Processing Pathway Deduced from the Structures of N-Glycans in Carica papaya

Makino, Yasushi; Shimazaki, Atsuyuki; Omichi, Kaoru; Okada, Shoji; Hase, Sumihiro

doi:10.1093/oxfordjournals.jbchem.a022707

Cited by 10 publications

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“…[10][11][12][13] Sugar structures were confirmed mainly by comparing with elution positions of the standard PA-sugar chains on both size and reversed-phase HPLCs. Exoglucosidases, such as jack bean -mannosidase, and A. oryzae 1,2-mannosidase were used when the structure was uncertain.…”

Section: Methodsmentioning

confidence: 97%

“…Most of the standard PA-oligosaccharides were described in previous papers. [10][11][12][13] G3M9 from starfish eggs was kindly donated by T. Endo.…”

Section: Methodsmentioning

confidence: 99%

“…Normally, 2-3 mg of a dried sample was heated in 0.5 ml of hydrazine at 100 C for 10 h. Released sugar chains in the hydrazine solution were transferred to another tube, N-acetylated, and pyridylaminated according to a previously reported method. 10,11) Two to four leaves were used for the analysis of samples suppressed with 5 mM CST, 0.2 mM N-MeDNJ, and 1 mM DNJ. Pyridylaminated oligosaccharides were purified by phenol-chloroform extraction and passed through a small column (0:5 Â 1 cm) packed with Dowex 50 NH 4 þ , and analyzed by size-fractionation HPLC on a Shodex NH2P-50 column (2 Â 150 mm).…”

Section: Methodsmentioning

confidence: 99%

“…Left sides (A) are the chromatograms of control and right sides (B) are those of 5 mM CST. Sugar structures in some peaks were elucidated by comparing their elution positions with those of the standard PA-oligosaccharides [10][11][12][13] and glycosidase digestions (Table 1). Structures in parentheses were not confirmed by glycosidase digestion.…”

Section: )mentioning

confidence: 99%

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Glucose Trimming ofN-Glycan in Endoplasmic Reticulum Is Indispensable for the Growth ofRaphanus sativusSeedling (kaiware radish)

Mega

2005

Bioscience, Biotechnology, and Biochemistry

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Recently I found that glycosidase inhibitors such as castanospermine, deoxynojirimycin, swainsonine, 2-acetamindo 2,3-dideoxynojirimycin, and deoxymannojirimycin change the N-glycan structure of root glycoproteins, and that the glucosidase inhibitors castanospermine and deoxynojirimycin suppress the growth of Raphanus sativus seedlings (Mega, T., J. Biochem., 2004). The present study undertook to see whether the growth suppression is due to the inhibition of glucose trimming in endoplasmic reticulum (ER). The study, using three glucosidase inhibitors, castanospermine, Nmethyl deoxynojirimycin, and deoxynojirimycin, upon the growth of R. sativus foliage leaf, made clear that glucose trimming is indispensable for plant growth, because the inhibition of glucose trimming correlated with leaf growth. On the other hand, processing inhibition in the Golgi apparatus by other glycosidase inhibitors had little effect on plant growth, although Nglycan processing was disrupted depending on inhibitor specificity. These results suggest that N-glycan processing after glucosidase processing is dispensable for plant growth and cell differentiation.Key words: castanospermine; deoxynojirimycin; N-glycan; glycosidase inhibitors; Raphanus sativusThe functions of plant N-glycan remain unclarified, 1) although plant glycoproteins, glycosidases, and lectins have been well studied. To identify the functions of enzymes, two approaches are usually taken: the use of mutants lacking the target enzyme activity, and the use of enzyme inhibitors. The method using enzyme inhibitors is easy to apply and enables easy control of the degree of an effect by regulating the amount administered. But we must consider the effects of inhibitors on as yet unidentified enzymes or interactions in living organisms.It has been reported that the glucosidase I mutant of Arabidopsis cannot form seeds, 2) and produces a diminished amount of cellulose.3) The Arabidopsis glucosidase II mutant can grow with a radial swollen root and is heat labile.4) In a previous study, R. sativus seed was germinated in the presence of glycosidase inhibitors, and the effect of these inhibitors on the Nglycan structure in the root was examined. Structural analysis showed that glycosidase inhibitors inhibited N-glycan processing in both the ER and the Golgi apparatus, as shown in Scheme 1. 5) That is, glycosidase inhibitors inhibit the processing glycosidases according to inhibitor specificity. For example, dMNJ (deoxymannojirimycin) let almost all the N-glycans in a plant retain oligomannose-type structures, but, germination of seeds and growth were normal. Glucosidase inhibitors such as CST and DNJ inhibit glucosidase I and/or II of plants, and also plant growth. Usually in cultured cells these inhibitors influence the N-glycan structure but not growth.These glucosidase inhibitors are also known to inhibit sucrase and -glucosidase.6,7) It is reasonable to speculate that they inhibit one or more glucose related enzymes which lead to the growth inhibition, because (a) sucrose has an ...

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Section: Methodsmentioning

confidence: 97%

“…Most of the standard PA-oligosaccharides were described in previous papers. [10][11][12][13] G3M9 from starfish eggs was kindly donated by T. Endo.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: )mentioning

confidence: 99%

See 2 more Smart Citations

Glucose Trimming ofN-Glycan in Endoplasmic Reticulum Is Indispensable for the Growth ofRaphanus sativusSeedling (kaiware radish)

Mega

2005

Bioscience, Biotechnology, and Biochemistry

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“…Dried samples, mainly leaves of the plants (2-6 mg), were heated at 100 C for 10 h in 0.4 ml of hydrazine in a tube. Oligosaccharides released in hydrazine were N-acetylated, pyridylaminated, and analyzed by HPLC after purification by phenol-chloroform extraction and Dowex 50 NH 4þ filtration with H 2 O according to the conventional method used in Hase's laboratory, [17][18][19] and as also described in my previous papers.10,14) SF-HPLC was performed with an Asahipak NH2P-50 column (2:0 Â 150 mm) at a flow rate of 75 ml/min. RP-HPLC was performed with a Cosmosil 5C18-P column (1:5 Â 250 mm) at a flow rate of 150 ml/ min.…”

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confidence: 99%

Plant-TypeN-Glycans Containing Fucose and Xylose in Bryophyta (Mosses) and Tracheophyta (Ferns)

Mega

2007

Bioscience, Biotechnology, and Biochemistry

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The presence of typical plant-type N-glycans (eg, M3FX, Gn2M3FX, and Le a 2M3FX) in mosses, ferns, and other organisms was examined to determine which plant initially acquired glycosyltransferases to produce plant-type N-glycans during organic evolution. No M3FX-type N-glycan was detected in lichens (Cladonia humilis) or in any one of the three preland plants Enteromorpha prolifera, Ulva pertusa Kjellman, and Chara braunii Gmelin. In Bryophyta, M3FX-type Nglycan was detected at trace amounts in Anthocerotopsida (hornworts) and at certain amounts in Bryopsida (mosses), but not in Hepaticopsida (liverworts). Le a 2M3FX was detected in some Bryopsida of relatively high M3FX content. Most Tracheophyta (ferns and higher plants) contained the three typical M3FX-type glycans as the main N-glycans in different ratios. These results suggest that organisms acquired xylosyltransferase and fucosyltransferase during the development of mosses from liverworts, and that later all plants retained both enzymes. Bryopsida have also obtained galactosyltransferase and fucosyltransferase to synthesize the Le a antigen.Key words: N-glycan; fern; moss; M3FX Gn2M3FX and Le a M3FX; Le a antigenSince the structure of the oligosaccharide of pineapple stem bromelain was elucidated, 1) the determined fucoseand xylose-containing core structure has been found universally in plant glycoproteins. M3FX, consisting of three Man (M), two GlcNAc (Gn), and one each of Fuc (F) and Xyl (X) residues is the most common structure in plants. The oligosaccharides GnM3FX and Gn2M3FX, composed of one and two GlcNAc residues respectively attached to M3FX, are also found universally in plants. The oligosaccharide with the Lewis a antigen, Le a 2M3FX, in which galactose and fucose residues are attached to both GlcNAc residues of Gn2M3FX, is also a main N-glycan universally found in plants, [2][3][4][5] except for Brassicaceae plants such as Arabidopsis thaliana and Raphanus sativus (kaiware raddish). Accordingly, the processing of plant N-glycans in the Golgi apparatus as shown in Fig. 1 is assumed to be almost universal. 5,6) N-Glycan processing byglucosidases I and II in ER is important in plants, [7][8][9][10] whereas N-glycan processing in the Golgi apparatus has little effect on plant growth, as determined in the varous studies: Arabidopsis mutants that lack N-acetylglucosaminyltransferase I 11) or both fucosyltrasferase and xylosyltransferase activities 12) grow normally and produce normal seeds. Fucosyltransferase and xylosyltransferase knockout plants of Bryopsida (Physcomitrella patens) grow normally.13) In my study using glycosidase inhibitors on germination, the seedling and leaf growth of R. sativus showed that ER-glucosidase inhibition inhibited the growth of the plant, although the inhibition of N-glycan processing in Golgi by deoxymannojirimycin, swainsonine, and 1,2-dideoxy-2-acetamido-nojirimysin had no eminent effect except for a change in the N-glycan structure. 10,14) These results indicate that the final N-glycan structure is not directly as...

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