title: Comparison of RP-HPLC modes to analyse the N-glycome of the free-living nematode Pristionchus pacificus.
abstract: Pristionchus pacificus is a free-living nematode increasingly used as an organism for comparison to the more familiar model Caenorhabditis elegans. In this study, we examined the N-glycans of this organism isolated after serial release with peptide:N-glycosidases F and A; after fluorescent labelling with 2-aminopyridine, chromatographic fractionation by three types of RP-HPLC (with either classical C18, fused core C18 or alkylamide-bonded phases) followed by mass spectrometric analyses revealed key features of its N-glycome. In addition to paucimannosidic and oligomannosidic glycans typical of invertebrates, N-glycans with two core fucose residues were detected. Furthermore, a range of glycans carrying up to three phosphorylcholine residues was observed whereas, unlike C. elegans, no tetrafucosylated N-glycans were detected. Structures with three fucose residues, unusual methylation of core α1,3-fucose or with galactosylated fucose motifs were found in low amounts; these features may correlate with a different ensemble or expression of glycosyltransferase genes as compared to C. elegans. From an analytical perspective, both the alkylamide RP-amide and fused core C18 columns, as compared to a classical C18 material, offer advantages in terms of resolution and of elution properties, as some minor pyridylamino-labelled glycans (e.g. those carrying phosphorylcholine) appear in earlier fractions and so potential losses of such structures due to insufficient gradient length can be avoided.
authors: Department für Chemie, Universität für Bodenkultur, Wien, Austria.
authors: Shi
authors: S
authors: Yan
authors: Department für Chemie, Universität für Bodenkultur, Wien, Austria.
authors: Iain B H
authors: IB
authors: Wilson
authors: Department für Chemie, Universität für Bodenkultur, Wien, Austria.
authors: Katharina
authors: K
authors: Paschinger
mesh_terms: D000818:Animals; D002241:Carbohydrates / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D056148:Chromatography, Reverse-Phase; D009348:Nematoda / Q000737:chemistry*
chemical_list: D002241:Carbohydrates
keywords: Fucose; N-glycans; Nematode; Phosphorylcholine; Pyridylamination; RP-HPLC
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