e-space
Manchester Metropolitan University's Research Repository

Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system.

Jervis, AJ, Wood, AG, Cain, JA, Butler, JA, Frost, H, Lord, E, Langdon, R, Cordwell, SJ, Wren, BW and Linton, D (2018) Functional analysis of the Helicobacter pullorum N-linked protein glycosylation system. Glycobiology, 28 (4). pp. 233-244. ISSN 0959-6658

[img]
Preview

Available under License Creative Commons Attribution Non-commercial.

Download (1MB) | Preview

Abstract

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.

Impact and Reach

Statistics

Activity Overview
6 month trend
283Downloads
6 month trend
262Hits

Additional statistics for this dataset are available via IRStats2.

Altmetric

Actions (login required)

View Item View Item