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Analysis of Caenorhabditis Protein Glycosylation

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Recombinant Glycoproteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2762))

Abstract

Glycoproteins result from post-translational modification of proteins by glycans attached to certain side chains, with possible heterogeneity due to different structures being possible at the same glycosylation site.

In contrast to the mammalian systems, analysis of invertebrate glycans presents a challenge in analysis as there exist unfamiliar epitopes and a high degree of structural and isomeric variation between different species—Caenorhabditis elegans is no exception. Simple screening using lectins and antibodies can yield hints regarding which glycan epitopes are present in wild-type and mutant strains, but detailed analysis is necessary for determining more exact glycomic information. Here, our analytical approach is to analyze N- and O-glycans involving “off-line” RP-HPLC MALDI-TOF MS/MS. Enrichment and labeling steps facilitate the analysis of single structures and provide isomeric separation. Thereby, the “simple” worm expresses over 200 N-glycan structures varying depending on culture conditions or the genetic background.

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Abbreviations

DTT:

dithiothreitol

HRP:

horseradish peroxidase

MALDI-TOF MS:

matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry

NPGC:

non-porous graphitized carbon

PA:

pyridylamino

PC:

phosphorylcholine

RP-HPLC:

reversed phase high-pressure liquid chromatography

SDS-PAGE:

sodium dodecyl sulfate polyacrylamide gel electrophoresis

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Acknowledgments

This work was supported by the Austrian Fonds zur Förderung der wissenschaftlichen Forschung (FWF; grants P32572 and P29466 to K.P and I.B.H.W.).

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Authors and Affiliations

  1. Institut für Biochemie, Department für Chemie, Universität für Bodenkultur, Vienna, Austria

    Katharina Paschinger, Jorick Vanbeselaere & Iain B. H. Wilson

Authors
  1. Katharina Paschinger
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  2. Jorick Vanbeselaere
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  3. Iain B. H. Wilson
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Corresponding author

Correspondence to Iain B. H. Wilson .

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Editors and Affiliations

  1. Center for Global Health, University of New Mexico Health Sciences Center, Albuquerque, NM, USA

    Steven B. Bradfute

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Paschinger, K., Vanbeselaere, J., Wilson, I.B.H. (2024). Analysis of Caenorhabditis Protein Glycosylation. In: Bradfute, S.B. (eds) Recombinant Glycoproteins. Methods in Molecular Biology, vol 2762. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3666-4_8

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  • DOI: https://doi.org/10.1007/978-1-0716-3666-4_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3665-7

  • Online ISBN: 978-1-0716-3666-4

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