Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 1.3.1.14 extracted from

  • Kahler, A.E.; Nielsen, F.S.; Switzer, R.L.
    Biochemical characterization of the heteromeric Bacillus subtilis dihydroorotate dehydrogenase and its isolated subunits (1999), Arch. Biochem. Biophys., 371, 191-201.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
-
Bacillus subtilis
expression in Escherichia coliBL21 DE3, expression of pyrDI alone and coexpression with pyrDII Bacillus subtilis

Protein Variants

Protein Variants Comment 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 Comment Organism Structure
Orotate
-
Bacillus subtilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment 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] Comment 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 Bacillus subtilis
114000
-
molecular mass of the holoenzyme, calculated mass 122400 Da Bacillus subtilis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(S)-dihydroorotate + acceptor Bacillus subtilis fourth step in UMP-biosynthesis orotate + reduced acceptor
-
?

Organism

Organism UniProt Comment Textmining
Bacillus subtilis
-
-
-
Mus musculus
-
Ehrlich ascites tumor cells
-

Purification (Commentary)

Purification (Comment) Organism
anion-exchange, gel filtration Bacillus subtilis

Reaction

Reaction Comment Organism Reaction ID
(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)

Renatured (Comment) Organism
activity can be recovered by mixing the purified subunits Bacillus subtilis
PyrDII-containing inclusion bodies are denatured and refolded through dialysis into buffer Bacillus subtilis

Storage Stability

Storage Stability Organism
4Ā°C, 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 Comment Substrates Organism Products Comment (Products) Rev. Reac.
(S)-dihydroorotate + acceptor different specific activities with potassium ferricyanide, O2, fumarate and NAD+ as electron acceptors for PyrDI and the holoenzyme Bacillus subtilis orotate + reduced acceptor
-
?
(S)-dihydroorotate + acceptor fourth step in UMP-biosynthesis Bacillus subtilis orotate + reduced acceptor
-
?
dihydroorotate + acceptor acceptor: NAD+ Bacillus subtilis orotate + reduced acceptor
-
?
dihydroorotate + acceptor acceptor: menadione Bacillus subtilis orotate + reduced acceptor
-
?
dihydroorotate + acceptor activity measurement in permeabilized Ehrlich ascites tumor cells, acceptor: nitroblue tetrazolium Mus musculus orotate + reduced acceptor
-
?

Subunits

Subunits Comment Organism
tetramer heterotetramer 2 * 33094 and 2 * 28099 predicted from seqeuence, 2 * 38000 and 2 * 31000, SDS-PAGE Bacillus subtilis
tetramer 2 * 33000 + 2 * 28000 Bacillus subtilis

Cofactor

Cofactor Comment Organism Structure
FAD 1 mol PyrDII binds 1 mol FAD and 1 mol [2Fe-2S] Bacillus subtilis
FAD 1 mol/mol of subunit PyrDII 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 Comment Organism Structure
0.0206
-
Orotate holoenzyme Bacillus subtilis
1
-
Orotate PyrDI Bacillus subtilis