Information on EC 3.5.2.17 - hydroxyisourate hydrolase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria

EC NUMBER
COMMENTARY hide
3.5.2.17
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RECOMMENDED NAME
GeneOntology No.
hydroxyisourate hydrolase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
5-hydroxyisourate + H2O = 5-hydroxy-2-oxo-4-ureido-2,5-dihydro-1H-imidazole-5-carboxylate
show the reaction diagram
The reaction is the first stage in the conversion of 5-hydroxyisourate into S-allantoin. This reaction will also occur spontaneously but more slowly
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Metabolic pathways
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Microbial metabolism in diverse environments
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Purine metabolism
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urate conversion to allantoin I
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urate conversion to allantoin II
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urate conversion to allantoin III
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allantoin degradation
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SYSTEMATIC NAME
IUBMB Comments
5-hydroxyisourate amidohydrolase
The reaction is the first stage in the conversion of 5-hydroxyisourate into S-allantoin. This reaction will also occur spontaneously but more slowly.
CAS REGISTRY NUMBER
COMMENTARY hide
255885-20-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Uniprot
Manually annotated by BRENDA team
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UniProt
Manually annotated by BRENDA team
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Uniprot
Manually annotated by BRENDA team
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Manually annotated by BRENDA team
HIUHase 1
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
the enzyme belongs to the 5-hydroxyisourate hydrolase/transthyretin superfamily: evolutionary and functional analyses, overview. Teleosts have highly diverged genomes that resulted from whole genome duplication, which leads to an extensive diversity of paralogous genes. Transthyretin, an extracellular thyroid hormone binding protein, is thought to have evolved from an ancestral 5-hydroxyisourate hydrolase by gene duplication at some stage of chordate evolution. Phylogenetic analysis of the teleost aa sequences reveals the presence of two HIUHase subfamilies, HIUHase 1 (which has an N-terminal peroxisomal targeting signal-2) and HIUHase 2 (which does not have an N-terminal PTS2), and one transthyretin family
metabolism
the enzyme catalyzes the hydrolysis of 5-hydroxyisourate in the purine degradation pathway, and transthyretin, a thyroid hormone binding protein
physiological function
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point mutation Y98Cin the gene encoding mouse HIU hydrolase, Urah, results in undetectable protein expression. Mice homozygous for this mutation develop elevated platelet counts secondary to excess thrombopoietin production and hepatomegaly. The majority of homozygous mutant mice also develop hepatocellular carcinoma, and tumor development is accelerated by exposure to radiation
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
5,6-diaminouracil + H2O
?
show the reaction diagram
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slow alternative substrate
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?
5-hydroxyisourate + H2O
2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline
show the reaction diagram
5-hydroxyisourate + H2O
5-hydroxy-2-oxo-4-ureido-2,5-dihydro-1H-imidazole-5-carboxylate
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
5-hydroxyisourate + H2O
5-hydroxy-2-oxo-4-ureido-2,5-dihydro-1H-imidazole-5-carboxylate
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
the enzyme is not affected by 1 mM of Cr3+, Mg2+, or Ca2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Zn2+
significant inhibition at 0.05 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.026
5,6-diaminouracil
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pH 7.5, 30C
0.015 - 11
5-hydroxyisourate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.21
5,6-diaminouracil
Glycine max
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pH 7.5, 30C
0.015 - 172
5-hydroxyisourate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.18 - 1700
5-hydroxyisourate
2901
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37650
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pH 7.0, 25C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 27
assay at
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.2
estimated value
additional information
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
highest enzyme level
Manually annotated by BRENDA team
additional information
tissue distribution pattern
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
at pH 2-12
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Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
12439
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x * 12439, calculated, x * 15000, SDS-PAGE of recombinant protein
14000
4 * 14000, SDS-PAGE
15000
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x * 12439, calculated, x * 15000, SDS-PAGE of recombinant protein
36600
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4 * 36600, calculated
40000
gel filtration
63730
calculated from cDNA sequence
137000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 12439, calculated, x * 15000, SDS-PAGE of recombinant protein
homotetramer
monomer
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1 * 68000, SDS-PAGE
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
at 4C by the sitting-drop, vapour diffusion method, using 0.1 M HEPES (pH 7.5), 20% (w/v) PEG 10000
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mutants Y116T, I16A/Y116T and mutant I16A/Y116T in complex with thyroxine, to 1.7 A, 2.3 A., and 1.95 A resoultion. Structural comparison of HIUase and transthyretin, TTR. Mutations Y116T and I16A are likely to be crucial events in order to induce, after a gene duplication event, the conversion of the enzyme HIUase into a binding protein, transthyretin. The mutations at the active sites of HIUase open up the two ends of the channel that transverses the entire tetrameric protein, generating two cavities accessible to the thyroxine molecule and abrogating, at the same time, the enzymatic activity
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to 1.8 A resolution, and modeling of substrate 5-hydroxyisourate into the active site. The four chains of the enzyme come together to form a homotetramer with 222 symmetry. The tetramer is a dimer of dimers, with two protomers arranged to create an extended beta-sheet that makes up the dimerdimer interface
hanging drop vapour diffusion method in 0.1 M sodium cacodylate (pH 6.0), 0.2 M MgCl2, 20% (w/v) polyethylene glycol 3350 at 22C
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3
below 3.0 separation into monomeric units
697901
4.6
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below pH 4.6 separation into monomeric units
697901
5
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below pH 5.0 separation into monomeric units
697901
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
66.7
melting temperature, below 66.7C structure stable
93
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melting temperature, below 93.0C structure stable
97.8
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melting temperature, below 97.8C structure stable
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
glutathione-agarose 4B in column mode
glutathione-Sepharose 4B column chromatography
Q-Sepharose column chromatography
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recombinant enzyme
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recombinant soluble His-tagged enzyme from Escherichia coli by metal affinity chromatography
Superose-12 gel filtration and HiTrap SP column chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned and expressed in Escherichia coli
expressed in Escherichia coli BL21 (DE3) cells
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expressed in Escherichia coli BL21-DE3 cells
expressed in Escherichia coli Rosetta BL21 cells
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expressed in Escherichia coli strani BL21
expression in Escherichia coli
expression in Escherichia coli strain BL21
His-tagged enzyme expessed in Escherichia coli BL21 (DE3)pLysS
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phylogenetic analysis, sequence comparisons, quantitative real-time PCR enzyme expression analysis, recombinant expression of soluble His-tagged enzyme in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D50N
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shows 50% activity of the wild type enzyme
I16A
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mutation at the active sites of HIUase, opens up one end of the channel that transverses the entire tetrameric protein, generating a cavity accessible to the thyroxine molecule and abrogating, at the same time, the enzymatic activity
I16A/Y116T
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mutations at the active sites of HIUase open up the two ends of the channel that transverses the entire tetrameric protein, generating two cavities accessible to the thyroxine molecule and abrogating, at the same time, the enzymatic activity
Y116T
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mutation at the active sites of HIUase, opens up one end of the channel that transverses the entire tetrameric protein, generating a cavity accessible to the thyroxine molecule and abrogating, at the same time, the enzymatic activity
H101A
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10% of wild-type activity
H7A
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3% of wild-type activity
Y114F
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22% of wild-type activity
H7N
dramatic decrease in activity
H92N
dramatic decrease in activity
R41K
about 90% decrease in activity
S108A
about 50% decrease in activity
H7N
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dramatic decrease in activity
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H92N
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dramatic decrease in activity
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R41K
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about 90% decrease in activity
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S108A
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about 50% decrease in activity
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H102N
10fold reduced activity
H11N
mutant almost abolishes activity
R51E
mutant almost abolishes activity
R51K
mutant fails to affect activity
S118A
mutation has no influence on activity
Y98C
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point mutation in the gene encoding mouse HIU hydrolase, Urah, that perturbes uric acid metabolism within the liver. The substitution of cysteine for tyrosine in a conserved helical region results in undetectable protein expression. Mice homozygous for this mutation develop elevated platelet counts secondary to excess thrombopoietin production and hepatomegaly. The majority of homozygous mutant mice also develop hepatocellular carcinoma, and tumor development is accelerated by exposure to radiation
additional information
the deletion mutation DELTAYRGS greatly affects activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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point mutation Y98Cin the gene encoding mouse HIU hydrolase, Urah, results in undetectable protein expression. Mice homozygous for this mutation develop elevated platelet counts secondary to excess thrombopoietin production and hepatomegaly. The majority of homozygous mutant mice also develop hepatocellular carcinoma, and tumor development is accelerated by exposure to radiation
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