Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.1.52 (PNGase F)
 1,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous works have shown that glycoconjugates with terminal fucose (Fuc) are located in the primordial germ cells (PGCs) of some mammals and might play a role in the migration and adhesion processes during development. The aim of this work was to identify the terminal Fuc moieties of Xenopus PGCs by means of three Fuc-binding lectins: from asparagus pea (LTA), gorse seed (UEA-I), and orange peel fungus (AAA). The histochemical procedures were also carried out after deglycosylation pretreatments: beta-elimination with NaOH to remove O-linked oligosaccharides; incubation with PNGase F to remove N-linked carbohydrate chains; and incubation with alpha(1,2)- and alpha(1,6)-fucosidase. The PGCs were always negative for LTA and UEA-I, two lectins that have the highest affinity for Fuc alpha(1,2)-linked. However, the PGCs were strongly labeled with AAA, which preferentially binds to Fuc with alpha(1,3) or alpha(1,4) linkages and to Fuc alpha(1,6)-linked to the proximal N-acetylglucosamine. There was fainter labeling with AAA when the sections were preincubated with alpha(1,6)-fucosidase, but the labeling remained strong when the sections were pretreated with alpha(1,2)fucosidase. When the beta-elimination procedure was carried out, the PGC labeling with AAA was slight. If the PNGase F incubation was performed, the PGCs remained moderately positive for AAA. These data suggest that the Xenopus PGCs have Fuc moieties in O- and N-linked oligosaccharides, including Fuc alpha(1,6) linked to the innermost GlcNAc, and that the Fuc was not in alpha(1,2)-linkage.
 ...
PMID:Lectin histochemistry shows fucosylated glycoconjugates in the primordial germ cells of Xenopus embryos. 1253 32

The AAA ATPase p97 is a ubiquitin-selective molecular machine involved in multiple cellular processes, including protein degradation through the ubiquitin-proteasome system and homotypic membrane fusion. Specific p97 functions are mediated by a variety of cofactors, among them peptide N-glycanase, an enzyme that removes glycans from misfolded glycoproteins. Here we report the three-dimensional structure of the aminoterminal PUB domain of human peptide N-glycanase. We demonstrate that the PUB domain is a novel p97 binding module interacting with the D1 and/or D2 ATPase domains of p97 and identify an evolutionary conserved surface patch required for p97 binding. Furthermore, we show that the PUB and UBX domains do not bind to p97 in a mutually exclusive manner. Our results suggest that PUB domain-containing proteins constitute a widespread family of diverse p97 cofactors.
 ...
PMID:The PUB domain functions as a p97 binding module in human peptide N-glycanase. 1680 42

During endoplasmic reticulum-associated degradation, the multifunctional AAA ATPase p97 is part of a protein degradation complex. p97 associates via its N-terminal domain with various cofactors to recruit ubiquitinated substrates. It also interacts with alternative substrate-processing cofactors, such as Ufd2, Ufd3, and peptide:N-glycanase (PNGase) in higher eukaryotes. These cofactors determine different fates of the substrates and they all bind outside of the N-terminal domain of p97. Here, we describe a cofactor-binding motif of p97 contained within the last 10 amino acid residues of the C terminus, which is both necessary and sufficient to mediate interactions of p97 with PNGase and Ufd3. The crystal structure of the N-terminal domain of PNGase in complex with this motif provides detailed insight into the interaction between p97 and its substrate-processing cofactors. Phosphorylation of p97's highly conserved penultimate tyrosine residue, which is the main phosphorylation site during T cell receptor stimulation, completely blocks binding of either PNGase or Ufd3 to p97. This observation suggests that phosphorylation of this residue modulates endoplasmic reticulum-associated protein degradation activity by discharging substrate-processing cofactors.
 ...
PMID:Studies on peptide:N-glycanase-p97 interaction suggest that p97 phosphorylation modulates endoplasmic reticulum-associated degradation. 1749 50

The endoplasmic reticulum (ER) glycoprotein HMG-CoA reductase (HMGR) catalyzes the rate-limiting step in sterols biosynthesis. Mammalian HMGR is ubiquitinated and degraded by the proteasome when sterols accumulate in cells, representing the best example for metabolically controlled ER-associated degradation (ERAD). This regulated degradation involves the short-lived ER protein Insig-1. Here, we investigated the dislocation of these ERAD substrates to the cytosol en route to proteasomal degradation. We show that the tagged HMGR membrane region, HMG(350)-HA, the endogenous HMGR, and Insig-1-Myc, all polytopic membrane proteins, dislocate to the cytosol as intact full-length polypeptides. Dislocation of HMG(350)-HA and Insig-1-Myc requires metabolic energy and involves the AAA-ATPase p97/VCP. Sterols stimulate HMG(350)-HA and HMGR release to the cytosol concurrent with removal of their N-glycan by cytosolic peptide:N-glycanase. Sterols neither accelerate dislocation nor stimulate deglycosylation of ubiquitination-defective HMG(350)-HA((K89 + 248R)) mutant. Dislocation of HMG(350)-HA depends on Insig-1-Myc, whose dislocation and degradation are sterol independent. Coimmunoprecipitation experiments demonstrate sterol-stimulated association between HMG(350)-HA and Insig-1-Myc. Sterols do not enhance binding to Insig-1-Myc of HMG(350)-HA mutated in its sterol-sensing domain or of HMG(350)-HA((K89 + 248R)). Wild-type HMG(350)-HA and Insig-1-Myc coimmunoprecipitate from the soluble fraction only when both proteins were coexpressed in the same cell, indicating their encounter before or during dislocation, raising the possibility that they are dislocated as a tightly bound complex.
 ...
PMID:Dislocation of HMG-CoA reductase and Insig-1, two polytopic endoplasmic reticulum proteins, en route to proteasomal degradation. 1945 99

Glycoconjugates play roles in many physiological and pathological processes. Previous works have shown important functions mediated by glycans in spermatogenesis, and the carbohydrate composition of testis has been studied by several approaches, including lectin-histochemical methods. However, the testis of Xenopus laevis, an animal model extensively employed in biochemical, cell and developmental research, has not yet been analysed. The aim of this work was to carry out a histochemical study of the fucose (Fuc)-containing glycoconjugates of Xenopus testis by means of lectins, combined with deglycosylation pretreatments. Four Fuc-binding lectins were used: orange peel (Aleuria aurantia) lectin (AAL), gorse seed (Ulex europaeus) agglutinin-I (UEA-I), fresh water eel (Anguilla anguilla) agglutinin (AAA), and asparagus pea (Lotus tetragonolobus) agglutinin (LTA), each recognizing different forms of fucosylated glycans. Labelling with UEA-I, which preferably binds Fucalpha(1,2) containing oligosaccharides, did not show any appreciable staining. LTA, specific for Fucalpha(1,3), and AAA, which binds Fucalpha(1,2), labelled spermatocytes and spermatids, but no labelling was seen when the histochemical procedure was carried out after either beta-elimination (which removes O-linked oligosaccharides) or incubation with PNGase F (which removes N-linked oligosaccharides), suggesting that fucosylated glycans are of both N- and O-linked types. AAL, which has its highest affinity to Fucalpha(1,6), but also recognizes Fucalpha(1,2) and Fucalpha(1,3), labelled the whole testis, and the staining remained when the histochemical method was performed after either beta-elimination or incubation with PNGase F. Labelling with AAL could be explained by the fact that this lectin could be binding to diverse fucosylated glycans in N- and O-glycans, and even in glycolipids. The importance of these glycans is discussed.
 ...
PMID:Identification of fucosylated glycoconjugates in Xenopus laevis testis by lectin histochemistry. 2061 42

Dendritic cells use a specialized pathway called cross-presentation to activate CD8+ T cells by presenting peptides from exogenous protein antigens on major histocompatibility complex class I molecules. Considerable evidence suggests that internalized antigens cross endocytic membranes to access cytosolic proteasomes for processing. The mechanism of protein dislocation represents a major unsolved problem. Here we describe the development of a sensitive reporter substrate, an N-glycosylated variant of Renilla luciferase fused to the Fc region of human IgG1. The luciferase variant is designed to be enzymatically inactive when glycosylated, but active after the asparagine to aspartic acid conversion that occurs upon deglycosylation by the cytosolic enzyme N-glycanase-1. The generation of cytosolic luminescence depends on internalization, deglycosylation, the cytosolic AAA-ATPase VCP/p97, and the cytosolic chaperone HSP90. By incorporating a T cell epitope into the fusion protein, we demonstrate that antigen dislocation into the cytosol is the rate limiting step in cross-presentation.
 ...
PMID:A novel probe to assess cytosolic entry of exogenous proteins. 3008 32