Disease relevance of DHODH

• Their in vivo biological activity determined in rat and mouse delayed type hypersensitivity has been found to correlate well with their in vitro DHODH potency [1].
• As rapidly proliferating human T cells have an exceptional requirement for de novo pyrimidine biosynthesis, small molecule DHODH inhibitors constitute an attractive therapeutic approach to autoimmune diseases, immunosuppression, and cancer [2].
• Based on our recent functional expression of the full-length rat and human dihydroorotate dehydrogenase, here we expressed N-terminal-truncated C-terminal-histidine-tagged constructs of the mouse, rat and human enzymes in Escherichia coli [3].
• Functional expression of a fragment of human dihydroorotate dehydrogenase by means of the baculovirus expression vector system, and kinetic investigation of the purified recombinant enzyme [4].
• We report the discovery of a class of pyrazole-based compounds that are potent inhibitors of the dihydroorotate dehydrogenase of Helicobacter pylori but that do not inhibit the cognate enzymes from Gram-positive bacteria or humans [5].

High impact information on DHODH

• Single-molecule kinetics reveals signatures of half-sites reactivity in dihydroorotate dehydrogenase A catalysis [6].
• Brequinar sodium is a 4-quinolinecarboxylic acid analogue that inhibits dihydroorotate dehydrogenase and subsequent de novo pyrimidine biosynthesis [7].
• CAD was not concentrated near the mitochondria where the next enzyme of the de novo pathway, dihydroorotate dehydrogenase, is localized, which indicates that the intermediate dihydroorotate is not channeled, but rather dissociates from CAD and diffuses through the bulk cellular fluid [8].
• The results showed that NSC 368390 was a potent inhibitor of dihydroorotate dehydrogenase, the fourth enzyme in the pathway; thus, this study demonstrates that NSC 368390 exerts its tumoricidal effect by inhibiting a step in de novo pyrimidine biosynthesis resulting in the depletion of critical precursors for RNA and DNA synthesis [9].
• The structural basis for the species selective enzyme inhibition is explained by the variable amino acid sequence in this binding site, making DHODH a particularly strong candidate for the development of new anti-malarial compounds [10].

Chemical compound and disease context of DHODH

• Dihydroorotate dehydrogenase and succinate dehydrogenase activity in Ehrlich ascites tumor cells grown in suspension culture were quantified by application of nitroblue tetrazolium or cyanotolyl tetrazolium and subsequent extraction of the insoluble formazans with organic solvents [11].

Biological context of DHODH

• Import of rat DHODH into isolated yeast mitochondria required inner membrane potential and was at least partially dependent upon matrix ATP, indicating that its localization uses the well described import machinery of the mitochondrial inner membrane [12].
• The active site of human DHODH is generally similar to that of the previously reported bacterial active site [2].
• Mammalian dihydroorotate dehydrogenase, the fourth enzyme of pyrimidine de novo synthesis is an integral protein of the inner mitochondrial membrane that faces the intermembrane space and is functionally connected to the respiratory chain via ubiquinone [3].
• Structure-activity relationship studies explained that potent inhibitors of animal DHODH do not significantly affect the plant enzyme [13].
• Discovery of a novel series of DHODH inhibitors by a docking procedure and QSAR refinement [14].

Anatomical context of DHODH

• Thus the inhibition of DHODH by A77 1726 prevents lymphocytes from accumulating sufficient pyrimidines to support DNA synthesis [15].
• Deletion of the cationic portion of this N-terminal sequence from the rat DHODH protein blocked its import into isolated yeast mitochondria, whereas deletion of the adjacent hydrophobic segment resulted in import of the protein into the matrix [12].
• Analysis of fractionated rat liver mitochondria revealed that DHODH is an integral membrane protein exposed to the intermembrane space [12].
• Rabbit anti-DHODH IgG, which was generated using an immunization protocol with truncated recombinant human DHODH protein and purified by an immunosorbent method, was used for immunocytochemical detection and localization of this enzyme in ejaculated human spermatozoa [16].
• The MNAs inhibit both T-cell and B-cell function by blocking de novo pyrimidine synthesis, through blockade of the pivotal mitochondrial enzyme dihyroorotic acid dehydrogenase (DHODH), and the inhibition of tyrosine kinase activity [17].

Associations of DHODH with chemical compounds

• In the cell DHODH catalyzes the rate-limiting step of the de novo pyrimidine biosynthesis [18].
• The active metabolite (2) of the novel immunosuppressive agent leflunomide (1) has been shown to inhibit the enzyme dihydroorotate dehydrogenase (DHODH) [1].
• The initial set of phases was obtained using multiwavelength anomalous diffraction phasing with selenomethionine-containing DHODH [2].
• DHODH inhibition occurs at lower concentrations of A77 1726 than that of tyrosine kinases and is currently considered the major mode of action [19].
• Whereas the presence of the N-terminus of dihydroorotate dehydrogenase was essentially irrelevant for the efficacy of the malononitrilamides A77-1726, MNA715 and MNA279 with the rat and human enzyme, the N-termini were found to be important for the efficacy of the dianisidine derivative redoxal [3].

Regulatory relationships of DHODH

• We report here that inhibition of cytochrome c oxidase by nitric oxide (NO) indirectly inhibits DHODH activity [20].

Other interactions of DHODH

• The inhibition of human DHODH by A77 1726, the active metabolite of leflunomide, occurs at levels (approximately 600 nM) that are achieved during treatment of RA [21].
• A leflunomide metabolite and Brequinar inhibit dihydroorotate dehydrogenase, thereby suppressing pyrimidine nucleotide synthesis.Cyclosporine and FK-506 (Tacrolimus) inhibit the phosphatase activity of calcineurin, thereby suppressing the production of IL-2 and other cytokines [22].
• Dicoumarol-treated cells accumulated in S phase due to the impairment of pyrimidine biosynthesis at dihydroorotate dehydrogenase step because blockade was overcome by addition of exogenous uridine or orotate, but not by dihydroorotate [23].
• 312-bp and 983-bp regions of the human dihydroorotate dehydrogenase sequence (1496 bp) were amplified by the polymerase chain reaction, and subcloned into the expression vector pQE 32 [24].
• 5-Fluoroorotic acid (H(3)FOro) is a potent inhibitor for some metalloproteins such as dihydroorotase and dihydroorotate dehydrogenase and for thymidylate synthase (nonmetalloprotein) in the human malaria parasite Plasmodium falciparum [25].

Analytical, diagnostic and therapeutic context of DHODH

• The presence of DHODH protein was demonstrated by Western blotting of solubilized membrane fractions with peroxidase conjugated anti-rabbit IgG in combination with chemiluminescence detection [16].
• Anti-arthritic effects of KF20444, a new immunosuppressive compound inhibiting dihydroorotate dehydrogenase, on rat collagen-induced arthritis [26].
• Furthermore, the minimum inhibitory concentrations of these compounds increase upon addition of orotate to the culture medium in a concentration-dependent manner, consistent with dihydroorotate dehydrogenase inhibition as the mechanism of cellular inhibition [5].
• Activities as inhibitors of dihydroorotate dehydrogenase and the mixed lymphocyte reaction were related to the overall shape and lipophilicity of the 2-substituent [27].
• The antiviral activity of FK778 was reversed in cell culture by treatment with uridine, consistent with specific inhibition of dihydroorotate dehydrogenase (DHODH), a required enzyme in the de novo biosynthesis of pyrimidines [28].

References