Characterization, localization, essentiality, and high-resolution crystal structure of glucosamine 6-phosphate N-acetyltransferase from Trypanosoma brucei

Karina Marino, M. Lucia Sampaio Guther, Amy K. Wernimont, Wei Qiu, Raymond Hui, Michael A. J. Ferguson

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    23 Citations (Scopus)

    Abstract

    A gene predicted to encode Trypanosoma brucei glucosamine 6-phosphate N-acetyltransferase (TbGNA1; EC 2.3.1.4) was cloned and expressed in Escherichia coli. The recombinant protein was enzymatically active, and its high-resolution crystal structure was obtained at 1.86 angstrom. Endogenous TbGNA1 protein was localized to the peroxisome-like microbody, the glycosome. A bloodstream-form T. brucei GNA1 conditional null mutant was constructed and shown to be unable to sustain growth in vitro under nonpermissive conditions, demonstrating that there are no metabolic or nutritional routes to UDP-GlcNAc other than via GlcNAc-6-phosphate. Analysis of the protein glycosylation phenotype of the TbGNA1 mutant under nonpermissive conditions revealed that poly-N-acetyllactosamine structures were greatly reduced in the parasite and that the glycosylation profile of the principal parasite surface coat component, the variant surface glycoprotein (VSG), was modified. The significance of results and the potential of TbGNA1 as a novel drug target for African sleeping sickness are discussed.

    Original languageEnglish
    Pages (from-to)985-997
    Number of pages13
    JournalEukaryotic Cell
    Volume10
    Issue number7
    DOIs
    Publication statusPublished - Jul 2011

    Keywords

    • BLOOD-STREAM FORM
    • PHOSPHATIDYLINOSITOL MEMBRANE ANCHOR
    • GDP-MANNOSE PYROPHOSPHORYLASE
    • VARIANT SURFACE GLYCOPROTEINS
    • GALACTOSE METABOLISM
    • GLUCOSAMINE-6-PHOSPHATE ACETYLTRANSFERASE
    • LEISHMANIA-MAJOR
    • PROTEIN P67
    • CELL-GROWTH
    • GLYCOSYLATION

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