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The identification of an integral membrane, cytochrome c urate oxidase completes the catalytic repertoire of a therapeutic enzyme.

Doniselli N, Monzeglio E, Dal Palù A, Merli A, Percudani R - Sci Rep (2015)

Bottom Line: In contrast with the known soluble Uox, the identified gene (puuD) encodes a membrane protein with a C-terminal cytochrome c.The 8-helix transmembrane domain corresponds to DUF989, a family without similarity to known proteins.These findings identify a missing link in purine catabolism, assign a biochemical activity to a domain of unknown function (DUF989), and complete the catalytic repertoire of an enzyme useful for human therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, University of Parma, Italy.

ABSTRACT
In living organisms, the conversion of urate into allantoin requires three consecutive enzymes. The pathway was lost in hominid, predisposing humans to hyperuricemia and gout. Among other species, the genomic distribution of the two last enzymes of the pathway is wider than that of urate oxidase (Uox), suggesting the presence of unknown genes encoding Uox. Here we combine gene network analysis with association rule learning to identify the missing urate oxidase. In contrast with the known soluble Uox, the identified gene (puuD) encodes a membrane protein with a C-terminal cytochrome c. The 8-helix transmembrane domain corresponds to DUF989, a family without similarity to known proteins. Gene deletion in a PuuD-encoding organism (Agrobacterium fabrum) abolished urate degradation capacity; the phenotype was fully restored by complementation with a cytosolic Uox from zebrafish. Consistent with H2O2 production by zfUox, urate oxidation in the complemented strain caused a four-fold increase of catalase. No increase was observed in the wild-type, suggesting that urate oxidation by PuuD proceeds through cytochrome c-mediated electron transfer. These findings identify a missing link in purine catabolism, assign a biochemical activity to a domain of unknown function (DUF989), and complete the catalytic repertoire of an enzyme useful for human therapy.

No MeSH data available.


Related in: MedlinePlus

Identification of COG3748 as urate oxidase.(a) Pathway for the conversion of urate into allantoin. (b) String association network obtained with COG3648, COG2351, and COG3195. Nodes represent gene families according to the COG classification. Edges represent predicted functional associations; stronger associations are shown as thicker lines. The node identified as a candidate urate oxidase (COG3748) is indicated with an arrow. (c) Map of urate oxidation capacity in complete genomes. The tree represents 431 distinct species possessing either the uox, hpxO, hpyO, or COG3748 genes or both the urah and urad genes. The presence (red) or the absence (blue) of genes in complete genomes is shown alongside the organism tree. Main taxonomic groups and organisms discussed in the text are indicated.
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f1: Identification of COG3748 as urate oxidase.(a) Pathway for the conversion of urate into allantoin. (b) String association network obtained with COG3648, COG2351, and COG3195. Nodes represent gene families according to the COG classification. Edges represent predicted functional associations; stronger associations are shown as thicker lines. The node identified as a candidate urate oxidase (COG3748) is indicated with an arrow. (c) Map of urate oxidation capacity in complete genomes. The tree represents 431 distinct species possessing either the uox, hpxO, hpyO, or COG3748 genes or both the urah and urad genes. The presence (red) or the absence (blue) of genes in complete genomes is shown alongside the organism tree. Main taxonomic groups and organisms discussed in the text are indicated.

Mentions: In Bacteria, Archaea, and Eukaryotes, where present, the oxidative conversion of urate into allantoin proceeds through a three-step enzymatic pathway5. In the first step, uric acid (urate at neutral pH) is converted to 5-hydroxyisourate (HIU) in an oxygen-dependent reaction6. In the second step, HIU is hydrolysed to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU), which is decarboxylated to give dextrorotatory allantoin in the third step (Fig. 1a). As HIU and OHCU are metastable compounds with a half-life of 7.2 and 9.6 minutes at physiological conditions, racemic allantoin is obtained in vitro as a final product of the Uox reaction. However, in nature the presence of Uox is almost invariably associated with both HIU hydrolase (Urah) and OHCU decarboxylase (Urad). Besides affecting the reaction stereochemistry7, the presence of these enzymes appears to be important for the rapid elimination of the metastable intermediates of urate oxidation8.


The identification of an integral membrane, cytochrome c urate oxidase completes the catalytic repertoire of a therapeutic enzyme.

Doniselli N, Monzeglio E, Dal Palù A, Merli A, Percudani R - Sci Rep (2015)

Identification of COG3748 as urate oxidase.(a) Pathway for the conversion of urate into allantoin. (b) String association network obtained with COG3648, COG2351, and COG3195. Nodes represent gene families according to the COG classification. Edges represent predicted functional associations; stronger associations are shown as thicker lines. The node identified as a candidate urate oxidase (COG3748) is indicated with an arrow. (c) Map of urate oxidation capacity in complete genomes. The tree represents 431 distinct species possessing either the uox, hpxO, hpyO, or COG3748 genes or both the urah and urad genes. The presence (red) or the absence (blue) of genes in complete genomes is shown alongside the organism tree. Main taxonomic groups and organisms discussed in the text are indicated.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4562309&req=5

f1: Identification of COG3748 as urate oxidase.(a) Pathway for the conversion of urate into allantoin. (b) String association network obtained with COG3648, COG2351, and COG3195. Nodes represent gene families according to the COG classification. Edges represent predicted functional associations; stronger associations are shown as thicker lines. The node identified as a candidate urate oxidase (COG3748) is indicated with an arrow. (c) Map of urate oxidation capacity in complete genomes. The tree represents 431 distinct species possessing either the uox, hpxO, hpyO, or COG3748 genes or both the urah and urad genes. The presence (red) or the absence (blue) of genes in complete genomes is shown alongside the organism tree. Main taxonomic groups and organisms discussed in the text are indicated.
Mentions: In Bacteria, Archaea, and Eukaryotes, where present, the oxidative conversion of urate into allantoin proceeds through a three-step enzymatic pathway5. In the first step, uric acid (urate at neutral pH) is converted to 5-hydroxyisourate (HIU) in an oxygen-dependent reaction6. In the second step, HIU is hydrolysed to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU), which is decarboxylated to give dextrorotatory allantoin in the third step (Fig. 1a). As HIU and OHCU are metastable compounds with a half-life of 7.2 and 9.6 minutes at physiological conditions, racemic allantoin is obtained in vitro as a final product of the Uox reaction. However, in nature the presence of Uox is almost invariably associated with both HIU hydrolase (Urah) and OHCU decarboxylase (Urad). Besides affecting the reaction stereochemistry7, the presence of these enzymes appears to be important for the rapid elimination of the metastable intermediates of urate oxidation8.

Bottom Line: In contrast with the known soluble Uox, the identified gene (puuD) encodes a membrane protein with a C-terminal cytochrome c.The 8-helix transmembrane domain corresponds to DUF989, a family without similarity to known proteins.These findings identify a missing link in purine catabolism, assign a biochemical activity to a domain of unknown function (DUF989), and complete the catalytic repertoire of an enzyme useful for human therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, University of Parma, Italy.

ABSTRACT
In living organisms, the conversion of urate into allantoin requires three consecutive enzymes. The pathway was lost in hominid, predisposing humans to hyperuricemia and gout. Among other species, the genomic distribution of the two last enzymes of the pathway is wider than that of urate oxidase (Uox), suggesting the presence of unknown genes encoding Uox. Here we combine gene network analysis with association rule learning to identify the missing urate oxidase. In contrast with the known soluble Uox, the identified gene (puuD) encodes a membrane protein with a C-terminal cytochrome c. The 8-helix transmembrane domain corresponds to DUF989, a family without similarity to known proteins. Gene deletion in a PuuD-encoding organism (Agrobacterium fabrum) abolished urate degradation capacity; the phenotype was fully restored by complementation with a cytosolic Uox from zebrafish. Consistent with H2O2 production by zfUox, urate oxidation in the complemented strain caused a four-fold increase of catalase. No increase was observed in the wild-type, suggesting that urate oxidation by PuuD proceeds through cytochrome c-mediated electron transfer. These findings identify a missing link in purine catabolism, assign a biochemical activity to a domain of unknown function (DUF989), and complete the catalytic repertoire of an enzyme useful for human therapy.

No MeSH data available.


Related in: MedlinePlus