This Class 2 dihydroorotate dehydrogenase enzyme contains FMN . The enzyme is found in eukaryotes in the mitochondrial membrane, in cyanobacteria, and in some Gram-negative and Gram-positive bacteria associated with the cytoplasmic membrane [2,5,6]. The reaction is the only redox reaction in the de-novo biosynthesis of pyrimidine nucleotides [2,4]. The best quinone electron acceptors for the enzyme from bovine liver are ubiquinone-6 and ubiquinone-7, although simple quinones, such as benzoquinone, can also act as acceptor at lower rates . Methyl-, ethyl-, tert-butyl and benzyl (S)-dihydroorotates are also substrates, but methyl esters of (S)-1-methyl and (S)-3-methyl and (S)-1,3-dimethyldihydroorotates are not . Class 1 dihydroorotate dehydrogenases use either fumarate (EC 1.3.98.1), NAD+ (EC 1.3.1.14) or NADP+ (EC 1.3.1.15) as electron acceptor.
(S)-dihydroorotate + a quinone = orotate + a quinol
existence of 3 families differing in their selectivity for oxidizing substrates: 1A are soluble, containing FMN and use fumarate, 1B are soluble, one subunit contains an iron-sulfur center, FAD and reduces NAD+, family 2 enzymes are membrane-bound, contain FMN and are oxidized by ubiquinone. Mechanism consists of 3 reaction phases. Different binding-mechanisms for enzyme the reaction-steps are suggested
(S)-dihydroorotate + a quinone = orotate + a quinol
different binding sites for dihydroorotate and the electron acceptor, two-site ping-pong mechanism. Cleavage site at R182 is conserved between the two major families of dihydroorotate dehydrogenases, it is positioned in a loop, which is crucial for catalysis but irrelevant for protein stability
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SYSTEMATIC NAME
IUBMB Comments
(S)-dihydroorotate:quinone oxidoreductase
This Class 2 dihydroorotate dehydrogenase enzyme contains FMN [4]. The enzyme is found in eukaryotes in the mitochondrial membrane, in cyanobacteria, and in some Gram-negative and Gram-positive bacteria associated with the cytoplasmic membrane [2,5,6]. The reaction is the only redox reaction in the de-novo biosynthesis of pyrimidine nucleotides [2,4]. The best quinone electron acceptors for the enzyme from bovine liver are ubiquinone-6 and ubiquinone-7, although simple quinones, such as benzoquinone, can also act as acceptor at lower rates [2]. Methyl-, ethyl-, tert-butyl and benzyl (S)-dihydroorotates are also substrates, but methyl esters of (S)-1-methyl and (S)-3-methyl and (S)-1,3-dimethyldihydroorotates are not [2]. Class 1 dihydroorotate dehydrogenases use either fumarate (EC 1.3.98.1), NAD+ (EC 1.3.1.14) or NADP+ (EC 1.3.1.15) as electron acceptor.
electron spin resonance spectra show that the N-terminal binds to membranes and experiences a somewhat high flexibility that could be related to the role of this region as a molecular lid controlling the entrance of the enzyme's active site and thus allowing the enzyme to give access to quinones that are dispersed in the membrane and that are necessary for the catalysis
mutant incorporates into 1-dipalmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/Triton X-100 mixed vesicles and expected to be located right in the core of the more hydrophobic region of the model membrane. Mutated amino acids are either in a strongly immobilized regime or subjected to a fast motion
mutant incorporates into 1-dipalmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/Triton X-100 mixed vesicles. Mutated residues experience a high degree of freedom that is compatible with their location in the beginning of the protein chain. Mutated amino acids are either in a strongly immobilized regime or subjected to a fast motion
mutant incorporates into 1-dipalmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/Triton X-100 mixed vesicles and expected to be located right in the core of the more hydrophobic region of the model membrane. Mutated amino acids are either in a strongly immobilized regime or subjected to a fast motion
mutant incorporates into 1-dipalmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/Triton X-100 mixed vesicles. Mutated residues experience a high degree of freedom that is compatible with their location in the beginning of the protein chain
overexpression of Escherichia coli dihyroorotate dehyrogenase in same strain, partially deleted for the chromosomal pyrD gene, clone selection followed by ampicillin and by complementation of the pyrimidine requirement
kinetic isotope effects on flavin reduction in anaerobic stopped-flow experiments, are about 3fold for DHO labeled at the 5-position, about 4fold for DHO labeled at the 6-position, and about 6-7fold for DHO labeled at both the 5- and 6-positions, at a pH value above the pKa controlling reduction, no isotope effect was observed for DHO deuterated at the 5-position, which is consistent with a stepwise reaction, above the kinetic pKa, the deprotonation of C5 is fast enough that it does not contribute to the observed rate constant and, therefore, is not isotopically sensitive
The activity of Escherichia coli dihydroorotate dehydrogenase is dependent on a conserved loop identified by sequence homology, mutagenesis, and limited proteolysis