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Information on EC 3.5.2.3 - dihydroorotase and Organism(s) Escherichia coli and UniProt Accession P05020

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EC Tree
     3 Hydrolases
         3.5 Acting on carbon-nitrogen bonds, other than peptide bonds
             3.5.2 In cyclic amides
                3.5.2.3 dihydroorotase
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This record set is specific for:
Escherichia coli
UNIPROT: P05020 not found.
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Word Map
The taxonomic range for the selected organisms is: Escherichia coli
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
dihydroorotase, dho, dhoase, dihydroorotase domain, hudhoase, type i dhoase, human dhoase domain, type ii dho, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
carbamoylaspartic dehydrase
-
-
-
-
DHOase
-
-
-
-
dihydroorotate dehydrolase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(S)-dihydroorotate + H2O = N-carbamoyl-L-aspartate
show the reaction diagram
unique catalytic mechanism
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of cyclic amides
-
-
-
-
formation of cyclic amides
-
-
-
-
PATHWAY SOURCE
PATHWAYS
-
-, -, -
SYSTEMATIC NAME
IUBMB Comments
(S)-dihydroorotate amidohydrolase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9024-93-5
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(S)-dihydroorotate + H2O
N-carbamoyl-L-aspartate
show the reaction diagram
-
-
-
r
dihydroorotate + H2O
N-carbamoyl-L-aspartate
show the reaction diagram
-
-
-
r
L-dihydroorotate + H2O
N-carbamoyl-L-aspartate
show the reaction diagram
N-carbamoyl-L-aspartate
(S)-dihydroorotate + H2O
show the reaction diagram
-
-
-
r
N-carbamoyl-L-aspartate
L-dihydroorotate + H2O
show the reaction diagram
thio-dihydroorotate + H2O
?
show the reaction diagram
-
-
-
?
dihydro-DL-orotate + H2O
N-carbamoyl-L-aspartate
show the reaction diagram
-
-
-
-
r
L-carbamoylaspartate
L-dihydroorotate + H2O
show the reaction diagram
-
-
-
-
r
L-dihydroorotate + H2O
N-carbamoyl-L-aspartate
show the reaction diagram
-
-
-
-
r
N-carbamoyl-DL-aspartate
dihydroorotate + H2O
show the reaction diagram
-
-
-
-
r
N-carbamoyl-L-aspartate
L-dihydroorotate + H2O
show the reaction diagram
-
-
-
-
r
thio-dihydroorotate + H2O
?
show the reaction diagram
-
-
-
-
r
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
N-carbamoyl-L-aspartate
L-dihydroorotate + H2O
show the reaction diagram
-
-
-
-
r
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-oxo-1,2,3,6-tetrahydropyrimidine-4,6-dicarboxylate
-
5-Fluoroorotate
(S)-1,2,3,6-tetrahydro-6-oxopyridine-2-carboxylic acid
-
competitive inhibitor versus dihydroorotate and thio-dihydroorotate
4(R)-hydroxy-6-oxo-piperidine-2(S)-carboxylic acid
-
competitive inhibitor versus dihydroorotate and thio-dihydroorotate
4(S)-hydroxy-6-oxo-piperidine-2(S)-carboxylic acid
-
competitive inhibitor versus dihydroorotate and thio-dihydroorotate
4,6-dioxo-piperidine-2-(S)-carboxylic acid
-
most active competitive inhibitor versus dihydroorotate and thio-dihydroorotate
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.51 - 15
carbamoyl aspartate
0.08 - 0.7
dihydroorotate
1.5 - 6
N-(aminocarbonyl)-L-aspartic acid
0.009 - 0.03
thiodihydroorotate
0.076
dihydro-DL-orotate
-
-
1.07
N-carbamoyl-DL-aspartate
-
-
additional information
carbamoyl aspartate
mutants D250A, D250H, D250N, R20Q, R20M, N44A, H254N inactive
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
11.5 - 181.1
N-(aminocarbonyl)-L-aspartic acid
127
dihydro-DL-orotate
-
-
195
N-carbamoyl-DL-aspartate
-
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.3
(S)-1,2,3,6-tetrahydro-6-oxopyridine-2-carboxylic acid
-
with thio-dihydroorotate as substrate, pH. 8.0
1.6
4(R)-hydroxy-6-oxo-piperidine-2(S)-carboxylic acid
-
with thio-dihydroorotate as substrate, pH. 8.0
3
4(S)-hydroxy-6-oxo-piperidine-2(S)-carboxylic acid
-
with thio-dihydroorotate as substrate, pH. 8.0
0.076 - 0.44
4,6-dioxo-piperidine-2-(S)-carboxylic acid
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.8
-
cyclisation of L-carbamoylaspartate
8
-
hydrolysis of L-dihydroorotate
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 6.9
synthesis reaction
7.8 - 8.5
hydrolysis reaction
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
38300
-
2 * 38300, SDS-PAGE
80900
-
ultracentrifugation
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
dimer
-
2 * 38300, SDS-PAGE
homodimer
-
crystallization data
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystallized without ligand and in the presence of the inhibitors 2-oxo-1,2,3,6-tetrahydropyrimidine-4,6-dicarboxylate and 5-fluoroorotate, hanging drop vapour diffusion method with 15-20% (w/v) polyethylene glycol 3350, 0.1 M MES (pH 6.0-6.5), 75 mM MgCl2, 150 mM KCl (with ligand) or 20-25% polyethylene glycol 3350, 0.1 M Na HEPES (pH 7) and 0.2 M NaF (without ligand)
hanging drop vapour diffusion method with 15-20% polyethylene glycol 3350, 0.1 M MES, pH 6.0-6.5, 75 mM MgCl2, and 150 mM KCl
in complex with 5-fluoroorotate, hanging drop vapour diffusion method 14-16% PEG 3350, 0.1 M MES pH 6.25, 25 mM MgCl2, 0.2 M KCl and 30% sucrose
in the presence of enantiomerically pure L-dihydroorotate, refined at 1.9 Angstrom resolution
x-ray structure
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D250A
less than 1% of the activity possessed by the wild type enzyme
D250E
reduced catalytic activity compared to wild type
D250H
less than 1% of the activity possessed by the wild type enzyme
D250N
less than 1% of the activity possessed by the wild type enzyme
D250S
reduced catalytic activity compared to wild type
H254N
less than 1% of the activity possessed by the wild type enzyme
N44A
less than 1% of the activity possessed by the wild type enzyme
R20K
reduced catalytic activity compared to wild type
R20M
catalytically inactive
R20Q
less than 1% of the activity possessed by the wild type enzyme
T109A
decreased activity
T109G
negligible activity
T109S
T109V
strongly decreased activity
T110A
negligible activity
T110S
strongly decreased activity
T110V
negligible activity
C221S/C263S/C265S/C268S
-
this quadruple mutant is relatively stable to oxidation and has kinetic parameters consistent with reported values for wild type DHO
additional information
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80°C, Bis-Tris propane buffer, pH 7.0, 20% glycerol
4°C, under nitrogen, 100 mM Tris phosphate buffer, pH 7, metal free 10 mM N-carbamoyl-DL-aspartate, at least 2 months
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
chromatography, gel filtration, resource-Q anion exchange column
Mono Q column chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli strain X7014a
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Washabough, M.W.; Collins, K.D.
Dihydroorotase from Escherichia colii. Sulfhydryl group-metal ion interactions
J. Biol. Chem.
261
5920-5929
1986
Escherichia coli
Manually annotated by BRENDA team
Washabaugh, M.W.; Collins, K.D.
Dihydroorotase from Escherichia coli. Purification and characterization
J. Biol. Chem.
259
3293-3298
1984
Escherichia coli
Manually annotated by BRENDA team
Thoden, J.B.; Phillips, G.N., Jr.; Neal, T.M.; Raushel, F.M.; Holden, H.M.
Molecular structure of dihydroorotase: a paradigm for catalysis through the use of a binuclear metal center
Biochemistry
40
6989-6997
2001
Escherichia coli
Manually annotated by BRENDA team
Porter, T.N.; Li, Y.; Raushel, F.M.
Mechanism of the dihydroorotase reaction
Biochemistry
43
16285-16292
2004
Escherichia coli (P05020)
Manually annotated by BRENDA team
Li, Y.; Raushel, F.M.
Inhibitors designed for the active site of dihydroorotase
Bioorg. Chem.
33
470-483
2005
Escherichia coli
Manually annotated by BRENDA team
Lee, M.; Chan, C.W.; Mitchell Guss, J.; Christopherson, R.I.; Maher, M.J.
Dihydroorotase from Escherichia coli: loop movement and cooperativity between subunits
J. Mol. Biol.
348
523-533
2005
Arabidopsis thaliana (O04904), Escherichia coli (P05020), Escherichia coli, Mesocricetus auratus (P08955), Saccharomyces cerevisiae (P20051), Homo sapiens (P27708), Helicobacter pylori (P56465), Plasmodium falciparum (Q8IKA9)
Manually annotated by BRENDA team
Robles Lopez, S.M.; Hortua Triana, M.A.; Zimmermann, B.H.
Cloning and preliminary characterization of the dihydroorotase from Toxoplasma gondii
Mol. Biochem. Parasitol.
148
93-98
2006
Saccharomyces cerevisiae, Mycobacterium leprae, Plasmodium berghei (A0A509AX79), Arabidopsis thaliana (O04904), Trypanosoma cruzi (O76138), Escherichia coli (P05020), Mesocricetus auratus (P08955), Homo sapiens (P27708), Ustilago maydis (P31301), Toxoplasma gondii (Q8MXA5), Toxoplasma gondii
Manually annotated by BRENDA team
Lee, M.; Maher, M.J.; Guss, J.M.
Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-fluoroorotate: catalytic activity is reflected by the crystal form
Acta Crystallogr. Sect. F
F63
154-161
2007
Escherichia coli (P05020), Escherichia coli
Manually annotated by BRENDA team
Lee, M.; Maher, M.J.; Christopherson, R.I.; Guss, J.M.
Kinetic and structural analysis of mutant Escherichia coli dihydroorotases: a flexible loop stabilizes the transition state
Biochemistry
46
10538-10550
2007
Escherichia coli (P05020), Escherichia coli
Manually annotated by BRENDA team
Lee, M.; Chan, C.W.; Graham, S.C.; Christopherson, R.I.; Guss, J.M.; Maher, M.J.
Structures of ligand-free and inhibitor complexes of dihydroorotase from Escherichia coli: implications for loop movement in inhibitor design
J. Mol. Biol.
370
812-825
2007
Escherichia coli (P05020), Escherichia coli
Manually annotated by BRENDA team