Building a PGC-LC-MS N-glycan retention library and elution mapping resource

Jodie L. Abrahams, Matthew P. Campbell, Nicolle H. Packer*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    57 Citations (Scopus)

    Abstract

    Porous graphitised carbon-liquid chromatography (PGC-LC) has been proven to be a powerful technique for the analysis and characterisation of complex mixtures of isomeric and isobaric glycan structures. Here we evaluate the elution behaviour of N-glycans on PGC-LC and thereby provide the potential of using chromatographic separation properties, together with mass spectrometry (MS) fragmentation, to determine glycan structure assignments more easily. We used previously reported N-glycan structures released from the purified glycoproteins Immunoglobulin G (IgG), Immunoglobulin A (IgA), lactoferrin, α1-acid glycoprotein, Ribonuclease B (RNase B), fetuin and ovalbumin to profile their behaviour on capillary PGC-LC-MS. Over 100 glycan structures were determined by MS/MS, and together with targeted exoglycosidase digestions, created a N-glycan PGC retention library covering a full spectrum of biologically significant N-glycans from pauci mannose to sialylated tetra-antennary classes. The resultant PGC retention library (http://www.glycostore.org/showPgc) incorporates retention times and supporting fragmentation spectra including exoglycosidase digestion products, and provides detailed knowledge on the elution properties of N-glycans by PGC-LC. Consequently, this platform should serve as a valuable resource for facilitating the detailed analysis of the glycosylation of both purified recombinant, and complex mixtures of, glycoproteins using established workflows.

    Original languageEnglish
    Pages (from-to)15-29
    Number of pages15
    JournalGlycoconjugate Journal
    Volume35
    Issue number1
    Early online date13 Sep 2017
    DOIs
    Publication statusPublished - Feb 2018

    Keywords

    • N-glycan
    • Glycoprotein
    • Porous graphitised carbon
    • Liquid chromatography
    • Mass spectrometry

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