title: Babesia bovis dihydroorotate dehydrogenase (BboDHODH) is a novel molecular target of drug for bovine babesiosis.
abstract: The emergence of drug resistance and adverse side effects of current bovine babesiosis treatment suggest that the search of new drug targets and development of safer and effective compounds are required. This study focuses on dihydroorotate dehydrogenase (DHODH), the fourth enzyme of pyrimidine biosynthesis pathway as a potential drug target for bovine babesiosis. Recombinant Babesia bovis DHODH protein (rBboDHODH) was produced in Escherichia coli and used for characterization and measurement of enzymatic activity. Furthermore, the effects of DHODH inhibitors were evaluated in vitro. The recombinant B. bovis DHODH histidine fusion protein (rBboDHODH) had 42.4-kDa molecular weight and exhibited a specific activity of 475.7 ± 245 Unit/mg, a Km = 276.2 µM for L-dihydroorotate and a Km= 94.41 µM for decylubiquinone. A 44-kDa band of native BboDHODH was detected by Western blot analysis and found in parasites mitochondria using a confocal microscope. Among DHODH inhibitors, atovaquone (ATV) and leflunomide (LFN) significantly inhibited the activity of rBboDHODH as well as the growth of B. bovis in vitro. The half maximal inhibitory concentration (IC50) of ATV and LFN was 2.38 ± 0.53 nM and 52.41 ± 11.47 µM, respectively. These results suggest that BboDHODH might be a novel target for development of new drug for treatment of B. bovis infection.
authors.affiliation: National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
authors.forename: Ketsarin
authors.initials: K
authors.lastname: Kamyingkird
authors.forename: Shinuo
authors.initials: S
authors.lastname: Cao
authors.forename: Tatsunori
authors.initials: T
authors.lastname: Masatani
authors.forename: Paul Franck Adjou
authors.initials: PF
authors.lastname: Moumouni
authors.forename: Patrick
authors.initials: P
authors.lastname: Vudriko
authors.forename: Ahmed Abd El Moniem
authors.initials: AA
authors.lastname: Mousa
authors.forename: Mohamad Alaa
authors.initials: MA
authors.lastname: Terkawi
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Nishikawa
authors.forename: Ikuo
authors.initials: I
authors.lastname: Igarashi
authors.forename: Xuenan
authors.initials: X
authors.lastname: Xuan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016793:Babesia bovis / Q000201:enzymology*; D001404:Babesiosis / Q000188:drug therapy* / Q000378:metabolism; D015153:Blotting, Western; D002417:Cattle; D002418:Cattle Diseases / Q000188:drug therapy* / Q000469:parasitology*; D003001:Cloning, Molecular; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D000090722:Dihydroorotate Dehydrogenase; D016503:Drug Delivery Systems; D055808:Drug Discovery; D004791:Enzyme Inhibitors / Q000378:metabolism*; D004926:Escherichia coli; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D017931:DNA Primers; D000090722:Dihydroorotate Dehydrogenase; D004791:Enzyme Inhibitors; D011993:Recombinant Fusion Proteins; D044925:Oxidoreductases Acting on CH-CH Group Donors