Hot and sweet: protein glycosylation in Crenarchaeota

2013 ◽  
Vol 41 (1) ◽  
pp. 384-392 ◽  
Author(s):  
Benjamin H. Meyer ◽  
Sonja-Verena Albers
Keyword(s):  
Living Cell ◽  
Protein Glycosylation ◽  
Haloferax Volcanii ◽  
Present Review ◽  
Sulfolobus Acidocaldarius ◽  
Glycosylphosphatidylinositol Anchor ◽  
Sweet Protein ◽  
Domains Of Life ◽  
Glycosylation Pathway ◽  
Sugar Chains

Every living cell is covered with a dense and complex array of covalently attached sugars or sugar chains. The majority of these glycans are linked to proteins via the so-called glycosylation process. Protein glycosylation is found in all three domains of life: Eukarya, Bacteria and Archaea. However, on the basis of the limit in analytic tools for glycobiology and genetics in Archaea, only in the last few years has research on archaeal glycosylation pathways started mainly in the Euryarchaeota Haloferax volcanii, Methanocaldococcus maripaludis and Methanococcus voltae. Recently, major steps of the crenarchaeal glycosylation process of the thermoacidophilic archaeon Sulfolobus acidocaldarius have been described. The present review summarizes the proposed N-glycosylation pathway of S. acidocaldarius, describing the phenotypes of the mutants disrupted in N-glycan biosynthesis as well as giving insights into the archaeal O-linked and glycosylphosphatidylinositol anchor glycosylation process.

Add salt, add sugar: N-glycosylation in Haloferax volcanii

2013 ◽  
Vol 41 (1) ◽  
pp. 432-435 ◽  
Author(s):  
Lina Kaminski ◽  
Shai Naparstek ◽  
Lina Kandiba ◽  
Chen Cohen-Rosenzweig ◽  
Adi Arbiv ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Molecular Biology ◽  
Haloferax Volcanii ◽  
Present Review ◽  
Post Translational Modification ◽  
Environmental Salinity ◽  
Native Proteins ◽  
Domains Of Life ◽  
Adaptive Role

Although performed by members of all three domains of life, the archaeal version of N-glycosylation remains the least understood. Studies on Haloferax volcanii have, however, begun to correct this situation. A combination of bioinformatics, molecular biology, biochemical and mass spectrometry approaches have served to delineate the Agl pathway responsible for N-glycosylation of the S-layer glycoprotein, a reporter of this post-translational modification in Hfx. volcanii. More recently, differential N-glycosylation of the S-layer glycoprotein as a function of environmental salinity was demonstrated, showing that this post-translational modification serves an adaptive role in Hfx. volcanii. Furthermore, manipulation of the Agl pathway, together with the capability of Hfx. volcanii to N-glycosylate non-native proteins, forms the basis for establishing this species as a glyco-engineering platform. In the present review, these and other recent findings are addressed.

scholarly journals N-Glycosylation of Haloferax volcanii Flagellins Requires Known Agl Proteins and Is Essential for Biosynthesis of Stable Flagella

2012 ◽  
Vol 194 (18) ◽  
pp. 4876-4887 ◽  
Author(s):  
Manuela Tripepi ◽  
Jason You ◽  
Sevcan Temel ◽  
Özlem Önder ◽  
Dustin Brisson ◽  
...  
Keyword(s):  
Gradient Centrifugation ◽  
Haloferax Volcanii ◽  
Content Type ◽  
Glycosylation Sites ◽  
Cscl Density Gradient ◽  
Domains Of Life ◽  
Covalently Linked ◽  
Glycosylation Pathway ◽  
A Minor ◽  
And Function

ABSTRACTN-glycosylation, a posttranslational modification required for the accurate folding and stability of many proteins, has been observed in organisms of all domains of life. Although the haloarchaeal S-layer glycoprotein was the first prokaryotic glycoprotein identified, little is known about the glycosylation of other haloarchaeal proteins. We demonstrate here that the glycosylation ofHaloferax volcaniiflagellins requires archaeal glycosylation (Agl) components involved in S-layer glycosylation and that the deletion of anyHfx. volcaniiaglgene impairs its swimming motility to various extents. A comparison of proteins in CsCl density gradient centrifugation fractions from supernatants of wild-typeHfx. volcaniiand deletion mutants lacking the oligosaccharyltransferase AglB suggests that when the Agl glycosylation pathway is disrupted, cells lack stable flagella, which purification studies indicate consist of a major flagellin, FlgA1, and a minor flagellin, FlgA2. Mass spectrometric analyses of FlgA1 confirm that its three predicted N-glycosylation sites are modified with covalently linked pentasaccharides having the same mass as that modifying its S-layer glycoprotein. Finally, the replacement of any of three predicted N-glycosylated asparagines of FlgA1 renders cells nonmotile, providing direct evidence for the first time that the N-glycosylation of archaeal flagellins is critical for motility. These results provide insight into the role that glycosylation plays in the assembly and function ofHfx. volcaniiflagella and demonstrate thatHfx. volcaniiflagellins are excellent reporter proteins for the study of haloarchaeal glycosylation processes.

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

Glycobiology ◽  
2018 ◽  
Vol 28 (4) ◽  
pp. 233-244 ◽  
Author(s):  
Adrian J Jervis ◽  
Alison G Wood ◽  
Joel A Cain ◽  
Jonathan A Butler ◽  
Helen Frost ◽  
...  
Keyword(s):  
Polyclonal Antiserum ◽  
Protein Glycosylation ◽  
Chromosomal Locus ◽  
Individual Gene ◽  
Knockout Mutants ◽  
Domains Of Life ◽  
General Protein ◽  
Glycosylation Pathway

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.

Salty and Sweet: Protein Glycosylation in Haloferax volcanii

2011 ◽  
pp. 227-235
Author(s):  
Jerry Eichler ◽  
Doron Calo ◽  
Lina Kaminski ◽  
Lina Kandiba ◽  
Zvia Konrad ◽  
...  
Keyword(s):  
Protein Glycosylation ◽  
Haloferax Volcanii ◽  
Sweet Protein

scholarly journals Mgat5 Deficiency in T Cells and Experimental Autoimmune Encephalomyelitis

2011 ◽  
Vol 2011 ◽  
pp. 1-6 ◽  
Author(s):  
Ani Grigorian ◽  
Michael Demetriou
Keyword(s):  
T Cells ◽  
T Cell ◽  
Experimental Autoimmune Encephalomyelitis ◽  
Demyelinating Disease ◽  
Cell Receptor ◽  
Protein Glycosylation ◽  
Autoimmune Encephalomyelitis ◽  
Glycosylation Pathway ◽  
Experimental Autoimmune ◽  
Surface Glycoproteins

Multiple sclerosis (MS) is an inflammatory demyelinating and neurodegenerative disease initiated by autoreactive T cells. Mgat5, a gene in the Asn (N-) linked protein glycosylation pathway, associates with MS severity and negatively regulates experimental autoimmune encephalomyelitis (EAE) and spontaneous inflammatory demyelination in mice. N-glycan branching by Mgat5 regulates interaction of surface glycoproteins with galectins, forming a molecular lattice that differentially controls the concentration of surface glycoproteins. T-cell receptor signaling, T-cell proliferation, TH1 differentiation, and CTLA-4 endocytosis are inhibited by Mgat5 branching. Non-T cells also contribute to MS pathogenesis and express abundant Mgat5 branched N-glycans. Here we explore whether Mgat5 deficiency in myelin-reactive T cells is sufficient to promote demyelinating disease. Adoptive transfer of myelin-reactive Mgat5−/− T cells into Mgat5+/+ versus Mgat5−/− recipients revealed more severe EAE in the latter, suggesting that Mgat5 branching deficiency in recipient naive T cells and/or non-T cells contribute to disease pathogenesis.

scholarly journals The Hitchhiker's guide to glycoproteomics

2021 ◽  
Author(s):  
Tiago Oliveira ◽  
Morten Thaysen-Andersen ◽  
Nicolle H. Packer ◽  
Daniel Kolarich
Keyword(s):  
Functional Analysis ◽  
Cell Function ◽  
Prognostic Biomarker ◽  
Protein Glycosylation ◽  
Life Changes ◽  
Carrier Proteins ◽  
Post Translational Modifications ◽  
Starting Point ◽  
Domains Of Life ◽  
The Common

Protein glycosylation is one of the most common post-translational modifications that are essential for cell function across all domains of life. Changes in glycosylation are considered a hallmark of many diseases, thus making glycoproteins important diagnostic and prognostic biomarker candidates and therapeutic targets. Glycoproteomics, the study of glycans and their carrier proteins in a system-wide context, is becoming a powerful tool in glycobiology that enables the functional analysis of protein glycosylation. This ‘Hitchhiker's guide to glycoproteomics’ is intended as a starting point for anyone who wants to explore the emerging world of glycoproteomics. The review moves from the techniques that have been developed for the characterisation of single glycoproteins to technologies that may be used for a successful complex glycoproteome characterisation. Examples of the variety of approaches, methodologies, and technologies currently used in the field are given. This review introduces the common strategies to capture glycoprotein-specific and system-wide glycoproteome data from tissues, body fluids, or cells, and a perspective on how integration into a multi-omics workflow enables a deep identification and characterisation of glycoproteins — a class of biomolecules essential in regulating cell function.

scholarly journals Cj1496c Encodes a Campylobacter jejuni Glycoprotein That Influences Invasion of Human Epithelial Cells and Colonization of the Chick Gastrointestinal Tract

2006 ◽  
Vol 74 (8) ◽  
pp. 4715-4723 ◽  
Author(s):  
Tsutomu Kakuda ◽  
Victor J. DiRita
Keyword(s):  
Campylobacter Jejuni ◽  
Cell Invasion ◽  
Protein Glycosylation ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
Human Intestinal Epithelial Cell ◽  
Site Specific ◽  
Glycosylation Pathway ◽  
Gastrointestinal Colonization ◽  
Insertion Mutants

ABSTRACT Campylobacter jejuni has an N-linked protein glycosylation pathway that is required for efficient cell invasion and chick gastrointestinal colonization by the microbe. In this study, we constructed insertion mutants of 22 putative glycoprotein genes and examined the ability of each to invade the human intestinal epithelial cell line INT-407. Among the mutants tested, one carrying an insertion in Cj1496c was defective for invasion into INT-407 cells; this defect was also observed in an in-frame deletion mutant of Cj1496c (ΔCj1496c). The ΔCj1496c mutant C. jejuni also showed a reduced ability to colonize chick ceca. Site-specific mutagenesis combined with Western blot analysis suggested that the Cj1496c protein is glycosylated at N73 and N169. However, the ΔCj1496c mutant expressing a nonglycosylated form of Cj1496c exhibited levels of invasion and colonization equivalent to those of the parent strain, suggesting that glycans are not directly involved in the function of Cj1496c.

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