Characterization of the novel Trypanosoma brucei inosine 5 '-monophosphate dehydrogenase

Tomoaki Bessho, Shoko Morii, Toshihide Kusumoto, Takahiro Shinohara, Masanori Noda, Susumu Uchiyama, Satoshi Shuto, Shigenori Nishimura, Appolinaire Djikeng, Michael Duszenko, Samuel K. Martin, Takashi Inui*, Kilunga B. Kubata

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

There is an alarming rate of human African trypanosomiasis recrudescence in many parts of sub-Saharan Africa. Yet, the disease has no successful chemotherapy. Trypanosoma lacks the enzymatic machinery for the de novo synthesis of purine nucleotides, and is critically dependent on salvage mechanisms. Inosine 5'-monophosphate dehydrogenase (IMPDH) is responsible for the rate-limiting step in guanine nucleotide metabolism. Here, we characterize recombinant Trypanosoma brucei IMPDH (TbIMPDH) to investigate the enzymatic differences between TbIMPDH and host IMPDH. Size-exclusion chromatography and analytical ultracentrifugation sedimentation velocity experiments reveal that TbIMPDH forms a heptamer, different from type 1 and 2 mammalian tetrameric IMPDHs. Kinetic analysis reveals calculated K-m values of 30 and 1300 mu M for IMP and NAD, respectively. The obtained K-m value of TbIMPDH for NAD is approximately 20-200-fold higher than that of mammalian enzymes and indicative of a different NAD binding mode between trypanosomal and mammalian IMPDHs. Inhibition studies show K-i values of 3.2 mu M, 21 nM and 3.3 nM for ribavirin 5'-monophosphate, mycophenolic acid and mizoribine 5'-monophosphate, respectively. Our results show that TbIMPDH is different from its mammalian counterpart and thus may be a good target for further studies on anti-trypanosomal drugs.

Original languageEnglish
Pages (from-to)735-745
Number of pages11
JournalParasitology
Volume140
Issue number6
DOIs
Publication statusPublished - May 2013

Keywords

  • trypanosome
  • enzyme kinetics
  • enzyme inhibitor
  • nucleoside nucleotide analogues
  • drug design
  • HUMAN AFRICAN TRYPANOSOMIASIS
  • TRANSITION-STATE ANALOG
  • HEPATITIS-C VIRUS
  • MONOPHOSPHATE DEHYDROGENASE
  • TRITRICHOMONAS-FETUS
  • IMP DEHYDROGENASE
  • CRYSTAL-STRUCTURE
  • ANTIVIRAL ACTIVITY
  • KINETIC MECHANISM
  • DRUG DESIGN

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