Identification and Origin of <i>N</i>-Linked β-<scp>d</scp>-<i>N-</i>Acetylglucosamine Monosaccharide Modifications on Arabidopsis Proteins

Kim, Young Cheon; Jahren, Neal; Stone, Matthew D.; Udeshi, Namrata D.; Markowski, Todd W.; Witthuhn, Bruce A.; Shabanowitz, Jeffrey; Hunt, Donald F.; Olszewski, Neil E.

doi:10.1104/pp.112.208900

Cited by 27 publications

(24 citation statements)

References 35 publications

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“…However, there is experimental evidence showing the intracellular occurrence of N-GlcNAc proteins (i.e., potential cytoplasmic ENGase reaction products) in murine synapses (38)(39)(40). It was also clearly shown that at least some of the N-GlcNAc proteins are formed by the cytoplasmic ENGase activity in plants (20). These results indicate that formation of N-GlcNAc proteins by ENGase may not be a rare event in cells.…”

Section: Discussionmentioning

confidence: 92%

“…Although the ENGase is believed to be involved in the catabolism of cytosolic free oligosaccharides, recent evidence shows that it can deglycosylate glycoproteins in vivo to generate N-GlcNAc-bearing proteins in Arabidopsis thaliana (20), raising the possibility that this enzyme may also act as a deglycosylation enzyme for misfolded glycoproteins in the cytosol (21,22) (Fig. 1A).…”

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confidence: 99%

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Endo-β- N -acetylglucosaminidase forms N -GlcNAc protein aggregates during ER-associated degradation in Ngly1-defective cells

Huang

Harada

Hosomi

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

108

115

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The cytoplasmic peptide:N-glycanase (PNGase; Ngly1 in mice) is a deglycosylating enzyme involved in the endoplasmic reticulum (ER)-associated degradation (ERAD) process. The precise role of Ngly1 in the ERAD process, however, remains unclear in mammals. The findings reported herein, using mouse embryonic fibroblast (MEF) cells, that the ablation of Ngly1 causes dysregulation of the ERAD process. Interestingly, not only delayed degradation but also the deglycosylation of a misfolded glycoprotein was observed in Ngly1 −/− MEF cells. The unconventional deglycosylation reaction was found to be catalyzed by the cytosolic endo-β-N-acetylglucosaminidase (ENGase), generating aggregation-prone N-GlcNAc proteins. The ERAD dysregulation in cells lacking Ngly1 was restored by the additional knockout of ENGase gene. Thus, our study underscores the functional importance of Ngly1 in the ERAD process and provides a potential mechanism underlying the phenotypic consequences of a newly emerging genetic disorder caused by mutation of the human NGLY1 gene.PNGase (Ngly1) | ENGase | protein aggregates | glycoprotein | ERAD

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Section: Discussionmentioning

confidence: 92%

mentioning

confidence: 99%

Endo-β- N -acetylglucosaminidase forms N -GlcNAc protein aggregates during ER-associated degradation in Ngly1-defective cells

Huang

Harada

Hosomi

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

108

115

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“…A. thaliana proteins bearing this modification were explained as the product of the reaction of the enzyme endo-β-N-acetylglucosaminidase on larger high mannose N-glycans [31]. In animal cells, endoglycosidases are encoded in the genome even if it is not clear if their activity is directed towards glycoproteins, glycopeptides, or free oligosaccharide [32,33].…”

Section: Sugar-peptide Analogs Of Csf114mentioning

confidence: 99%

Multi-Stage Mass Spectrometry Analysis of Sugar-Conjugated β-Turn Structures to be Used as Probes in Autoimmune Diseases

Gentilini

Auberger

Rentier

et al. 2016

J. Am. Soc. Mass Spectrom.

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Abstract. Synthetic sugar-modified peptides were identified as antigenic probes in the context of autoimmune diseases. The aim of this work is to provide a mechanistic study on the fragmentation of different glycosylated analogs of a synthetic antigenic probe able to detect antibodies in a subpopulation of multiple sclerosis patients. In particular the N-glucosylated type I′ β-turn peptide structure called CSF114(Glc) was used as a model to find signature fragmentations exploring the potential of multistage mass spectrometry by MALDI-LTQ Orbitrap. Here we compare the fragmentation of the glucosylated form of the synthetic peptide CSF114(Glc), bearing a glucose moiety on an asparagine residue, with less or non-immunoreactive forms, bearing different sugar-modifications, such as CSF114(GlcNAc), modified with a residue of N-acetylglucosamine, and CSF114[Lys 7 (1-deoxyfructopyranosyl)], this last one modified with a 1-deoxyfructopyranosyl moiety on a lysine at position 7. The analysis was set up using a synthetic compound specifically deuterated on the C-1 to compare its fragmentation with the fragmentation of the undeuterated form, and thus ascertain with confidence the presence on an Asn(Glc) within a peptide sequence. At the end of the study, our analysis led to the identification of signature neutral losses inside the sugar moieties to characterize the different types of glycosylation/glycation. The interest of this study lies in the possibility of applyimg this approach to the discovery of biomarkers and in the diagnosis of autoimmune diseases.

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“…During a study of antibodies expressed in transgenic plants MALDI-TOF analysis allowed the detection of fragments with masses consistent with N-GlcNAc modification [130]. Recently Arabidopsis thaliana proteins that are predicted to transit through the endoplasmic reticulum and Golgi were identified with such N-GlcNAc modifications [131]. During this study a mass labelling method was developed and used to study N-GlcNAc modification of two myrosinases (thioglucoside glucohydrolases, TGG), TGG1 and TGG2, that are also substituted by high-mannose (Man)-type glycans.…”

Section: Plant N-glycan-engasesmentioning

confidence: 99%

“…These data support the hypothesis that ENGases hydrolyze high-Man glycans to produce some of the N-GlcNAc modifications but also suggest that some modifications are generated by another mechanism. It was finally concluded that, since the modification was detected at only one site on each myrosinase, the production of the N-GlcNAc modification may be regulated [131]. This prompts the investigation of the anomeric configuration of the GlcNAc linkage, and also the occurrence of enzymes that transfer GlcNAc to consensus Asn-X-Ser/Thr sites, equivalent to the Haemophilus influenzae glycosyltransferase, able to transfer hexose (Gal or Glc) residues with no requirement for a lipid donor intermediate that has been identified recently [132].…”

Section: Plant N-glycan-engasesmentioning

confidence: 99%

Endo-N-Acetyl-β-D-Glucosaminidases and Peptide-N4-(N-acetyl-β-D-Glucosaminyl) Asparagine Amidases: More Than Just Tools

Karamanos¹

2013

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Abstract:Since the discovery of endo-N-acetyl-β-D-glucosaminidases (ENGase) and peptide-N4-(N-acetyl-β-D-glucosaminyl) asparagine amidases (PNGase) most of the published work described their use for structural studies. Less attention was given to the potential roles of those enzymes in the physiology of the cells/organisms they produced them. The scope of this review is firstly to analyse the data on the occurrence and characteristics of murein-, chitin-, and N-glycan-ENGases acting on GlcNAc-containing polymers in three structural families, namely murein, chitin, and N-glycosylproteins, and of PNGases, only acting on N-glycosylproteins, and secondly to discuss the biological roles of the enzymes in the producing cells. The analysis demonstrates the remarkable diversity of the enzymes, and simultaneously the interest of studying their substrate specificity and their structural features. Many examples illustrate the importance of the structure/function relationships studies. Diverse biological roles were anticipated, e.g. they are useful for feeding purposes, are implicated in pathogenesis processes, modulate the activity of macromolecules, and help in the destruction of misfolded proteins. Their effect can be direct or indirect, through the reaction products. Current knowledge only partially explains the biological roles of ENGases and PNGases, thus further studies are expected for determining novel possibilities and elucidating other cell pathways.

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Identification and Origin of N-Linked β-d-N-Acetylglucosamine Monosaccharide Modifications on Arabidopsis Proteins

Cited by 27 publications

References 35 publications

Endo-β- N -acetylglucosaminidase forms N -GlcNAc protein aggregates during ER-associated degradation in Ngly1-defective cells

Endo-β- N -acetylglucosaminidase forms N -GlcNAc protein aggregates during ER-associated degradation in Ngly1-defective cells

Multi-Stage Mass Spectrometry Analysis of Sugar-Conjugated β-Turn Structures to be Used as Probes in Autoimmune Diseases

Endo-N-Acetyl-β-D-Glucosaminidases and Peptide-N4-(N-acetyl-β-D-Glucosaminyl) Asparagine Amidases: More Than Just Tools

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