Probing the substrate specificity of Trypanosoma brucei GlcNAc-PI de-N-acetylase with synthetic substrate analogues

Amy S. Capes, Arthur Crossman, Michael D. Urbaniak, Sophie H. Gilbert, Michael A. J. Ferguson (Lead / Corresponding author), Ian H. Gilbert (Lead / Corresponding author)

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    Abstract

    A series of synthetic analogues of 1-D-(2-amino-2-deoxy-?-D-glucopyranosyl)-myo-inositol 1-(1,2-di-O-hexadecanoyl-sn-glycerol 3-phosphate), consisting of 7 variants of either the d-myo-inositol, d-GlcpN or the phospholipid components, were prepared and tested as substrates and inhibitors of GlcNAc-PI de-N-acetylase, a genetically validated drug target enzyme responsible for the second step in the glycosylphosphatidylinositol (GPI) biosynthetic pathway of Trypanosoma brucei. The D-myo-inositol in the physiological substrate was successfully replaced by cyclohexanediol and is still a substrate for T. brucei GlcNAc-PI de-N-acetylase. However, this compound became sensitive to the stereochemistry of the glycoside linkage (the ß-anomer was neither substrate or inhibitor) and the structure of the lipid moiety (the hexadecyl derivatives were inhibitors). Chemistry was successfully developed to replace the phosphate with a sulphonamide, but the compound was neither a substrate or an inhibitor, confirming the importance of the phosphate for molecular recognition. We also replaced the glucosamine by an acyclic analogue, but this also was inactive, both as a substrate and inhibitor. These findings add significantly to our understanding of substrate and inhibitor binding to the GlcNAc-PI de-N-acetylase enzyme and will have a bearing on the design of future inhibitors.

    Original languageEnglish
    Pages (from-to)1919-1934
    Number of pages16
    JournalOrganic and Biomolecular Chemistry
    Volume12
    Issue number12
    DOIs
    Publication statusPublished - Mar 2014

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