Article | Published:

Hijacking a biosynthetic pathway yields a glycosyltransferase inhibitor within cells

Nature Chemical Biology volume 7, pages 174181 (2011) | Download Citation

Abstract

Glycosyltransferases are ubiquitous enzymes that catalyze the assembly of glycoconjugates throughout all kingdoms of nature. A long-standing problem is the rational design of probes that can be used to manipulate glycosyltransferase activity in cells and tissues. Here we describe the rational design and synthesis of a nucleotide sugar analog that inhibits, with high potency both in vitro and in cells, the human glycosyltransferase responsible for the reversible post-translational modification of nucleocytoplasmic proteins with O-linked N-acetylglucosamine residues (O-GlcNAc). We show that the enzymes of the hexosamine biosynthetic pathway can transform, both in vitro and in cells, a synthetic carbohydrate precursor into the nucleotide sugar analog. Treatment of cells with the precursor lowers O-GlcNAc in a targeted manner with a single-digit micromolar EC50. This approach to inhibition of glycosyltransferases should be applicable to other members of this superfamily of enzymes and enable their manipulation in a biological setting.

  • Compound C8H15NO6

    2-Acetamido-2-deoxy-D-glucopyranose

  • Compound C17H25N3O17P22-

    Uridine diphospho-N-acetylglucosamine

  • Compound C8H15NO5S

    2-Acetamido-2-deoxy-5-thio-D-glucopyranose

  • Compound C17H25N3O16P2S2-

    Uridine diphospho-5-thio-N-acetylglucosamine

  • Compound C16H23NO9S

    2-Acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-5-thio-α-D-glucopyranose

  • Compound C15H21NO7

    p-Methoxyphenyl 2-acetamido-2-deoxy-β-D-glucopyranoside

  • Compound C15H21NO6S

    p-Methoxyphenyl 2-acetamido-2-deoxy-5-thio-β-D-glucopyranoside

  • Compound C16H23NO10

    2-Acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-α-D-glucopyranose

  • Compound C9H17NO6

    Methyl 2-acetamido-2-deoxy-β-D-glucopyranoside

  • Compound C9H17NO5S

    Methyl 2-acetamido-2-deoxy-5-thio-β-D-glucopyranoside

  • Compound C8H14N4O6

    2-Azidoacetamido-2-deoxy-D-glucopyranose

  • Compound C17H24N6O17P22-

    Uridine diphospho-N-azidoacetylglucosamine

  • Compound C8H14N4O5S

    2-Azidoacetamido-2-deoxy-5-thio-D-glucopyranose

  • Compound C16H22N4O9S

    1,3,4,6-Tetra-O-acetyl-2-azidoacetamido-2-deoxy-5-thio-α-D-glucopyranose

  • Compound C16H22N4O10

    1,3,4,6-Tetra-O-acetyl-2-azidoacetamido-2-deoxy-α-D-glucopyranose

  • Compound C17H24N6O16P2S2-

    Uridine diphospho-5-thio-N-azidoacetylglucosamine

  • Compound C16H29NO7

    2-Acetamido-2-deoxy-3,4,5,6-di-O-isopropylidine-aldehydo-D-glucose dimethyl acetal

  • Compound C13H25NO7

    2-Acetamido-2-deoxy-3,4-O-isopropylidine-aldehydo-D-glucose dimethyl acetal

  • Compound C20H29NO8

    2-Acetamido-6-O-benzoyl-2-deoxy-3,4-O-isopropylidene-aldehydo-D-glucose dimethyl acetal

  • Compound C21H31NO10S

    2-Acetamido-6-O-benzoyl-2-deoxy-3,4-O-isopropylidine-5-O-mesyl-aldehydo-D-glucose dimethyl acetal

  • Compound C13H23NO6

    2-Acetamido-5,6-anhydro-2-deoxy-3,4-O-isopropylidene-aldehydo-L-idose dimethyl acetal

  • Compound C13H23NO5S

    2-Acetamido-2,5,6-trideoxy-5,6-epithio-3,4-O-isopropylidine-aldehydo-D-glucose dimethyl acetal

  • Compound C17H29NO8S

    2-Acetamido-6-O-acetyl-5-S-acetyl-2-deoxy-3,4-O-isopropylidine-5-thio-aldehydo-D-glucose dimethyl acetal

  • Compound C14H21NO8S

    2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-5-thio-α-D-glucopyranose

  • Compound C16H21Cl3N2O8S

    2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-5-thio-α-D-glucopyranosyl trichloroacetimidate

  • Compound C22H29NO8S

    p-Methoxyphenyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-5-thio-β-D-glucopyranoside

  • Compound C15H23NO8S

    Methyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-5-thio-β-D-glucopyranoside

  • Compound C14H19N3O8S

    1,3,4,6-Tetra-O-acetyl-2-azido-2-deoxy-5-thio-α-D-glucopyranoside

  • Compound C14H22ClNO8S

    2-Amino-1,3,4,6-tetra-O-acetyl-5-thio-α-D-glucopyranose hydrochloride

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Acknowledgements

The plasmid encoding GNK was a gift from M. Berger (Charite Universitatsmedizin Berlin) and S. Hinderlich (University of Applied Sciences, Berlin), and the plasmid encoding AGX1 was a gift from V. Piller (Centre de Recherche Scientifique, UPR 4301). Polyclonal antibody to OGA was a gift from G. Hart (John Hopkins University). EMEG32 cells were a gift from T. Mak (University of Toronto). We thank G. Davies (University of York) for the E. coli expression construct of BtGH84. T.M.G. is a Sir Henry Wellcome postdoctoral fellow and a Michael Smith for Health Research (MSFHR) trainee award holder. D.J.V. is a scholar of MSFHR and holds a Canada Research Chair in Chemical Glycobiology. We thank the Natural Sciences and Engineering Research Council of Canada and Simon Fraser University for funding support.

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Affiliations

  1. Department of Chemistry, Simon Fraser University, Burnaby, British Columbia, Canada.

    • Tracey M Gloster
    • , Wesley F Zandberg
    • , Julia E Heinonen
    • , Lehua Deng
    •  & David J Vocadlo
  2. Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, British Columbia, Canada.

    • David L Shen
    •  & David J Vocadlo

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Contributions

T.M.G. and D.J.V. designed experiments. T.M.G., W.F.Z., J.E.H., D.L.S. and L.D. carried out experiments. T.M.G., W.F.Z., D.L.S. and D.J.V. analyzed results. T.M.G. and D.J.V wrote the manuscript with input from all authors.

Competing interests

Provisional patent applications covering this work have been filed.

Corresponding author

Correspondence to David J Vocadlo.

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DOI

https://doi.org/10.1038/nchembio.520

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