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Data-Independent Acquisition for Yeast Glycoproteomics

  • Lucía F. Zacchi
  • Benjamin L. SchulzEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2049)

Abstract

Glycosylation is a complex posttranslational modification that is critical for regulating the functions of diverse proteins. Analysis of protein glycosylation is made challenging by the high degree of heterogeneity in both glycan occupancy and structure. Here, we describe methods for data-independent acquisition (SWATH) mass spectrometry analysis of structure and occupancy of N-glycans from yeast cell wall glycoproteins.

Key words

Glycosylation Glycoprotein Mass spectrometry Data independent acquisition DIA SWATH Occupancy Macroheterogeneity Structure Microheterogeneity Yeast Cell wall 

Notes

Acknowledgments

Funding: This work was supported by Australian Research Council Discovery Project grant DP160102766 and Australian Research Council Industrial Transformation and Training Centre Grant IC160100027 to BLS. BLS holds an Australian National Health and Medical Research Council RD Wright Biomedical Fellowship APP1087975.

References

  1. 1.
    Aebersold R, Mann M (2016) Mass-spectrometric exploration of proteome structure and function. Nature 537(7620):347–355CrossRefGoogle Scholar
  2. 2.
    Giansanti P et al (2016) Six alternative proteases for mass spectrometry-based proteomics beyond trypsin. Nat Protoc 11(5):993–1006CrossRefGoogle Scholar
  3. 3.
    Varki A et al (2017) Essentials of glycobiology, 3rd edn. Cold Spring Harbor, NYGoogle Scholar
  4. 4.
    Rudd P et al (2015) Glycomics and glycoproteomics. In: Varki A et al (eds) Essentials of glycobiology. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. Copyright 2015 by the consortium of Glycobiology editors, La Jolla, CA. All rights reservedGoogle Scholar
  5. 5.
    Schwarz F, Aebi M (2011) Mechanisms and principles of N-linked protein glycosylation. Curr Opin Struct Biol 21(5):576–582CrossRefGoogle Scholar
  6. 6.
    Kowarik M et al (2006) N-linked glycosylation of folded proteins by the bacterial oligosaccharyltransferase. Science 314(5802):1148–1150CrossRefGoogle Scholar
  7. 7.
    Medus ML et al (2017) N-glycosylation triggers a dual selection pressure in eukaryotic secretory proteins. Sci Rep 7(1):8788CrossRefGoogle Scholar
  8. 8.
    Zacchi LF, Schulz BL (2016) N-glycoprotein macroheterogeneity: biological implications and proteomic characterization. Glycoconj J 33(3):359–376CrossRefGoogle Scholar
  9. 9.
    Caramelo JJ, Parodi AJ (2015) A sweet code for glycoprotein folding. FEBS Lett 589(22):3379–3387CrossRefGoogle Scholar
  10. 10.
    Tate MD et al (2014) Playing hide and seek: how glycosylation of the influenza virus hemagglutinin can modulate the immune response to infection. Viruses 6(3):1294–1316CrossRefGoogle Scholar
  11. 11.
    Freeze HH et al (2014) Solving glycosylation disorders: fundamental approaches reveal complicated pathways. Am J Hum Genet 94(2):161–175CrossRefGoogle Scholar
  12. 12.
    Thaysen-Andersen M, Packer NH, Schulz BL (2016) Maturing glycoproteomics technologies provide unique structural insights into the N-glycoproteome and its regulation in health and disease. Mol Cell Proteomics 15(6):1773–1790CrossRefGoogle Scholar
  13. 13.
    Zacchi LF, Schulz BL (2016) SWATH-MS glycoproteomics reveals consequences of defects in the glycosylation machinery. Mol Cell Proteomics 15(7):2435–2447CrossRefGoogle Scholar
  14. 14.
    Wilson NL et al (2002) Sequential analysis of N- and O-linked glycosylation of 2D-PAGE separated glycoproteins. J Proteome Res 1(6):521–529CrossRefGoogle Scholar
  15. 15.
    Schulz BL et al (2009) Oxidoreductase activity of oligosaccharyltransferase subunits Ost3p and Ost6p defines site-specific glycosylation efficiency. Proc Natl Acad Sci U S A 106(27):11061–11066CrossRefGoogle Scholar
  16. 16.
    Bailey UM, Jamaluddin MF, Schulz BL (2012) Analysis of congenital disorder of glycosylation-id in a yeast model system shows diverse site-specific under-glycosylation of glycoproteins. J Proteome Res 11(11):5376–5383CrossRefGoogle Scholar
  17. 17.
    Gillet LC et al (2012) Targeted data extraction of the MS/MS spectra generated by data-independent acquisition: a new concept for consistent and accurate proteome analysis. Mol Cell Proteomics 11(6):O111.016717CrossRefGoogle Scholar
  18. 18.
    Treco DA, Lundblad V (2000) Basic techniques of yeast genetics, Current protocols in molecular biology. John Wiley and Sons, Piscataway NJ, pp 13.1.1–13.1.7Google Scholar
  19. 19.
    Tran JR, Brodsky JL (2012) Assays to measure ER-associated degradation in yeast. Methods Mol Biol 832:505–518CrossRefGoogle Scholar
  20. 20.
    Loo RR, Dales N, Andrews PC (1996) The effect of detergents on proteins analyzed by electrospray ionization. Methods Mol Biol 61:141–160PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.ARC Training Centre for Biopharmaceutical InnovationThe University of QueenslandSt. LuciaAustralia
  2. 2.School of Chemistry and Molecular BiosciencesThe University of QueenslandSt. LuciaAustralia

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