BRENDA - Enzyme Database
show all sequences of 1.3.1.14

Biochemical characterization of the heteromeric Bacillus subtilis dihydroorotate dehydrogenase and its isolated subunits

Kahler, A.E.; Nielsen, F.S.; Switzer, R.L.; Arch. Biochem. Biophys. 371, 191-201 (1999)

Data extracted from this reference:

Cloned(Commentary)
Commentary
Organism
; expression in Escherichia coliBL21 DE3, expression of pyrDI alone and coexpression with pyrDII
Bacillus subtilis
Engineering
Amino acid exchange
Commentary
Organism
C220A
pyrDII mutant, conserved, essential for activity
Bacillus subtilis
C225A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
C228A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
C230A
pyrDII mutant nonconserved
Bacillus subtilis
C243A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
Inhibitors
Inhibitors
Commentary
Organism
Structure
Orotate
-
Bacillus subtilis
KM Value [mM]
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.027
-
S-dihydroorotate
holoenzyme with 2,6-dichlorophenolindophenol
Bacillus subtilis
0.0317
-
S-dihydroorotate
holoenzyme with menadione
Bacillus subtilis
0.5
1
S-dihydroorotate
PyrDI
Bacillus subtilis
Molecular Weight [Da]
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
28000
-
2 * 33000 + 2 * 28000
Bacillus subtilis
28099
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
31000
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
33000
-
2 * 33000 + 2 * 28000
Bacillus subtilis
33094
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
38000
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
85000
-
native molecular mass of PyrDI
Bacillus subtilis
114000
-
gel filtration; molecular mass of the holoenzyme, calculated mass 122400 Da
Bacillus subtilis
Natural Substrates/ Products (Substrates)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
(S)-dihydroorotate + acceptor
Bacillus subtilis
fourth step in UMP-biosynthesis
orotate + reduced acceptor
-
Bacillus subtilis
?
Organism
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
Bacillus subtilis
-
-
-
Mus musculus
-
Ehrlich ascites tumor cells
-
Purification (Commentary)
Commentary
Organism
anion-exchange, gel filtration
Bacillus subtilis
Reaction
Reaction
Commentary
Organism
(S)-dihydroorotate + NAD+ = orotate + NADH + H+
PyrDI contains the dihydroorotate-binding site, but PyrDII is required for full activity in vivo. Holoenzyme joins a NAD+-reductase activity
Bacillus subtilis
Renatured (Commentary)
Commentary
Organism
activity can be recovered by mixing the purified subunits; PyrDII-containing inclusion bodies are denatured and refolded through dialysis into buffer
Bacillus subtilis
Storage Stability
Storage Stability
Organism
4C, holoenzyme, stable for many days
Bacillus subtilis
on ice at room temperature, PyrDI, gradually loses activity over a period of hours
Bacillus subtilis
room temperature, holoenzyme, stable for many hours
Bacillus subtilis
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
(S)-dihydroorotate + acceptor
different specific activities with potassium ferricyanide, O2, fumarate and NAD+ as electron acceptors for PyrDI and the holoenzyme
390928
Bacillus subtilis
orotate + reduced acceptor
-
390928
Bacillus subtilis
?
(S)-dihydroorotate + acceptor
fourth step in UMP-biosynthesis
390928
Bacillus subtilis
orotate + reduced acceptor
-
390928
Bacillus subtilis
?
dihydroorotate + acceptor
acceptor: NAD+
390928
Bacillus subtilis
orotate + reduced acceptor
-
-
-
?
dihydroorotate + acceptor
acceptor: menadione
390928
Bacillus subtilis
orotate + reduced acceptor
-
-
-
?
dihydroorotate + acceptor
activity measurement in permeabilized Ehrlich ascites tumor cells, acceptor: nitroblue tetrazolium
390928
Mus musculus
orotate + reduced acceptor
-
-
-
?
Subunits
Subunits
Commentary
Organism
tetramer
2 * 33000 + 2 * 28000; heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
Cofactor
Cofactor
Commentary
Organism
Structure
FAD
1 mol/mol of subunit PyrDII; 1 mol PyrDII binds 1 mol FAD and 1 mol [2Fe-2S]
Bacillus subtilis
FMN
PyrDI is an FMN-containing iron-sulfur flavoprotein, 1 FMN molecule per PyrDI molecule
Bacillus subtilis
NAD+
for holoenzyme
Bacillus subtilis
Ki Value [mM]
Ki Value [mM]
Ki Value maximum [mM]
Inhibitor
Commentary
Organism
Structure
0.0206
-
Orotate
holoenzyme
Bacillus subtilis
1
-
Orotate
PyrDI
Bacillus subtilis
Cloned(Commentary) (protein specific)
Commentary
Organism
; expression in Escherichia coliBL21 DE3, expression of pyrDI alone and coexpression with pyrDII
Bacillus subtilis
Cofactor (protein specific)
Cofactor
Commentary
Organism
Structure
FAD
1 mol/mol of subunit PyrDII; 1 mol PyrDII binds 1 mol FAD and 1 mol [2Fe-2S]
Bacillus subtilis
FMN
PyrDI is an FMN-containing iron-sulfur flavoprotein, 1 FMN molecule per PyrDI molecule
Bacillus subtilis
NAD+
for holoenzyme
Bacillus subtilis
Engineering (protein specific)
Amino acid exchange
Commentary
Organism
C220A
pyrDII mutant, conserved, essential for activity
Bacillus subtilis
C225A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
C228A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
C230A
pyrDII mutant nonconserved
Bacillus subtilis
C243A
pyrDII mutant conserved, essential for activity
Bacillus subtilis
Inhibitors (protein specific)
Inhibitors
Commentary
Organism
Structure
Orotate
-
Bacillus subtilis
Ki Value [mM] (protein specific)
Ki Value [mM]
Ki Value maximum [mM]
Inhibitor
Commentary
Organism
Structure
0.0206
-
Orotate
holoenzyme
Bacillus subtilis
1
-
Orotate
PyrDI
Bacillus subtilis
KM Value [mM] (protein specific)
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
0.027
-
S-dihydroorotate
holoenzyme with 2,6-dichlorophenolindophenol
Bacillus subtilis
0.0317
-
S-dihydroorotate
holoenzyme with menadione
Bacillus subtilis
0.5
1
S-dihydroorotate
PyrDI
Bacillus subtilis
Molecular Weight [Da] (protein specific)
Molecular Weight [Da]
Molecular Weight Maximum [Da]
Commentary
Organism
28000
-
2 * 33000 + 2 * 28000
Bacillus subtilis
28099
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
31000
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
33000
-
2 * 33000 + 2 * 28000
Bacillus subtilis
33094
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
38000
-
heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
85000
-
native molecular mass of PyrDI
Bacillus subtilis
114000
-
gel filtration; molecular mass of the holoenzyme, calculated mass 122400 Da
Bacillus subtilis
Natural Substrates/ Products (Substrates) (protein specific)
Natural Substrates
Organism
Commentary (Nat. Sub.)
Natural Products
Commentary (Nat. Pro.)
Organism (Nat. Pro.)
Reversibility
(S)-dihydroorotate + acceptor
Bacillus subtilis
fourth step in UMP-biosynthesis
orotate + reduced acceptor
-
Bacillus subtilis
?
Purification (Commentary) (protein specific)
Commentary
Organism
anion-exchange, gel filtration
Bacillus subtilis
Renatured (Commentary) (protein specific)
Commentary
Organism
activity can be recovered by mixing the purified subunits; PyrDII-containing inclusion bodies are denatured and refolded through dialysis into buffer
Bacillus subtilis
Storage Stability (protein specific)
Storage Stability
Organism
4C, holoenzyme, stable for many days
Bacillus subtilis
on ice at room temperature, PyrDI, gradually loses activity over a period of hours
Bacillus subtilis
room temperature, holoenzyme, stable for many hours
Bacillus subtilis
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
(S)-dihydroorotate + acceptor
different specific activities with potassium ferricyanide, O2, fumarate and NAD+ as electron acceptors for PyrDI and the holoenzyme
390928
Bacillus subtilis
orotate + reduced acceptor
-
390928
Bacillus subtilis
?
(S)-dihydroorotate + acceptor
fourth step in UMP-biosynthesis
390928
Bacillus subtilis
orotate + reduced acceptor
-
390928
Bacillus subtilis
?
dihydroorotate + acceptor
acceptor: NAD+
390928
Bacillus subtilis
orotate + reduced acceptor
-
-
-
?
dihydroorotate + acceptor
acceptor: menadione
390928
Bacillus subtilis
orotate + reduced acceptor
-
-
-
?
dihydroorotate + acceptor
activity measurement in permeabilized Ehrlich ascites tumor cells, acceptor: nitroblue tetrazolium
390928
Mus musculus
orotate + reduced acceptor
-
-
-
?
Subunits (protein specific)
Subunits
Commentary
Organism
tetramer
2 * 33000 + 2 * 28000; heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE
Bacillus subtilis
Other publictions for EC 1.3.1.14
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Temperature Optimum [C]
Temperature Range [C]
Temperature Stability [C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [C] (protein specific)
Temperature Range [C] (protein specific)
Temperature Stability [C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
701809
Kawasaki
b-Type dihydroorotate dehydrog ...
Bifidobacterium bifidum
Appl. Environ. Microbiol.
75
629-636
2009
-
-
-
-
-
-
4
-
-
-
1
-
-
4
-
-
1
-
-
-
-
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-
1
-
-
-
-
2
1
-
2
-
-
-
-
-
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2
-
-
-
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4
-
-
-
-
1
-
-
-
-
1
-
-
-
-
-
1
-
-
-
-
2
1
-
-
-
-
-
-
-
-
674586
Combe
Lys-D48 is required for charge ...
Lactococcus lactis
J. Biol. Chem.
281
17977-17988
2006
-
-
-
-
4
-
-
9
-
-
-
-
-
1
-
-
-
-
-
-
-
-
1
-
-
-
-
9
-
-
-
3
-
-
-
-
-
-
3
-
4
-
-
-
-
9
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
9
-
-
-
-
-
-
-
-
-
-
654759
Mohsen
Thermodynamic basis of electro ...
Lactococcus lactis
Biochemistry
43
6498-6510
2004
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
1
-
-
-
-
1
-
-
-
-
-
-
-
-
3
-
-
-
-
-
-
3
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390568
Argyrou
Dihydroorotate dehydrogenase f ...
Bacillus subtilis, Enterococcus faecalis, Faecalicatena orotica, Lactococcus lactis
Biochemistry
39
10373-10384
2000
6
-
2
-
-
-
-
2
4
-
3
3
-
4
-
-
1
3
-
-
1
1
13
3
1
-
-
3
-
1
-
12
-
-
-
6
-
2
12
-
-
-
-
-
-
2
4
-
3
3
-
-
-
1
-
-
1
1
13
3
1
-
-
3
-
1
-
-
-
-
-
-
-
-
390569
Rowland
Structure of dihydroorotate de ...
Bacillus subtilis, Enterococcus faecalis, Faecalicatena orotica, Lactococcus lactis
Structure
8
1227-1238
2000
-
5
-
1
-
-
-
-
4
-
-
3
-
4
-
-
1
1
-
-
-
-
11
3
-
-
-
-
-
-
-
12
-
-
-
-
5
-
12
1
-
-
-
-
-
-
4
-
-
3
-
-
-
1
-
-
-
-
11
3
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390928
Kahler
Biochemical characterization o ...
Bacillus subtilis, Mus musculus
Arch. Biochem. Biophys.
371
191-201
1999
-
-
1
-
5
-
1
3
-
-
8
1
-
2
-
-
1
1
1
-
-
3
5
1
-
-
-
-
-
-
-
3
2
-
-
-
-
1
3
-
5
-
-
1
2
3
-
-
8
1
-
-
-
1
1
-
-
3
5
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
391244
Marcinkeviciene
Dihydroorotate dehydrogenase B ...
Enterococcus faecalis
Biochemistry
38
13129-13137
1999
-
-
-
-
-
-
-
-
-
-
3
-
-
3
-
-
1
1
-
-
-
-
2
1
-
-
-
-
-
-
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
3
-
-
-
-
1
-
-
-
-
2
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390917
Andersen
Two different dihydroorotate d ...
Lactococcus lactis
J. Bacteriol.
176
3975-3982
1994
-
-
1
-
-
-
-
-
-
-
1
-
-
2
-
-
-
-
-
-
4
-
4
-
-
-
-
-
-
-
-
1
-
-
-
-
-
2
1
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
4
-
4
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390565
Buntain
-
Latent Inhibitors. Part 4. Irr ...
Faecalicatena orotica
J. Chem. Soc. Perkin Trans. I
1988
3175-3182
1988
-
2
-
-
-
1
12
-
-
-
-
1
-
1
-
-
-
-
-
-
-
-
2
-
1
-
-
-
1
-
-
1
8
-
-
-
2
-
1
-
-
1
-
12
8
-
-
-
-
1
-
-
-
-
-
-
-
-
2
-
1
-
-
-
1
-
-
-
-
-
-
-
-
-
390564
Friedmann
-
Bestimmung mit Dihydroorotat-D ...
Faecalicatena orotica
Methods Enzym. Anal. , 3rd Ed. (Bergmeyer, H. U. , ed. )
2
2010-2014
1984
1
-
-
-
-
-
4
2
-
-
-
-
-
1
-
-
1
-
-
-
-
1
3
-
-
-
-
1
1
-
-
4
1
-
-
1
-
-
4
-
-
-
-
4
1
2
-
-
-
-
-
-
-
1
-
-
-
1
3
-
-
-
-
1
1
-
-
-
-
-
-
-
-
-
390563
Blattmann
Stereospecificity of the dihyd ...
Faecalicatena orotica
Eur. J. Biochem.
30
130-137
1972
-
-
-
-
-
-
-
-
-
-
-
-
-
1
-
-
-
1
-
-
1
1
2
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
1
1
2
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390567
Nelson
Preparation of bovine xanthine ...
Faecalicatena orotica
J. Biol. Chem.
243
5368-5373
1968
-
-
-
-
-
-
-
-
-
-
1
-
-
1
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
2
-
-
-
-
-
-
2
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
-
-
-
-
390566
Friedmann
Crystalline dihydroorotic dehy ...
Faecalicatena orotica
J. Biol. Chem.
235
1526-1530
1960
1
-
-
1
-
-
5
-
-
-
1
-
-
1
-
-
1
-
-
-
1
1
2
-
2
-
-
1
1
-
-
4
-
-
-
1
-
-
4
1
-
-
-
5
-
-
-
-
1
-
-
-
-
1
-
-
1
1
2
-
2
-
-
1
1
-
-
-
-
-
-
-
-
-
390561
Friedmann
Purification and properties of ...
Faecalicatena orotica
J. Biol. Chem.
233
1398-1406
1958
3
-
-
-
-
-
5
2
-
-
-
1
-
1
-
-
1
-
-
-
1
-
5
-
-
-
-
1
-
-
-
3
-
-
-
3
-
-
3
-
-
-
-
5
-
2
-
-
-
1
-
-
-
1
-
-
1
-
5
-
-
-
-
1
-
-
-
-
-
-
-
-
-
-
390562
Lieberman
Enzymic synthesis and breakdow ...
Faecalicatena orotica
Biochim. Biophys. Acta
12
223-234
1953
1
-
-
-
-
-
1
1
-
-
-
1
-
1
-
-
1
-
-
-
1
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3
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1
1
2
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1
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1
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1
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1
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