期刊
GLYCOBIOLOGY
卷 28, 期 4, 页码 233-+出版社
OXFORD UNIV PRESS INC
DOI: 10.1093/glycob/cwx110
关键词
bacteria; glycoprotein; glycosylation; Helicobacter; N-linked
资金
- UK Biotechnology and Biological Science Research Council [BB/F009321/1, BB/H017542/1]
- Wellcome Trust [102979/13/Z]
- Australian Research Council [DP110103573]
- BBSRC DTP studentship
- Australian Postgraduate Award
- BBSRC [BB/H017437/1, BB/F009321/1, BB/N001591/1, BB/H017542/1] Funding Source: UKRI
- Biotechnology and Biological Sciences Research Council [BB/H017437/1, BB/H017542/1, BBS/B/04862, BB/F009321/1, E20372, BB/N001591/1, EGA16167] Funding Source: researchfish
- Wellcome Trust [102979/Z/13/Z] Funding Source: researchfish
N-linked protein glycosylation systems operate in species from all three domains of life. The model bacterial N-linked glycosylation system from Campylobacter jejuni is encoded by pgl genes present at a single chromosomal locus. This gene cluster includes the pglB oligosaccharyltransferase responsible for transfer of glycan from lipid carrier to protein. Although all genomes from species of the Campylobacter genus contain a pgl locus, among the related Helicobacter genus only three evolutionarily related species (H. pullorum, H. canadensis and H. winghamensis) potentially encode N-linked protein glycosylation systems. Helicobacter putative pgl genes are scattered in five chromosomal loci and include two putative oligosaccharyltransferase-encoding pglB genes per genome. We have previously demonstrated the in vitro N-linked glycosylation activity of H. pullorum resulting in transfer of a pentasaccharide to a peptide at asparagine within the sequon (D/E) XNXS/T. In this study, we identified the first H. pullorum N-linked glycoprotein, termed HgpA. Production of histidine-tagged HgpA in the background of insertional knockout mutants of H. pullorum pgl/wbp genes followed by analysis of HgpA glycan structures demonstrated the role of individual gene products in the PglB1-dependent N-linked protein glycosylation pathway. Glycopeptide purification by zwitterionic-hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry identified six glycosites from five H. pullorum proteins, which was consistent with proteins reactive with a polyclonal antiserum generated against glycosylated HgpA. This study demonstrates functioning of a H. pullorum N-linked general protein glycosylation system.
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