Identification of two discrete peptide: N-glycanases inOryzias latipes during embryogenesis

Seko, Akira; Kitajima, Ken; Isobe, Takashi; Inoue, Yasuo; Inoue, Sadako

doi:10.1093/glycob/9.9.887

Cited by 30 publications

(23 citation statements)

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“…germination) and the subsequent ontogenesis (e.g. post-germinative development) as we have shown for the embryonic development of fish (40). (F1, F2, and F3) and centrifugation method (C1, C2, and C3) were used to locate the expressed enzyme activity (see "Experimental Procedures").…”

Section: Discussionmentioning

confidence: 99%

“…14). (e) We were successful in identifying two discrete PNGases in Oryzias latipes during embryogenesis, and their physiological roles were discussed (15). Ever since we unveiled the presence of PNGase and proposed its functional significance in animal cells, an increasing number of reports have appeared to implicate a possible involvement of PNGase in a wide range of biological phenomena (18 -22).…”

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confidence: 99%

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Developmentally Regulated Expression of a Peptide:N-Glycanase during Germination of Rice Seeds (Oryza sativa) and Its Purification and Characterization

Chang

Kuo

Khoo

et al. 2000

Journal of Biological Chemistry

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Peptide:N-glycanase (PNGase; EC 3.5.1.52) activity was detected in dormant rice seeds (Oryza sativa) and the imbibed rice grains. Time-course studies revealed that the enzyme activity remained almost constant until about 30 h after imbibition in both of endosperm-and embryo tissue-containing areas, and started to increase only in growing germ part, reached a peak at about 3-day stage, followed by a gradual decrease concomitant with a sharp increase in the coleoptile. The specific activity increased about 6-fold at about 3-day stage. PNGase was purified to electrophoretic homogeneity from the extracts of germinated rice seeds at 24 h, and the apparent molecular weight of the purified enzyme, estimated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), was about 80,000. The purified enzyme was designated PNGase Os to denote its origin. The N-terminal sequence of the 10 residues was determined to be SYN-VASVAGL. The purified PNGase Os in SDS-PAGE appeared as a rather broad band, consistent with the presence of multiple glycoforms as indicated by chromatographic behavior on a Sephadex G-75 column. PNGase expressed in coleoptile under anoxia condition was also purified, and both of the purified enzymes were found to exhibit very similar, if not identical, electrophoretic mobility in SDS-PAGE. PNGase Os exhibited a broad pH-activity profile with an optimum of 4 -5 and, interestingly, was significantly inactivated by K ؉ and Na ؉ at near the physiological concentration, 100 mM. These results are discussed in relation to other work.-(N-acetyl-␤-Dglucosaminyl) asparagine amidase, EC 3.5.1.52) had only been known to occur in some plant seeds (1-5) and bacteria (6) and used as a useful reagent in a number of studies of structure and function of glycoproteins having N-linked glycan chains until we demonstrated for the first time its occurrence in the early embryos of Medaka fish, Oryzias latipes in 1991 (7). Following this discovery, we began to focus our interest on the physiological significance of this enzyme in living organisms because little attention had been paid to it. In view of the unique structural change in a given functional protein by converting the glycosylated asparagine residue to the aspartic acid residue upon de-N-glycosylation catalyzed by PNGase as the posttranslational remodification of protein, we anticipated that N-glycosylation of proteins by oligosaccharyltransferase and their de-N-glycosylation by PNGase constitute a basic biological mechanism of functioning within cells (8, 9). For understanding the biological meaning of the occurrence of PNGase in eukaryotes, we have been involved in a series of studies on animal-derived PNGases (10 -15) and reported the following. (a) PNGase activities were shown to occur in mammalianderived cell lines including human origin (10). (b) PNGase activities were detected ubiquitously in various organs and tissues of mouse (13). (c) PNGase was purified to homogeneity from the confluent stage of C3H mouse fibroblast L-929 cells and characterized to serve not only a...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Developmentally Regulated Expression of a Peptide:N-Glycanase during Germination of Rice Seeds (Oryza sativa) and Its Purification and Characterization

Chang

Kuo

Khoo

et al. 2000

Journal of Biological Chemistry

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“…The substrate used in this study was the glycoasparagine Asn(Man 4 GlcNAc 4 ) (GP-IVB; ref. 28) and was 14 C-labeded as described (27,29). The specific radioactivity of 14 C-labeled GP-IVB was 1.2 ϫ 10 5 cpm͞nmol.…”

Section: Methodsmentioning

confidence: 99%

Endo-β- N -acetylglucosaminidase, an enzyme involved in processing of free oligosaccharides in the cytosol

Suzuki

Yano

Sugimoto

et al. 2002

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

129

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Formation of oligosaccharides occurs both in the cytosol and in the lumen of the endoplasmic reticulum (ER). Luminal oligosaccharides are transported into the cytosol to ensure that they do not interfere with proper functioning of the glycan-dependent quality control machinery in the lumen of the ER for newly synthesized glycoproteins. Once in the cytosol, free oligosaccharides are catabolized, possibly to maximize the reutilization of the component sugars. An endo-␤-N-acetylglucosaminidase (ENGase) is a key enzyme involved in the processing of free oligosaccharides in the cytosol. This enzyme activity has been widely described in animal cells, but the gene encoding this enzyme activity has not been reported. Here, we report the identification of the gene encoding human cytosolic ENGase. After 11 steps, the enzyme was purified 150,000-fold to homogeneity from hen oviduct, and several internal amino acid sequences were analyzed. Based on the internal sequence and examination of expressed sequence tag (EST) databases, we identified the human orthologue of the purified protein.The human protein consists of 743 aa and has no apparent signal sequence, supporting the idea that this enzyme is localized in the cytosol. By expressing the cDNA of the putative human ENGase in COS-7 cells, the enzyme activity in the soluble fraction was enhanced 100-fold over the basal level, confirming that the human gene identified indeed encodes for ENGase. Careful gene database surveys revealed the occurrence of ENGase homologues in Drosophila melanogaster, Caenorhabditis elegans, and Arabidopsis thaliana, indicating the broad occurrence of ENGase in higher eukaryotes. This gene was expressed in a variety of human tissues, suggesting that this enzyme is involved in basic biological processes in eukaryotic cells.

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“…While the fish enzyme is believed to be of lysosomal origin [156], mammalian PNGase is a cytoplasmic enzyme and its optimal activity is at neutral pH [154]. A gene encoding cytoplasmic PNGase (PNG1) was identified in Saccharomyces cerevisiae and mapped to the left arm of chromosome XVI [157].…”

Section: Occurrence Characteristics and Functions Of Pngasesmentioning

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 of two discrete peptide: N-glycanases inOryzias latipes during embryogenesis

Cited by 30 publications

References 32 publications

Developmentally Regulated Expression of a Peptide:N-Glycanase during Germination of Rice Seeds (Oryza sativa) and Its Purification and Characterization

Developmentally Regulated Expression of a Peptide:N-Glycanase during Germination of Rice Seeds (Oryza sativa) and Its Purification and Characterization

Endo-β- N -acetylglucosaminidase, an enzyme involved in processing of free oligosaccharides in the cytosol

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

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