Biochemical and genetic characterization of Trypanosoma cruzi N-myristoyltransferase

Adam J. Roberts, Leah S. Torrie, Susan Wyllie, Alan H. Fairlamb (Lead / Corresponding author)

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    Abstract

    Co- and post-translational N-myristoylation is known to play a role in the correct subcellular localization of specific proteins in eukaryotes. The enzyme that catalyses this reaction, NMT (N-myristoyltransferase), has been pharmacologically validated as a drug target in the African trypanosome, Trypanosoma brucei. In the present study, we evaluate NMT as a potential drug target in Trypanosoma cruzi, the causative agent of Chagas' disease, using chemical and genetic approaches. Replacement of both allelic copies of TcNMT (T. cruzi NMT) was only possible in the presence of a constitutively expressed ectopic copy of the gene, indicating that this gene is essential for survival of T. cruzi epimastigotes. The pyrazole sulphonamide NMT inhibitor DDD85646 is 13-23-fold less potent against recombinant TcNMT than TbNMT (T. brucei NMT), with Ki values of 12.7 and 22.8 nM respectively, by scintillation proximity or coupled assay methods. DDD85646 also inhibits growth of T. cruzi epimastigotes (EC = 6.9 µM), but is ~1000-fold less potent than that reported for T. brucei. On-target activity is demonstrated by shifts in cell potency in lines that over- and under-express NMT and by inhibition of intracellular N-myristoylation of several proteins in a dose-dependent manner. Collectively, our findings suggest that N-myristoylation is an essential and druggable target in T. cruzi.

    Original languageEnglish
    Pages (from-to)323-332
    Number of pages10
    JournalBiochemical Journal
    Volume459
    Issue number2
    DOIs
    Publication statusPublished - 15 Apr 2014

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    Assessment of N-myristoyltransferase and the N-myristoylomeas a potential chemotherapeutic target inTrypanosoma cruzi

    Author: Roberts, A., 2014

    Supervisor: Fairlamb, A. (Supervisor)

    Student thesis: Doctoral ThesisDoctor of Philosophy

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