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Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system
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 responsib...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025236/ https://www.ncbi.nlm.nih.gov/pubmed/29340583 http://dx.doi.org/10.1093/glycob/cwx110 |
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author | Jervis, Adrian J Wood, Alison G Cain, Joel A Butler, Jonathan A Frost, Helen Lord, Elizabeth Langdon, Rebecca Cordwell, Stuart J Wren, Brendan W Linton, Dennis |
author_facet | Jervis, Adrian J Wood, Alison G Cain, Joel A Butler, Jonathan A Frost, Helen Lord, Elizabeth Langdon, Rebecca Cordwell, Stuart J Wren, Brendan W Linton, Dennis |
author_sort | Jervis, Adrian J |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-6025236 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-60252362018-07-10 Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system Jervis, Adrian J Wood, Alison G Cain, Joel A Butler, Jonathan A Frost, Helen Lord, Elizabeth Langdon, Rebecca Cordwell, Stuart J Wren, Brendan W Linton, Dennis Glycobiology Regular Manuscripts 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. Oxford University Press 2018-01-11 /pmc/articles/PMC6025236/ /pubmed/29340583 http://dx.doi.org/10.1093/glycob/cwx110 Text en © The Author(s) 2018. Published by Oxford University Press. http://creativecommons.org/licenses/by/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Regular Manuscripts Jervis, Adrian J Wood, Alison G Cain, Joel A Butler, Jonathan A Frost, Helen Lord, Elizabeth Langdon, Rebecca Cordwell, Stuart J Wren, Brendan W Linton, Dennis Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title | Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title_full | Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title_fullStr | Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title_full_unstemmed | Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title_short | Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system |
title_sort | functional analysis of the helicobacter pullorum n-linked protein glycosylation system |
topic | Regular Manuscripts |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025236/ https://www.ncbi.nlm.nih.gov/pubmed/29340583 http://dx.doi.org/10.1093/glycob/cwx110 |
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