title: Cloning, characterization and validation of inosine 5'-monophosphate dehydrogenase of Babesia gibsoni as molecular drug target.
abstract: The inosine monophosphate dehydrogenase (IMPDH) enzyme has been characterized and validated as a molecular drug target in other apicomplexans but not in the genus Babesia. Subsequently, we cloned and expressed a Babesia gibsoni IMPDH (BgIMPDH) cDNA in Escherichia coli. We also determined the inhibitory effect of mycophenolic acid (MPA) on recombinant BgIMPDH (rBgIMPDH) activity and the Babesia-growths in vitro. The translated BgIMPDH peptide contained thirteen amino acid residues responsible for substrate and cofactor binding in its catalytic domain with Gly374 in BgIMPDH being replaced by Ser388 in mammalian IMPDH. The native BgIMPDH enzyme in the parasite was approximately 54-kDa a mass similar to His-tag rBgIMPDH protein. The Km values of the rBgIMPDH were 8.18±0.878 (mean±standard error of the mean) μM and 360.80±43.41μM for IMP and NAD(+), respectively. MPA inhibited the rBgIMPDH activity yielding a Ki value of 20.93±1.83μM with respect to NAD(+). For Babesia growths, the IC50s were 0.95±0.21 and 2.88±0.49μM for B. gibsoni and B. bovis, respectively. Therefore, our results suggest that MPA may inhibit the replication of Babesia parasites by targeting IMPDH enzyme of the purine pathway.
authors.affiliation: National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido, Japan.
authors.forename: Shinuo
authors.initials: S
authors.lastname: Cao
authors.forename: Gabriel Oluga
authors.initials: GO
authors.lastname: Aboge
authors.forename: Mohamad Alaa
authors.initials: MA
authors.lastname: Terkawi
authors.forename: Mo
authors.initials: M
authors.lastname: Zhou
authors.forename: Yuzi
authors.initials: Y
authors.lastname: Luo
authors.forename: Longzheng
authors.initials: L
authors.lastname: Yu
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
authors.forename: Younkyoung
authors.initials: Y
authors.lastname: Goo
authors.forename: Ketsarin
authors.initials: K
authors.lastname: Kamyingkird
authors.forename: Tatsunori
authors.initials: T
authors.lastname: Masatani
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Suzuki
authors.forename: Ikuo
authors.initials: I
authors.lastname: Igarashi
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Nishikawa
authors.forename: Xuenan
authors.initials: X
authors.lastname: Xuan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology*; D001403:Babesia / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D001483:Base Sequence; D002417:Cattle; D003001:Cloning, Molecular; D019295:Computational Biology; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004912:Erythrocytes / Q000469:parasitology; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005260:Female; D015870:Gene Expression; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D009173:Mycophenolic Acid / Q000494:pharmacology*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
chemical_list: D000913:Antibodies, Protozoan; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D007168:IMP Dehydrogenase; D009173:Mycophenolic Acid