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1.
CMP-N-acetylneuraminate hydroxylase was isolated from mouse liver high speed supernatant with a yield of 0.4% and an apparent 1000-fold purification. The enzyme is a monomeric protein with a molecular weight of 66 kDa, as determined by gel filtration and SDS-PAGE. The hydroxylase system was reconstituted with Triton X-100-solubilized mouse liver microsomes and purified soluble or microsomal forms of cytochrome b5 reductase and cytochrome b5. The systems were characterized in detail and kinetic parameters for each system were determined.Abbreviations Neu5Ac
N-acetyl--d-neuraminic acid
- Neu5Gc
N-glycoloyl--d-neuraminic acid
- CMP-Neu5Ac
cytidine-5-monophospho-N-acetylneuraminic acid
- CMP-Neu5Gc
cytidine-5-monophospho-N-glycoloylneuraminic acid
- TCA
trichloroacetic acid
- Chaps
3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulphonate
- SOD
superoxide dismutase
Enzymes: CMP-N-acetylneuraminate: NADH oxidoreductase (N-acetyl hydroxylating) (E.C. 1.14.13.45), CMP-Neu5Ac hydroxylase; NADH: cytochrome b5 oxidoreductase (E.C. 1.6.2.2), cytochrome b5 reductase; hydrogen peroxide: hydrogen peroxide oxidoreductase, catalase (E.C. 1.11.1.6); superoxide:superoxide oxidoreductase (E.C. 1.15.1.1), superoxide dismutase.This paper is dedicated to Professor Harry Schachter on the occasion of his 60th birthday. 相似文献
2.
We demonstrate that 9-amino-NeuAc transferred to asialo-1-acid glycoprotein resists cleavage by bacterial, viral and mammalian sialidases. This is the first synthetic sialic acid analogue, which can be activated and transferred to glycoprotein, but is not a sialidase (EC 3.2.1.18) substrate.Abbreviations HPLC
high performance liquid chromatography
- BSA
bovine serum albumin
- NeuAc
N-acetyl-d-neuraminic acid, 5-acetamido-3,5-dideoxy-d-glycero-d-galacto-non-2-ulosonic acid
- 9-Amino-NeuAc
9-amino-5-N-acetyl-d-neuraminic acid, 5-acetamido-9-trideoxy-d-glycero-d-galacto-non-2-ulosonic acid
- CMP-NeuAc
cytidine-5-monophospho-N-acetyl-d-neuraminic acid
- CMP-9-amino-NeuAc
cytidine-5-monophospho-9-amino-5-N-acetyl-d-neuraminic acid
- 9-azido-NeuAc
5-acetamido-9-azido-3,5,9-trideoxy-d-glycero-d-galacto-non-2-ulosonic acid.
Enzymes EC 3.2.1.18
sialidase, acylneuraminylhydrolase
- EC 2.4.99.1
Galß1-4GlcNAc a(2-6)-sialytransferase 相似文献
3.
The sialic acid analogue,N-acetyl-4-deoxy-neuraminic acid, is readily activated by CMP-sialic acid synthase from bovine brain. We also show that sialyl-transfer from CMP-N-acetyl-4-deoxy-neuraminic acid to asialo-
1-acid glycoprotein is achieved at a high rate using Gal1-4GlcNAc (2.6)-sialyltransferase from rat liver.In contrast toVibrio cholerae sialidase, fowl plague virus sialidase liberates boundN-acetyl-4-deoxy-neuraminic acid from the glycoprotein. Thus, as opposed to the general view, the action of neither synthase nor transferase depends on the presence of the hydroxy group at C-4 ofN-acetylneuraminic acid.Abbrevations BSA
bovine serum albumin
- DTE
dithioerythritol
- HPLC
high performance liquid chromatography
- NeuAc
N-acetyl-d-neuraminic acid
- 4-deoxy-NeuAc
N-acetyl-4-deoxy-d-neuraminic acid
- 4-epi-NeuAc
4-acetamido-3,5-dideoxy-d-glycero-d-talononulosonic acid
- CMP-NeuAc
Cytidine-5-monophospho-N-acetylneuraminic acid
- CMP-4-deoxy-NeuAc
Cytidine-5-monophospho-N-acetyl-4-deoxy-neuraminic acid
- FPV-sialidase
Fowl plague virus sialidase
- VCN
Vibrio cholerae neuraminidase 相似文献
4.
The purification and characterisation of viral, bacterial and mammalian sialidases (EC 3.2.1.18, neuraminidases, neuraminosylglycohydrolases) prompted a search for a colorimetric technique to localize the enzymes on electropherograms. The 5-bromo substituted indol-3-yl -ketoside of 5-N-acetyl-d-neuraminic acid (BIN), the synthesis of which is described here, seemed to be the appropriate substrate, because of its relative ease of enzymatic hydrolysis to 5-N-acetyl-d-neuraminic acid and 5-bromoindoxyl. The latter is readily transformed to the insoluble, intensely coloured 5,5-dibromo-indigo. This precipitates and can be seen readily at the sites of enzymatic activity. The new substrate is of definitive advantage as it provides a simple and direct method for the demonstration of sialidases without the need of a coupling reaction.Abbreviations Neu5Ac
5-N-acetyl-d-neuraminic acid
- BIN
5-bromo-indol-3-yl -ketoside ofN-acetyl-d-neuraminic acid 相似文献
5.
Pascal Reboul Pascal George Delphine Miquel Pierre Louisot Pierre Broquet 《Glycoconjugate journal》1996,13(1):69-79
When treated with retinoic acidin vivo, C6 glioma cells show an enhancement of CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase activity. A 300kDa glycoprotein was detected by lectin affinoblotting in retinoic acid-treated C6 cells which stained weakly or not at all in control cells. Comparative studies with different lectins demonstrated that this glycoprotein contains 2,3 Neu5Ac Gal-GalNAc O-glycan moieties. Cultures in the presence of an inhibitor of O-glycan synthesis (N-acetylgalactosaminide -O-benzyl) demonstrated that enhancement of staining of the 300 kDa glycoprotein was not due to the increase of the 2,3 sialyltransferase but to thede novo synthesis of the polypeptide chain of this glycoprotein.Abbreviations RA
retinoic acid
- Neu5Ac
N-acetylneuraminic acid
- CMP-Neu5Ac
cytidine 5 monophosphosialate
- 2,3 ST
CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase
- GalNAc-O-benzyl
N-acetylgalactosaminide -O-benzyl
- Gal1-3GalNAc-O-benzyl
Galactosyl 1-3N-acetylgalactosaminide -O-benzyl
- TBS
Tris-HCl buffer 50mm pH 7.5 containing NaCl 0.15m and Tween 20 0.05%
- B1 buffer
TBS containing MgCl2 1mm, MnCl2 1mm and CaCl2 1mm 相似文献
6.
Gerd Reuter Roland Schauer Reginaldo Prioli Miercio E A Pereira 《Glycoconjugate journal》1987,4(4):339-348
In the culture supernatant ofTrypanosoma rangeli, strain El Salvador, a sialidase was present with an activity of 0.1 U/mg protein as determined with the 4-methylumbelliferyl glycoside of -N-acetylneuraminic acid as substrate. This enzyme was purified about 700-fold almost to homogeneity by gel chromatography on Sephadex G-100 and Blue Sepharose, and affinity chromatographies on 2-deoxy-2,3-didehydroneuraminic acid and horse submandibular gland mucin, both immobilized on Sepharose. The pH optimum is at 5.4–5.6, and the molecular weight was determined by gel chromatography, high performance liquid chromatography and sodium dodecyl sulphate gel electrophoresis to be 70 000. The substrate specificity of the enzyme is comparable to bacterial, viral and mammalian sialidases with cleavage rates for the following substrates in decreasing order:
N-acetylneuraminyl-(2–3)-lactose>
N-glycoloylneuraminy-(2–3)-lactose>
N-acetylneuraminyl-(2–6)-lactose >sialoglycoproteins>gangliosides>9-O-acetylated sialoglycoproteins.4-O-Acetylated derivatives are resistant towards the action of this sialidase. The enzyme activity can be inhibited by 2-deoxy-2,3-didehydro-N-acetylneuraminic acid, Hg2+ ions, andp-nitrophenyloxamic acid; it is not dependent on the presence of Ca2+ Mn2+ or Mg2+ ions.Abbreviations BSA
bovine serum albumin
- BSM
bovine submandibular gland mucin
- CMP
cytidine monophosphate
- EDIA
ethylenediaminetetraacetic acid
- ESM
equine submandibular gland mucin
- HEPES
N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid
- HPLC
high performance liquid chromatography
- Lac
lactose
- MU-Neu5Ac
4-methylumbelliferyl glycoside of -N-acetylneuraminic acid
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid
- Neu4Ac5Gc
N-glycoloyl-4-O-acetylneuraminic acid
- Neu2en
2-deoxy-2,3-didehydroneuraminic acid
- Neu5Gc
N-glycoloylneuraminic acid
- PMSF
phenylmethylsulfonyl fluoride
- PSM
pig submandibular gland mucin
- SDS
sodium dodecyl sulfate
- Tris
tris-(hydroxymethyl)aminomethane
Dedicated to Professor Dr. Heinz Mühlpfordt on the occasion of his 65th birthday. 相似文献
7.
The inhibitory effect of various compounds on the activities of four types of rat sialidase was investigated. 2-Deoxy-2,3-dehydro-N-acetylneuraminic acid andN-acetylneuraminic acid were competitive inhibitors for the sialidases. The former was effective against cytosolic sialidase and intralysosomal sialidase more than two membrane-associated sialidases I and II, the latter being a much weaker inhibitor. A heavy metal ion such as Cu2+ (1mm) and thiol-modifying 4-hydroxymercuribenzoate (50 µm) caused complete inhibition of the activities of cytosolic sialidase and membrane sialidase I, while no decrease in the activities of intralysosomal sialidase and membrane sialidase II was observed. When 4-nitrophenyloxamic acid and siastatin B, inhibitors of bacterial sialidases, and synthetic thioglycoside GM3 analogue Neu5Ac-s-(2-6)Gal(1-4)Glc(1-1) ceramide, an inhibitor of influenza virus sialidase, were tested, they did not affect any activity of the rat sialidases. By the differential effect of these inhibitors, the four types of rat sialidase could be discriminated from one another and furthermore from viral and bacterial sialidases.Abbreviations Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-dehydro-N-acetylneuraminic acid
- 4MU-Neu5Ac
4-methylumbelliferyl--N-acetyl-d-neuraminic acid 相似文献
8.
A Gal1-4GlcNAc (2-6)-sialyltransferase from human liver was purified 34 340-fold with 18% yield by dye chromatography on Cibacron Blue F3GA and cation exchange FPLC. The enzyme preparation was free of other sialyltransferases. It did not contain CMP-NeuAc hydrolase, protease, or sialidase activity, and was stable at –20°C for at least eight months. The donor substrate specificity was examined with CMP-NeuAc analogues modified at C-5 or C-9 of theN-acetylneuraminic acid moiety. Affinity of the human enzyme for parent CMP-NeuAc and each CMP-NeuAc analogue was substantially higher than the corresponding Gal1-4GlcNAc (2-6)-sialyltransferase from rat liver.Abbreviations FPLC
fast protein liquid chromatography
- NeuAc
5-N-acetyl-d-neuraminic acid
- 9-amino-NeuAc
5-acetamido-9-amino-3,5,9-trideoxy-d-glycero-2-nonulosonic acid
- 9-acetamido-NeuAc
5,9-diacetamido-3,5,9-trideoxy-d-glycero--d-2-nonulosonic acid
- 9-benzamido-NeuAc
5-acetamido-9-benzamido-3,5,9-trideoxy-d-glycero--d-galacto-2-nonulosonic acid
- 9-fluoresceinyl-NeuAc
9-fluoresceinylthioureido-NeuAc
- 5-formyl-Neu
5-formyl--d-neuraminic acid
- 5-aminoacetyl-Neu
5-aminoacetyl--d-neuraminic acid
- CMP-NeuAc
cytidine-5-monophospho-N-acetylneuraminic acid
- GM1
Gal1-3GalNAc1-4(NeuAc2-3)Gal1-4Glc-ceramide
- ST
sialyltransferase
- DTE
1,4-dithioerythritol
Enzyme: Gal1-4GlcNAc (2-6)-sialyltransferase, EC 2.4.99.1. 相似文献
9.
Cornelis H. Hokke Jos G. M. van der Ven Johannis P. Kamerling Johannes F. G. Vliegenthart 《Glycoconjugate journal》1993,10(1):82-90
Incubation of synthetic Man\1-4GlcNAc-OMe, GalNAc1-4GlcNAc-OMe, Glc1-4GlcNAc-OMe, and GlcNAc1-4GlcNac-OMe with CMP-Neu5Ac and rat liver Gal1-4GlcNAc (2-6)-sialyltransferase resulted in the formation of Neu5Ac2-6Man1-4GlcNAc-OMe, Neu5Ac2-6GalNAc1-4GlcNAc-OMe, Neu5Ac2-6Glc1-4GlcNAc-OMe and Neu5Ac2-6GlcNAc1-4GlcNAc-OMe, respectively. Under conditions which led to quantitative conversion of Gal1-4GlcNAc-OEt into Neu5Ac2-6Gal1-4GlcNAc-OEt, the aforementioned products were obtained in yields of 4%, 48%, 16% and 8%, respectively. HPLC on Partisil 10 SAX was used to isolate the various sialyltrisaccharides, and identification was carried out using 1- and 2-dimensional 500-MHz1H-NMR spectroscopy.Abbreviations 2D
2-dimensional
- CMP
cytidine 5-monophosphate
- CMP-Neu5Ac
cytidine 5-monophospho--N-acetylneuraminic acid
- COSY
correlation spectroscopy
- DQF
double quantum filtered
- HOHAHA
homonuclear Hartmann-Hahn
- MLEV
composite pulse devised by M. Levitt
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid 相似文献
10.
The natural sialidase ofClostridium septicum was purified and characterized in parallel with the recombinant enzyme expressed byEscherichia coli. The two enzymes exhibit almost identical properties. The maximum hydrolytic activity was measured at 37 °C in 60mm sodium acetate buffer, pH 5.3. Glycoproteins like fetuin and saponified bovine submandibular gland mucin, most of them having (2-6) linked sialic acids, are preferred substrates, while sialic acids from gangliosides, sialyllactoses, or the (2-8) linked sialic acid polymer (colominic acid) are hydrolysed at lower rates. (2-3) Linkages are more rapidly hydrolysed than (2-6) bonds of sialyllactoses. The cleavage rate is markedly reduced by O-acetylation of the sialic acid moiety. These properties are similar to those of other secreted clostridial sialidases. The enzyme exists in mono-, di- and trimeric forms, the monomer exhibiting a molecular mass of 125 kDa, which is close to the protein mass of 111 kDa deduced from the nucleotide sequence of the cloned gene.Abbreviations BSM
bovine submandibular gland mucine
- CMM
cooked meat medium
- EDTA
ethylenediaminetetraacetic acid
- FPLC
fast performance liquid chromatography
- LB
Luria-Bertani
- MU-Neu5Ac
4-methylumbelliferyl--d-N-acetylneuraminic acid
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid
- Neu4,5Ac2
N-acetyl-4-O-acetylneuraminic acid
- pI
isoelectric point
- SDS
sodium dodecyl sulfate 相似文献
11.
The conformational properties of the oligosaccharide chain of GM1 ganglioside containingN-glycolyl-neuraminic acid, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer, were studied through NMR nuclear Overhauser effect investigations on the monomeric ganglioside in dimethylsulfoxide, and on mixed micelles of ganglioside and dodecylphosphocholine in water. Several interresidual contacts for the trisaccharide core--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-were found to fix the relative orientitation of the three saccharides, while the glycosidic linkage of the terminal -Gal-was found to be quite mobile as the -Gal-(1-3)--GalNAc-disaccharide exists in different conformations. These results are similar to those found for two GM1 gangliosides containingN-acetyl-neuraminic acid and neuraminic acid [1].Abbreviations Ganglioside nomenclature is in accordance with Svennerholm [23] and the IUPAC-IUB Recommendations [24] GM3(Neu5Ac)
II3Neu5AcLacCer, -Neu5Ac-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer
- GM3(Neu5Gc)
II3Neu5GcLacCer, -Neu5Gc-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer
- GM1(Neu5Ac)
II3Neu5AcGgOse4Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer
- GM1(Neu5Gc)
II3Neu5GcGgOse4Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer
- GM1(Neu)
II3NeuGgOse4Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu-(2-3)]--Glc-(1-1)-Cer
- GD1a
IV3Neu5AcII3Neu5AcGgOse4Cer, -Neu5Ac-(2-3)--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer
- GalNAc-GD1a
IV4GalNAcIV3Neu5AcII3Neu5AcGgOse4Cer, -GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer
- Neu
neuraminic acid
- Neu5Ac
N-acetyl-neuraminic acid
- Neu5Gc
N-glycolyl-neuraminic acid
- Cer
ceramide 相似文献
12.
Dong Gyun Kang Chang Sup Kim Hyung Joon Cha 《Biotechnology and bioengineering》2019,116(11):2815-2822
Recombinant glycoproteins expressed in Chinese hamster ovary (CHO) cells contain two forms of sialic acids; N-acetylneuraminic acid (Neu5Ac) as a major type and N-glycolylneuraminic acid (Neu5Gc) as a minor type. The Neu5Gc glycan moieties in therapeutic glycoproteins can elicit immune responses because they do not exist in human. In the present work, to reduce Neu5Gc levels of recombinant glycoproteins from CHO cell cultures, we coexpressed cytidine-5′-monophosphate-sialic acid transporter (CMP-SAT) that is an antiporter and transports cytosolic CMP-sialic acids (both forms) into Golgi lumen. When human erythropoietin was used as a target human glycoprotein, coexpression of CMP-SAT resulted in a significant decrease of Neu5Gc level by 41.4% and a notable increase of Neu5Ac level by 21.2%. This result could be reasonably explained by our hypothesis that the turnover rate of Neu5Ac to Neu5Gc catalyzed by CMP-Neu5Ac hydroxylase would be reduced through facilitated transportation of Neu5Ac into Golgi apparatus by coexpression of CMP-SAT. We confirmed the effects of CMP-SAT coexpression on the decrease of Neu5Gc level and the increase of Neu5Ac level using another glycoprotein human DNase I. Therefore, CMP-SAT coexpression might be an effective strategy to reduce the levels of undesired Neu5Gc in recombinant therapeutic glycoproteins from CHO cell cultures. 相似文献
13.
Dorothee Heuermann Peter Roggentin Reinhard G. Kleineidam Roland Schauer 《Glycoconjugate journal》1991,8(2):95-101
The sialidase secreted byClostridium chauvoei NC08596 was purified to apparent homogeneity by ion-exchange chromatography, gel filtration, hydrophobic interaction-chromatography, FPLC ion-exchange chromatography, and FPLC gel filtration. The enzyme was enriched about 10 200-fold, reaching a final specific activity of 24.4 U mg–1. It has a relatively high molecular mass of 300 kDa and consists of two subunits each of 150 kDa. The cations Mn2+, Mg2+, and Ca2+ and bovine serum albumin have a positive effect on the sialidase activity, while Hg2+, Cu2+, and Zn2+, chelating agents and salt decrease enzyme activity. The substrate specificity, kinetic data, and pH optimum of the enzyme are similar to those of other bacterial sialidases.Abbreviations FPLC
fast protein liquid chromatography
- NCTC
National Collection of Type Cultures
- ATCC
American Type Culture Collection
- MU-Neu5Ac
4-methylumbelliferyl--d-N-acetylneuraminic acid
- buffer A
0.02m piperazine, 0.01m CaCl2, pH 5.5
- buffer B
0.02m piperazine, 0.01m CaCl2, 1.0m NaCl, pH 5.5
- buffer C
0.1m sodium acetate, 0.01m CaCl2, pH 5.5
- SDS
sodium dodecyl sulfate
- PAGE
polyacrylamide gel electrophoresis
- Neu5Ac
N-acetylneuraminic acid
- BSM
bovine submandibular gland mucin
- GD1a
IV3Neu5Ac, II3Neu5Ac-GgOse4Cer
- GM1
II3Neu5Ac-GgOse4Cer
- MU-Neu4,5Ac2
4-methylumbelliferyl--d-N-acetyl-4-O-acetylneuraminic acid
- TLC
thin-layer chromatography
- HPTLC
high performance thin-layer chromatography
- EDTA
ethylenediamine tetraacetic acid
- EGTA
ethylene glycol bis(2-aminoethyl-ethen)-N,N,N,N-tetraacetic acid
- BSA
bovine serum albumin
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid
- IEF
isoelectric focusing
- IEP
isoelectric point 相似文献
14.
Puente-Polledo L Reglero A González-Clemente C Rodríguez-Aparicio LB Ferrero MA 《Glycoconjugate journal》1998,15(9):855-861
The capsular polysaccharide of Pasteurella haemolytica A2 consists of a linear polymer of N-acetylneuraminic acid (Neu5Ac) with (2–8) linkages. When the bacterium was grown at 37°C for 90 h in 250 ml shake flasks at 200 rpm in Brain heart infusion broth (BHIB), it accumulated, attaining a level of 60 g/ml. Release of this polymer was strictly regulated by the growth temperature, and above 40° no production was detected. The pathway for the biosynthesis of this sialic acid capsular polymer was also examined in P. haemolytica A2 and was seen to involve the sequential presence of three enzymatic activities: Neu5Ac lyase activity, which synthesizes Neu5Ac by condensation of N-acetyl-D-mannosamine and pyruvate with apparent Km values of 91 mM and 73 mM, respectively; a CMP-Neu5Ac synthetase, which catalyzes the production of CMP-Neu5Ac from Neu5Ac and CTP with apparent Km values of 2 mM and 0.5 mM, respectively, and finally a membrane-associated polysialyltransferase, which catalyzes the incorporation of sialic acid from CMP-Neu5Ac into polymeric products with an apparent CMP-Neu5Ac Km of 250 M. 相似文献
15.
Takashi Angata Shinobu Kitazume Takaho Terada Ken Kitajima Sadako Inoue Frederic A. Troy II Yasuo Inoue 《Glycoconjugate journal》1994,11(5):493-499
A novel glycosyltransferase which catalyses transfer of deaminated neuraminic acid, KDN (2-keto-3-deoxy-d-glycero-d-galacto-nononic acid) from CMP-KDN to the non-reducing termini of oligo-polysialyl chains of polysialoglycoprotein (PSGP), was discovered in the ovary of rainbow trout (Oncorhynchus mykiss). The KDN-transferase activity was optimal at neutral pH, and stimulated 2 to 2.5-fold by 2–5mm Mg2+ or Mn2+. Expression of KDN-transferase was developmentally regulated in parallel with expression of the 2 8-polysialytransferase, which catalyses synthesis of the oligo-polysialyl chains in PSGP. Incorporation of the KDN residues into the oligo-polysialyl chains prevented their further elongation, resulting in capping of the oligo-polysialyl chains. This is the first example of a glycosyltransferase that catalyses termination of 2 8-polysialylation in glycoproteins.Abbreviations KDN
2-keto-3-deoxy-d-glycero-d-galacto-nononic acid or naturally occurring deaminated neuraminic acid
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ge
N-glycolylneuraminic acid
- CMP-KDN
cytidine 5-(3-deoxy-d-glycero-d-galacto-2-nonulosonic phosphate) or cytidine 5-KDN phosphate
- CMP-NeuAc
cytidine 5-Neu5Ac phosphate; oligo-polySia, oligo- and/or polysialic acid
- PSGP
rainbow trout egg polysialoglycoprotein comprising 2 8-linked oligo- polyNeu5Gc
- PSGP (low Sia)
a precursor of PSGP present at early stages of oogenesis which contains mostly the disialyl group, Sia2 8Sia2 6-
- *K-PSGP
[14C]KDN-labelled PSGP obtained by incubating PSGP and CMP-[14C]KDN with the immature cortical vesicle fraction P1 containing KDN-transferase
- *A-PSGP
[14C]Neu5Ac-labelled PSGP obtained by incubating PSGP and CMP-[14C]Neu5Ac with the P1 fraction
-
A-*K-PSGP andK-*K-PSGP
the products obtained after incubating *K-PSGP with P1 fraction and unlabelled CMP-Neu5Ac or CMP-KDN, respectively
- *K-PSGP
cho
,A-*K-PSGP
cho
, andK-*K-PSGP
cho
mixture of oligosaccharide alditols obtained by alkaline borohydride treatment of *K-PSGP,A-*K-PSGP, and K-*K-PSGP, respectively
- *A-PSGP
cho
a mixture of oligosaccharide alditols obtained by alkaline borohydride treatment of [14C]Neu5Ac-labelled PSGP
- Endo-N
endo-N-acylneuraminidase
- DP
degree of polymerization
- GLC
gas-liquid chromatography
- HPLC
high performance liquid chromatography
- TLC
thin layer chromatography 相似文献
16.
The regional difference in the carbohydrate components of the ductus epididymis epithelium of a lizard was delineated by means of 13 lectins. Basal cells expressed only N-acetylglucosamine (GlcNAc). Throughout the ductus, the secretory cells showed oligosaccharides with terminal N-acetylneuraminic acid (Neu5Ac)(2,6)galactose (Gal)/N-acetylgalactosamine (GalNAc) and internal mannose (Man) and/or glucose (Glc) in the whole cytoplasm, oligosaccharides terminating in Neu5Ac(2,6)Gal(1,3)GalNAc, Neu5Ac(2,6)Gal (1,4)GlcNAc, GalNAc, GlcNAc, and fucose (Fuc) in the supra-nuclear zone, and also glycans terminating in Neu5Ac(2,3)Gal (1,4)GlcNAc, Neu5Ac(2,6)Gal(1,3)GalNAc, Gal (1,4)GlcNAc on the luminal surface. In the caput and corpus regions, the supra-nuclear cytoplasm was characterized by terminal Gal(1,4)GlcNAc and GalNAc, the luminal surface by GalNAc and Gal. The Golgi zone, showing oligosaccharides with terminal Neu5Ac(2,3)Gal (1,4)GlcNAc, Neu5Ac(2,6)Gal (1,3)GalNAc, Neu5Ac(2,6)Gal (1,4)GlcNAc, and internal GlcNAc, expressed terminal Gal (1,4)GlcNAc and GalNAc in the caput, and terminal GalNAc in the corpus. The granules showed all the investigated carbohydrates in their peripheral zone except terminal GalNAc and Fuc, whereas internal Man/Glc and terminal Gal were expressed in the central core, and Fuc throughout the ductus, terminal GlcNAc in the caput and corpus, and terminal GalNAc only in the corpus. 相似文献
17.
Summary N-Glycolylneuraminic acid (Neu5Gc) has been prepared by enzymatic hydrolysis of its -(28) linked homopolymer. The rate of hydrolysis of the natural poly -(28)-(Neu5Ac) and the semi-synthetic poly -(28)-(Neu5Gc) were compared with the neuraminidases fromClostridium perfringens andVibrio cholerae. The natural Neu5Ac polysaccharide was a better substrate for both enzymes. For comparison, acid hydrolysis of the two polysaccharides showed extensive degradation. 相似文献
18.
We have investigated the activity of CMP-Neu5Ac:Gal\1-3GalNAc -2,3-sialyltransferase (EC 2.4.99.4) in FR3T3 cells transformed by the Ha-ras oncogene in which we have previously demonstrated the higher expression of the -galactosidase -2,6-sialyltransferase (EC 2.4.99.1) [21]. We demonstrate that the presence of the activatedras gene decreases the activity of this specific -2,3-sialyltransferase fourfold. According to the kinetic parameters and to mixing experiments, we can assume that this decreased enzymatic activity reflects a decrease in the number of activeO-glycan -2,3-sialyltransferase polypeptides inras-transformed cells. However, no change in the binding of Peanut agglutinin was observed on the cell surface ofras-transformed FR3T3 suggesting that no change in the sialylation ofO-glycan core 1 appeared in these cells, although the activity of the -2,3-sialyltransferase was decreased.Abbreviations -2,3-ST(O)
CMP-Neu5Ac:Gal1-3GalNAc-R -2,3-sialyltransferase
- -2,3-ST(N/O)
CMP-Neu5Ac:Gal1-3/4GlcNAc-R -2,3-sialyltransferase
- -2,6-ST(N)
CMP-Neu5Ac:Gal1-4GlcNAc-R -2,6-sialyltransferase
- -2,6-ST(O)I
CMP-Neu5Ac:R-GalNAc(1-O)Ser -2,6-sialyltransferase
- -2,6-ST(O)II
CMP-Neu5Ac:Neu5Ac2-3Gal1-3GalNAc-R -2,6-sialyltransferase
- ASFet
asialofetuin
- FR3T3
Fisher rat fibroblast
- FRras
Ha-ras-transfected FR3T3 fibroblasts
- NaCl/Pi
sodium phosphate 10mm, NaCl 0.15m, pH 7.4, buffer
-
pNp
p-nitrophenol 相似文献
19.
As a precursor for the chemical synthesis of sialylated oligosaccharides, the trisaccharide glycoside Neu5Ac (2-8)Gal(1-4)GlcNAc(1-O)-pent-4-ene was synthesized starting from GlcNAc(1-O)-pent-4-ene, UDP-glucose andN-acetylneuraminic acid in a one pot reaction employing galactosyltransferase and (2-6)sialyl-transferase in a complete cofactor regeneration system.Abbreviations Neu5Ac
N-acetylneuraminic acid
- CMP-Neu5Ac
cytidine 5-monophosphosialate
- CMP
cytidine 5-monophosphate
- CDP
cytidine 5-diphosphate
- CTP
cytidine 5-triphosphate
- Gal
galactose
- GlcNAc
N-acetylglucosamine
- UDP
uridine 5-diphosphate
- UDP-Glc
uridine-5-diphosphoglucose
- UDP-Gal
uridine-5-diphosphogalactose
- PEP
phosphoenolpyruvate 相似文献
20.
Sialyl Lewis X ganglioside analogues containing 5-acetamido-3,5-dideoxy-l-arabino-2-heptulopyranosylonic acid (C7-Neu5Ac), 5-acetamido-3,5-dideoxy-d-galacto-2-octulopyranosylonic acid (C8-Neu5Ac), and 5-acetamido-3,5-dideoxy-l-glycero-d-galacto-1-2-nonulopyranosylonic acid (8-epi-Neu5Ac) in place ofN-acetylneuraminic acid (Neu5Ac) have been synthesized. Glycosylation of 2-(trimethylsilyl)ethyl 6-O-benzoyl--d-galactopyranoside with the phenyl or methyl 2-thioglycoside derivatives of the respective sialic acids, usingN-iodosuccinimide (NIS)-trifluoromethanesulfonic acid as a promoter in acetonitrile, gave the three required 2-(trimethylsilyl)ethyl (2S)-sialyl-(2 3)--galactopyranosides. These were converted viaO-benzoylation, selective transformation of the 2-(trimethylsilyl)ethyl group to acetyl, and introduction of the methylthio group with methylthiotrimethylsilane into the corresponding glycosyl donors. Glycosylation of 2-(trimethylsilyl)ethylO-(2,3,4-tri-O-benzyl--l-fucopyranosyl)-(1 3)-O-(2-acetamido-6-O-benzyl-2-deoxy--d-glucopyranosyl)-(1 3)-2,4,6-tri-O-benzyl--d-galactopyranoside with these donors in the presence of dimethyl(methylthio)sulfonium triflate (DMTST) afforded the expected -glycosides, which were converted into the corresponding -trichloroacetimidates, and these, on coupling with (2S, 3R, 4E)-2-azido-3-O-benzoyl-4-octadecene-1,3-diol, gave the required -glycosides. Finally, these were transformed via selective reduction of the azide group, condensation with octadecanoic acid,O-deacylation, and de-esterification into the target compounds in good yields. 相似文献