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Literature summary extracted from

  • Lohkamp, B.; Voevodskaya, N.; Lindqvist, Y.; Dobritzsch, D.
    Insights into the mechanism of dihydropyrimidine dehydrogenase from site-directed mutagenesis targeting the active site loop and redox cofactor coordination (2010), Biochim. Biophys. Acta, 1804, 2198-2206.
    View publication on PubMed

Application

EC Number Application Comment Organism
1.3.1.2 drug development the enzyme is an adjunct target in cancer therapy since it rapidly breaks down the anti-cancer drug 5-fluorouracil and related compounds Sus scrofa

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.3.1.2 expression of wild-type and mutant enzymes in Escherichia coli strain Tuner(DE3) Sus scrofa

Protein Variants

EC Number Protein Variants Comment Organism
1.3.1.2 C126A site-directed mutagenesis, a potential [4Fe-4S]-cluster binding residue, the mutant shows slightly increased activity compared to the wild-type enzyme Sus scrofa
1.3.1.2 H673N site-directed mutagenesis, active site loop residue, the mutant shows reduced activity compared to the wild-type enzyme Sus scrofa
1.3.1.2 H673Q site-directed mutagenesis, active site loop residue, the mutant shows reduced activity compared to the wild-type enzyme Sus scrofa
1.3.1.2 Q156E site-directed mutagenesis, [4Fe-4S]-cluster binding residue, inactive mutant Sus scrofa
1.3.1.2 R235A site-directed mutagenesis, FAD binding residue, inactive mutant Sus scrofa
1.3.1.2 R235K site-directed mutagenesis, FAD binding residue, inactive mutant Sus scrofa
1.3.1.2 S670A site-directed mutagenesis, active site loop residue, the mutant shows reduced activity compared to the wild-type enzyme Sus scrofa

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.3.1.2 additional information
-
 additional information pH dependence and kinetics of recombinant wild-type and mutant enzymes, overview Sus scrofa
1.3.1.2 0.0011
-
 Uracil pH 7.5, 30°C, recombinant mutant C126A Sus scrofa
1.3.1.2 0.0012
-
 Uracil pH 7.5, 30°C, recombinant wild-type enzyme Sus scrofa
1.3.1.2 0.0042
-
 Uracil pH 7.5, 30°C, recombinant mutant H673Q Sus scrofa
1.3.1.2 0.0057
-
 NADPH pH 7.5, 30°C, recombinant wild-type enzyme Sus scrofa
1.3.1.2 0.0061
-
 NADPH pH 7.5, 30°C, recombinant mutant C126A Sus scrofa
1.3.1.2 0.0062
-
 NADPH pH 7.5, 30°C, recombinant mutant H673Q Sus scrofa
1.3.1.2 0.007
-
 Uracil pH 7.5, 30°C, recombinant mutant H673N Sus scrofa
1.3.1.2 0.0075
-
 NADPH pH 7.5, 30°C, recombinant mutant H673N Sus scrofa
1.3.1.2 0.0188
-
 NADPH pH 7.5, 30°C, recombinant mutant S670A Sus scrofa
1.3.1.2 0.0366
-
 Uracil pH 7.5, 30°C, recombinant mutant S670A Sus scrofa

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.3.1.2 Fe2+ iron-sulfur cluster binding, overview Sus scrofa

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.3.1.2 dihydrofluorouracil + NADP+ Sus scrofa dihydrofluorouracil is further metabolized to 2'-fluoro-beta-alanine 5-fluorouracil + NADPH + H+
-
 ?
1.3.1.2 dihydrothymine + NADP+ Sus scrofa
-
 thymine + NADPH + H+
-
 ?
1.3.1.2 dihydrouracil + NADP+ Sus scrofa
-
 uracil + NADPH + H+
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.3.1.2 Sus scrofa Q28943
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.3.1.2 recombinant wild-type and mutant enzymes from Escherichia coli by affinity chromatography and gel filtration Sus scrofa

Source Tissue

EC Number Source Tissue Comment Organism Textmining
1.3.1.2 liver
-
 Sus scrofa
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.3.1.2 dihydrofluorouracil + NADP+ dihydrofluorouracil is further metabolized to 2'-fluoro-beta-alanine Sus scrofa 5-fluorouracil + NADPH + H+
-
 ?
1.3.1.2 dihydrofluorouracil + NADP+ pyrimidine binding to this enzyme is accompanied by active site loop closure that positions a catalytically crucial cysteine 671 residue. The deprotonation of the loop residue H673 is required for active site closure, while S670 is important for substrate recognition. R235 is crucial for FAD binding Sus scrofa 5-fluorouracil + NADPH + H+ dihydrofluorouracil is further metabolized to 2'-fluoro-beta-alanine ?
1.3.1.2 dihydrothymine + NADP+
-
 Sus scrofa thymine + NADPH + H+
-
 ?
1.3.1.2 dihydrouracil + NADP+
-
 Sus scrofa uracil + NADPH + H+
-
 ?
1.3.1.2 additional information the first FeS cluster, with unusual coordination, cannot be reduced and displays no activity when Q156 is mutated to glutamate. The active site loop comprising residues 670-683 are observed in open and closed conformational states, overview Sus scrofa ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
1.3.1.2 dihydropyrimidine dehydrogenase
-
 Sus scrofa
1.3.1.2 DPD
-
 Sus scrofa

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.3.1.2 30
-
 assay at Sus scrofa

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.3.1.2 0.068
-
 Uracil pH 7.5, 30°C, recombinant mutant H673N Sus scrofa
1.3.1.2 0.117
-
 Uracil pH 7.5, 30°C, recombinant mutant H673Q Sus scrofa
1.3.1.2 0.177
-
 Uracil pH 7.5, 30°C, recombinant mutant S670A Sus scrofa
1.3.1.2 0.242
-
 Uracil pH 7.5, 30°C, recombinant wild-type enzyme Sus scrofa
1.3.1.2 0.256
-
 Uracil pH 7.5, 30°C, recombinant mutant C126A Sus scrofa

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.3.1.2 additional information
-
 pH dependence and kinetics of recombinant wild-type and mutant enzymes, overview Sus scrofa
1.3.1.2 7.5
-
 assay at Sus scrofa

Cofactor

EC Number Cofactor Comment Organism Structure
1.3.1.2 FAD absolutely required, R235 is crucial for FAD binding Sus scrofa
1.3.1.2 NADPH dependent on Sus scrofa

General Information

EC Number General Information Comment Organism
1.3.1.2 metabolism in mammals, the pyrimidines uracil and thymine are metabolised by a three-step reductive degradation pathway. Dihydropyrimidine dehydrogenase catalyses its first and rate-limiting step, reducing uracil and thymine to the corresponding 5,6-dihydropyrimidines in an NADPH-dependent reaction Sus scrofa
1.3.1.2 additional information the N-terminal half of DPD is a member of a family of FAD-containing NADPH oxidoreductases, which transfer electrons to an acceptor protein or domain through [4Fe-4S] clusters of low to very low potential Sus scrofa

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.3.1.2 5000
-
 Uracil pH 7.5, 30°C, recombinant mutant S670A Sus scrofa
1.3.1.2 9000
-
 NADPH pH 7.5, 30°C, recombinant mutant H673N Sus scrofa
1.3.1.2 9000
-
 NADPH pH 7.5, 30°C, recombinant mutant S670A Sus scrofa
1.3.1.2 10000
-
 Uracil pH 7.5, 30°C, recombinant mutant H673N Sus scrofa
1.3.1.2 19000
-
 NADPH pH 7.5, 30°C, recombinant mutant H673Q Sus scrofa
1.3.1.2 28000
-
 Uracil pH 7.5, 30°C, recombinant mutant H673Q Sus scrofa
1.3.1.2 42000
-
 NADPH pH 7.5, 30°C, recombinant mutant C126A Sus scrofa
1.3.1.2 42000
-
 NADPH pH 7.5, 30°C, recombinant wild-type enzyme Sus scrofa
1.3.1.2 208000
-
 Uracil pH 7.5, 30°C, recombinant wild-type enzyme Sus scrofa
1.3.1.2 238000
-
 Uracil pH 7.5, 30°C, recombinant mutant C126A Sus scrofa