Identification of N-glycans from Ebola virus glycoproteins by matrix-assisted laser desorption/ionisation time-of-flight and negative ion electrospray tandem mass spectrometry

Authors

  • Gayle Ritchie,

    1. Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford UK
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  • David J. Harvey,

    Corresponding author
    1. Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford UK
    • Oxford Glycobiology Institute, Department of Biochemistry, South Parks Road, Oxford OX1 3QU, UK.
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  • Ute Stroeher,

    1. Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, Canada, R3E 3R2
    2. Department of Medical Microbiology, University of Manitoba, 543-730 William Avenue, Winnipeg, Manitoba, Canada, R3E 0W3
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  • Friederike Feldmann,

    1. Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, Canada, R3E 3R2
    2. Department of Medical Microbiology, University of Manitoba, 543-730 William Avenue, Winnipeg, Manitoba, Canada, R3E 0W3
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  • Heinz Feldmann,

    1. Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, Canada, R3E 3R2
    2. Department of Medical Microbiology, University of Manitoba, 543-730 William Avenue, Winnipeg, Manitoba, Canada, R3E 0W3
    Current affiliation:
    1. Rocky Mountain Laboratories, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 S 4th Street, Hamilton, Montana, United States 59840.
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  • Victoria Wahl-Jensen,

    1. Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, Canada, R3E 3R2
    2. Viral Therapeutics Branch, Virology Division, US Army Medical Research Institute for Infectious Diseases, 1425 Porter Street, Fort Detrick, Maryland 21702, USA
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  • Louise Royle,

    1. National Institute for Bioprocessing Research and Training, Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland
    Current affiliation:
    1. Ludger Ltd., Culham Science Centre, Oxfordshire OX14 3EB, UK.
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  • Raymond A. Dwek,

    1. Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford UK
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  • Pauline M. Rudd

    1. National Institute for Bioprocessing Research and Training, Conway Institute, University College Dublin, Belfield, Dublin 4, Ireland
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Abstract

The larger fragment of the transmembrane glycoprotein (GP1) and the soluble glycoprotein (sGP) of Ebola virus were expressed in human embryonic kidney cells and the secreted products were purified from the supernatant for carbohydrate analysis. The N-glycans were released with PNGase F from within sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS-PAGE) gels. Identification of the glycans was made with normal-phase high-performance liquid chromatography (HPLC), matrix-assisted laser desorption/ionisation mass spectrometry, negative ion electrospray ionisation fragmentation mass spectrometry and exoglycosidase digestion. Most glycans were complex bi-, tri- and tetra-antennary compounds with reduced amounts of galactose. No bisected compounds were detected. Triantennary glycans were branched on the 6-antenna; fucose was attached to the core GlcNAc residue. Sialylated glycans were present on sGP but were largely absent from GP1, the larger fragment of the transmembrane glycoprotein. Consistent with this was the generally higher level of processing of carbohydrates found on sGP as evidenced by a higher percentage of galactose and lower levels of high-mannose glycans than were found on GP1. These results confirm and expand previous findings on partial characterisation of the Ebola virus transmembrane glycoprotein. They represent the first detailed data on carbohydrate structures of the Ebola virus sGP. Copyright © 2010 John Wiley & Sons, Ltd.

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