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Literature summary extracted from

  • Argyrou, A.; Washabaugh, M.W.; Pickart, C.M.
    Dihydroorotate dehydrogenase from Clostridium oroticum is a class 1B enzyme and utilizes a concerted mechanism of catalysis (2000), Biochemistry, 39, 10373-10384.
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
1.3.1.14 2-mercaptoethanol activates Faecalicatena orotica
1.3.1.14 cysteine L-cysteine activates strongly, D-cysteine is also an activator, 1.8 mM L-cysteine is required at pH 8 for half-maximal activity, 10 mM for near-maximal activity Faecalicatena orotica
1.3.1.14 dithiothreitol activates Faecalicatena orotica
1.3.1.14 glutathione activates Faecalicatena orotica
1.3.1.14 Mercaptoacetic acid activates Faecalicatena orotica
1.3.1.14 additional information thiol activation involves reduction of side chain of a cysteine residue corresponding to Cys-130, which functions as a general base Faecalicatena orotica
1.3.1.14 additional information not activated by L-serine or L-alanine Faecalicatena orotica

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.3.1.14 genes encoding the two alpha and beta polypeptides, expression in Escherichia coli Bacillus subtilis
1.3.1.14 genes encoding the two alpha and beta polypeptides, expression in Escherichia coli Lactococcus lactis

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.3.1.14 0.09
-
 L-dihydroorotate pH 8, 25°C Faecalicatena orotica
1.3.1.14 0.13
-
 NAD+ pH 8, 25°C Faecalicatena orotica

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
1.3.1.14 cytosol
-
 Bacillus subtilis 5829
-
1.3.1.14 cytosol
-
 Lactococcus lactis 5829
-
1.3.1.14 cytosol
-
 Enterococcus faecalis 5829
-
1.3.1.14 cytosol
-
 Faecalicatena orotica 5829
-

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.3.1.14 28000
-
 alpha2,beta2, 2 * 32000 + 2 * 28000, SDS-PAGE Faecalicatena orotica
1.3.1.14 32000
-
 alpha2,beta2, 2 * 32000 + 2 * 28000, SDS-PAGE Faecalicatena orotica
1.3.1.14 120000
-
 analytical gel filtration Faecalicatena orotica

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.3.1.14 dihydroorotate + NAD+ Lactococcus lactis fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway orotate + NADH + H+
-
 r
1.3.1.14 dihydroorotate + NAD+ Enterococcus faecalis fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway orotate + NADH + H+
-
 r
1.3.1.14 dihydroorotate + NAD+ Faecalicatena orotica fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway orotate + NADH + H+
-
 r

Organism

EC Number Organism UniProt Comment Textmining
1.3.1.14 Bacillus subtilis
-
-
-
1.3.1.14 Enterococcus faecalis
-
-
-
1.3.1.14 Faecalicatena orotica
-
 identical with Clostridium oroticum
-
1.3.1.14 Lactococcus lactis
-
 family IB enzyme, also contains family IA enzyme
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.3.1.14
-
 Faecalicatena orotica

Reaction

EC Number Reaction Comment Organism Reaction ID
1.3.1.14 (S)-dihydroorotate + NAD+ = orotate + NADH + H+ electron transfer mechanism Lactococcus lactis
a
1.3.1.14 (S)-dihydroorotate + NAD+ = orotate + NADH + H+ ping-pong steady-state kinetic mechanism Enterococcus faecalis
a
1.3.1.14 (S)-dihydroorotate + NAD+ = orotate + NADH + H+ ping-pong steady-state kinetic mechanism Faecalicatena orotica
a

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.3.1.14 additional information
-
-
 Faecalicatena orotica

Storage Stability

EC Number Storage Stability Organism
1.3.1.14 -20°C, 0.27 M sodium phosphate, pH 6.3, over 6 months, less than 15% loss of activity Faecalicatena orotica

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.3.1.14 (S)-dihydroorotate + NAD+ equilibrium favours direction of orotate reduction Faecalicatena orotica orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ equilibrium favours direction of orotate reduction Faecalicatena orotica orotate + NADH + H+
-
 r
1.3.1.14 (S)-dihydroorotate + NAD+ mechanism of dehydrogenation, mechanism of electron transfer: L-dihydroorotate transfers a pair of electrons to FMN via iron-sulfur cluster via FAD to NAD+ Lactococcus lactis orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ mechanism of dehydrogenation, mechanism of electron transfer: L-dihydroorotate transfers a pair of electrons to FMN via iron-sulfur cluster via FAD to NAD+ Lactococcus lactis orotate + NADH + H+
-
 r
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Bacillus subtilis orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Lactococcus lactis orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Lactococcus lactis orotate + NADH + H+
-
 r
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Enterococcus faecalis orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Faecalicatena orotica orotate + NADH + H+
-
 ?
1.3.1.14 (S)-dihydroorotate + NAD+ NAD+ as ultimate electron acceptor Faecalicatena orotica orotate + NADH + H+
-
 r
1.3.1.14 dihydroorotate + NAD+ fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway Lactococcus lactis orotate + NADH + H+
-
 r
1.3.1.14 dihydroorotate + NAD+ fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway Enterococcus faecalis orotate + NADH + H+
-
 r
1.3.1.14 dihydroorotate + NAD+ fourth step and sole redox reaction in the pyrimidine de novo biosynthetic pathway Faecalicatena orotica orotate + NADH + H+
-
 r

Subunits

EC Number Subunits Comment Organism
1.3.1.14 heterotetramer alpha2,beta2 Bacillus subtilis
1.3.1.14 heterotetramer alpha2,beta2, 2 * 32000 + 2 * 28000, SDS-PAGE Faecalicatena orotica
1.3.1.14 heterotetramer alpha2,beta2, family IB enzyme Lactococcus lactis
1.3.1.14 heterotetramer alpha subunit is named PyrDB, beta subunit is named PyrK Bacillus subtilis
1.3.1.14 heterotetramer alpha subunit is named PyrDB, beta subunit is named PyrK Lactococcus lactis

Synonyms

EC Number Synonyms Comment Organism
1.3.1.14 More family IB dihydroorotate dehydrogenase Lactococcus lactis
1.3.1.14 More family IB dihydroorotate dehydrogenase Faecalicatena orotica

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.3.1.14 25
-
 assay at Faecalicatena orotica

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.3.1.14 3.97
-
 Orotate formation of L-dihydroorotate, in absence of NAD+ or NADH, pH 8, 10 mM cysteine Faecalicatena orotica
1.3.1.14 22
-
 L-dihydroorotate formation of orotate, in absence of NAD+ or NADH, pH 8, 10 mM cysteine Faecalicatena orotica
1.3.1.14 45
-
 L-dihydroorotate in presence of NAD+ Faecalicatena orotica

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.3.1.14 6.4 8.9 below and above the turnover number decreases Faecalicatena orotica

Cofactor

EC Number Cofactor Comment Organism Structure
1.3.1.14 4Fe-4S-center iron-sulfur cluster resides on the beta subunit Lactococcus lactis
1.3.1.14 4Fe-4S-center cofactor content: 4 non-heme iron and 4 labile sulfur atoms per heterotetramer Bacillus subtilis
1.3.1.14 4Fe-4S-center cofactor content: 4 non-heme iron and 4 labile sulfur atoms per heterotetramer Lactococcus lactis
1.3.1.14 4Fe-4S-center cofactor content: 4 non-heme iron and 4 labile sulfur atoms per heterotetramer Enterococcus faecalis
1.3.1.14 4Fe-4S-center cofactor content: 4 non-heme iron and 4 labile sulfur atoms per heterotetramer Faecalicatena orotica
1.3.1.14 FAD FAD is located on the beta subunit Lactococcus lactis
1.3.1.14 FAD two molecules FAD per heterotetramer Bacillus subtilis
1.3.1.14 FAD two molecules FAD per heterotetramer Lactococcus lactis
1.3.1.14 FAD two molecules FAD per heterotetramer Enterococcus faecalis
1.3.1.14 FAD two molecules FAD per heterotetramer Faecalicatena orotica
1.3.1.14 FAD flavoprotein, ratio FAD to FMN 1:1 Faecalicatena orotica
1.3.1.14 FMN flavoprotein, ratio FAD to FMN 1:1 Faecalicatena orotica
1.3.1.14 FMN two molecules of FMN per heterotetramer Bacillus subtilis
1.3.1.14 FMN two molecules of FMN per heterotetramer Lactococcus lactis
1.3.1.14 FMN two molecules of FMN per heterotetramer Enterococcus faecalis
1.3.1.14 FMN two molecules of FMN per heterotetramer Faecalicatena orotica
1.3.1.14 FMN FMN is located on the alpha subunit Lactococcus lactis