenase and aldehyde oxidase activities due to lack of a common sulfido-olybdenum cofactor (MoCo). Two mutations, both in the N-terminal domain of the Human Molybdenum Cofactor Sulfurase (HMCS), were reported in patients with type II xanthinuria. Whereas the N-terminal domain of HMCS was demonstrated to have cysteine desulfurase activity, the C-terminal domain hypothetically transfers the sulfur to the MoCo. We describe the first mutation in the C-terminal domain of HMCS identified in a Bedouin-Arab child presenting with urolithiasis and in an asymptomatic Jewish female. Patients were diagnosed with type II xanthinuria by homozygosity mapping and/or allopurinol loading test. The Bedouin-Arab child was homozygous for a c.2326C>T (p.Arg776Cys) mutation, while the female patient was compound heterozygous for this and a novel c.1034insA (p.Gln347fsStop379) mutation in the N-terminal domain of HMCS. Cosegregation of the homozygous mutant genotype with hypouricemia and hypouricosuria was demonstrated in the Bedouin family. Haplotype analysis indicated that p.Arg776Cys is a recurrent mutation. Arg776 together with six surrounding amino acid residues were found fully conserved and predicted to be buried in homologous eukaryotic MoCo sulfurases. Moreover, Arg776 is conserved in a diversity of eukaryotic and prokaryotic proteins that posses a domain homologous to the C-terminal domain of HMCS. Our findings suggest that Arg776 is essential for a core structure of the C-terminal domain of the HMCS and identification of a mutation at this site may contribute clarifying the mechanism of MoCo sulfuration.
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PMID:Identification and characterization of the first mutation (Arg776Cys) in the C-terminal domain of the Human Molybdenum Cofactor Sulfurase (HMCS) associated with type II classical xanthinuria. 1736 66

The molybdenum cofactor (Moco) containing enzymes aldehyde oxidase and xanthine dehydrogenase (XDH) require for activity a sulfuration step that inserts a terminal sulfur ligand into Moco. XdhC was shown to be essential for the production of active XDH in Rhodobacter capsulatus but is itself not a subunit of the purified enzyme. XdhC binds stoichiometric amounts of Moco and is further able to transfer its bound Moco to XDH. Previous work suggested that XdhC particularly stabilizes the sulfurated form of Moco before the insertion into XDH. In this work, we identify an R. capsulatus l-cysteine desulfurase, NifS4, which is involved in the formation of the Mo=S ligand of Moco. We show that NifS4 interacts with XdhC and not with XDH. NifS4 mobilizes sulfur from l-cysteine by formation of a protein-bound persulfide intermediate and transfers this sulfur further to Moco. This reaction was shown to be more effective than the chemical sulfuration of Moco using sulfide as sulfur source. Further studies clearly showed that Moco is sulfurated before the insertion into XDH, while it is bound to XdhC. Conclusively, XdhC has a versatile role in R. capsulatus: binding of Moco, interaction with NifS4 for the sulfuration of Moco, protection of sulfurated Moco from oxidation, and further transfer to XDH.
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PMID:Identification of a Rhodobacter capsulatus L-cysteine desulfurase that sulfurates the molybdenum cofactor when bound to XdhC and before its insertion into xanthine dehydrogenase. 1764 78

The molybdenum cofactor sulfurase ABA3 from Arabidopsis thaliana is needed for post-translational activation of aldehyde oxidase and xanthine dehydrogenase by transferring a sulfur atom to the desulfo-molybdenum cofactor of these enzymes. ABA3 is a two-domain protein consisting of an NH(2)-terminal NifS-like cysteine desulfurase domain and a C-terminal domain of yet undescribed function. The NH(2)-terminal domain of ABA3 decomposes l-cysteine to yield elemental sulfur, which subsequently is bound as persulfide to a conserved protein cysteinyl residue within this domain. In vivo, activation of aldehyde oxidase and xanthine dehydrogenase also depends on the function of the C-terminal domain, as can be concluded from the A. thaliana aba3/sir3-3 mutant. sir3-3 plants are strongly reduced in aldehyde oxidase and xanthine dehydrogenase activities due to a substitution of arginine 723 by a lysine within the C-terminal domain of the ABA3 protein. Here we present first evidence for the function of the C-terminal domain and show that molybdenum cofactor is bound to this domain with high affinity. Furthermore, cyanide-treated ABA3 C terminus was shown to release thiocyanate, indicating that the molybdenum cofactor bound to the C-terminal domain is present in the sulfurated form. Co-incubation of partially active aldehyde oxidase and xanthine dehydrogenase with ABA3 C terminus carrying sulfurated molybdenum cofactor resulted in stimulation of aldehyde oxidase and xanthine dehydrogenase activity. The data of this work suggest that the C-terminal domain of ABA3 might act as a scaffold protein where prebound desulfo-molybdenum cofactor is converted into sulfurated cofactor prior to activation of aldehyde oxidase and xanthine dehydrogenase.
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PMID:Binding of sulfurated molybdenum cofactor to the C-terminal domain of ABA3 from Arabidopsis thaliana provides insight into the mechanism of molybdenum cofactor sulfuration. 1825

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