- Synthesis and X-ray crystallographic investigation of N-(β-d-glycosyl) butanamides derived from GlcNAc and chitobiose as analogs of the conserved chitobiosylasparagine linkage of N-glycoproteins
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The linkage region, GlcNAcβAsn, is conserved in all eukaryotic N-glycoproteins. As a logical extension of a research endeavor aimed at understanding the structural significance of GlcNAc and Asn as the linkage region constituents, the newer analogs GlcNAcβNHBu and (GlcNAcβ(1-4) GlcNAc)alkanamides have been synthesized to assess the influence of aglycon as well as additional GlcNAc on the linkage region. X-ray crystallographic analysis of the GlcNAcβNHBu and (GlcNAcβ(1-4)GlcNAc)βNHBu is described. Comparative analysis of these structures with those of reported models and analogs shows that the deviation in N-glycosidic torsion, φN among the GlcNAc alkanamides is negligible (2) whereas (GlcNAcβ(1-4) GlcNAc)βNHBu deviates by ~15 as compared to GlcNAcβNHBu. Under the influence of the molecular packing, the conformation around the C1′-C2′ bond deviates from anti to gauche in (GlcNAcβ(1-4) GlcNAcβNHBu. Interestingly, C2-acetamido group in (GlcNAcβ(1-4)GlcNAc) NHBu orients differently as compared to GlcNAc alkanamides and this orientation was found to be almost similar to β-N,N′-diacetylchitobiose trihydrate. The bifurcated anti-parallel pattern involving N-HO and C-HO hydrogen bonds, a hallmark feature of the N-glycoprotein models, GlcNAcβNHAc and GlcNAcβAsn, is absent in both the title alkanamides. This is the first report on the crystal structure analysis of chitobiosyl alkanamide as analog of the N-glycoprotein linkage region, (GlcNAcβ(1-4) GlcNAc)βAsn.
- Mathiselvam, Manoharan,Srivastava, Amrita,Varghese, Babu,Pérez, Serge,Loganathan, Duraikkannu
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- Acetylated chitosan oligosaccharides act as antagonists against glutamate-induced PC12 cell death via Bcl-2/Bax signal pathway
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Chitosan oligosaccharides (COSs), depolymerized products of chitosan composed of β-(1→4) D-glucosamine units, have broad range of biological activities such as antitumour, antifungal, and antioxidant activities. In this study, peracetylated chitosan oligosaccharides (PACOs) and N-acetylated chitosan oligosaccharides (NACOs) were prepared from the COSs by chemcal modification. The structures of these monomers were identified using NMR and ESI-MS spectra. Their antagonist effects against glutamate-induced PC12 cell death were investigated. The results showed that pretreatment of PC12 cells with the PACOs markedly inhibited glutamate-induced cell death in a concentration-dependent manner. The PACOs were better glutamate antagonists compared to the COSs and the NACOs, suggesting the peracetylation is essential for the neuroprotective effects of chitosan oligosaccharides. In addition, the PACOs pretreatment significantly reduced lactate dehydrogenase release and reactive oxygen species production. It also attenuated the loss of mitochondrial membrane potential. Further studies indicated that the PACOs inhibited glutamate-induced cell death by preventing apoptosis through depressing the elevation of Bax/Bcl-2 ratio and caspase-3 activation. These results suggest that PACOs might be promising antagonists against glutamate-induced neural cell death.
- Hao, Cui,Gao, Lixia,Zhang, Yiran,Wang, Wei,Yu, Guangli,Guan, Huashi,Zhang, Lijuan,Li, Chunxia
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p. 1267 - 1289
(2015/04/14)
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- Improving glycopeptide synthesis: A convenient protocol for the preparation of β-glycosylamines and the synthesis of glycopeptides
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(Chemical Equation Presented) Herein we apply a recently introduced protocol using ammonium carbamate in methanol to the amination of crude chitobiose leading to 1,β-aminochitobiose. This simple, one-step procedure allows a facile preparation of unstable glycosylamines in contrast to the commonly implemented ammonium bicarbonate based amination of water-soluble carbohydrates. The new amination protocol leads to an improved synthesis of the key chitobiosyl-asparagine building block for the SPPS of glycopeptides. The utility of the method is demonstrated with the synthesis of a 39-amino acid glycoprotein.
- Hackenberger, Christian P. R.,O'Reilly, Mary K.,Imperiali, Barbara
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p. 3574 - 3578
(2007/10/03)
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- Probing the mechanism of a fungal glycosyltransferase essential for cell wall biosynthesis. UDP-Chitobiose is not a substrate for chitin synthase
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Chitin synthase is responsible for the biosynthesis of chitin, an essential component of the fungal cell wall. There is a long-standing question as to whether "processive" transferases such as chitin synthase operate in the same manner as non-processive transferases. The question arises from analysis of the polysaccharide structure - in chitin, for instance, each sugar residue is rotated ca. 180 deg relative to the preceding sugar in the chain. This requires that the enzyme account for the alternating "up/down" configuration during biosynthesis. An enzyme with a single active site, analogous to the non-processive transferases - would have to accommodate a distorted glycosidic linkage at every other synthetic step. An alternative proposal is that the enzyme might assemble the disaccharide donor, addressing the "up/down" conformational problem prior to polymer synthesis. We present compelling evidence that this latter hypothesis is incorrect.
- Chang, Robert,Yeager, Adam R.,Finney, Nathaniel S.
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- Chemo-enzymatic Synthesis of a Lipid-linked Core Trisaccharide of N-Linked Glycoproteins
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Two novel analogues of the mannosyl transferase acceptor substrate (GlcNAc)2-Pi-Pi-dolichyl 2 have been prepared in which dolichyl is replaced by phytanyl, 4, and lauryl, 5; both compounds 4 and 5 were synthesized using readily available chitin as the disaccharide precursor.Compound 4 acts as an efficient acceptor substrate for mannosyl transferase from porcine liver and saccharomyces cerevisiae.
- Flitsch, Sabine L.,Pinches, Heather L.,Taylor, James P.,Turner, Nicholas J.
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p. 2087 - 2094
(2007/10/02)
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- Synthesis of a Novel Acceptor Substrate for a Mannosyl Transferase
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Two novel analogues of the mannosyl transferase acceptor substrate (GlcNAc)2-pyrophosphate-dolichyl 2 have been prepared in which dolichyl is replaced by phytanyl 4 and lauryl 5; both 4 and 5 were synthesized using readily available chitin as the disaccharide precursor.
- Flitsch, Sabine L.,Taylor, James P.,Turner, Nicholas J.
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p. 380 - 382
(2007/10/02)
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- Synthesis of Partial Structures of N-Glycopeptides Representing the Linkage Regions of the Transmembrane Neuraminidase of an Influenca Virus and of Factor B of the Human Complement System
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The synthesis of the protected N-chitobiosylasparaganine peptides 12a and 14, which correspond to the linkage regions of factor B of the human complement system and of the transmembrane neuraminidase of an influenza virus, was accomplished by applying the protecting group combination Boc group/allyl ester together with acetyl groups in the carbohydrate part.From 12a the allyl ester and the acetates were simultaneously removed using ammonia in methanol.The Boc group was cleaved without side reactions using anhydrous trifluoroacetic acid.Attempts to remove the Boc-protecting group from the glycopeptides already deblocked in the carbohydrate part resulted predominantly in the cleavage of the O-glycosidic bond.
- Kunz, Horst,Waldmann, Herbert,Maerz, Joachim
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- Synthesis of trisaccharide determinants of enterobacterial common antigens
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In the presence of silver silicate, the reaction of 3,4,6-tri-O-acetyl-2-azido-2-deoxy-alpha-D-mannopyranosyl bromide with 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-beta-D-glucopyranose gave beta-glycosidically linked 1,6-anhydro-2-azido-3-O-benzyl-2-deoxy-4-O-(3,4,6- tri-O-acetyl-2-azido-2-deoxy-beta-D-mannopyranosyl)-beta-D- glucopyranose. After deacetylation and catalytic oxidation with oxygen and platinum, 1,6-anhydro-2-azido-4-O-[(2-azido-2-deoxy-beta-D-mannopyranosyl) uronic acid]-3-O-benzyl-2-deoxy-beta-D-glucopyranose was obtained. A series of intermediate steps led to the glycosyl donor 6-O-acetyl-2-azido-3-O-benzyl-4-O-[benzyl (3,4-di-O-acetyl-2-azido-2-deoxy-beta-D-mannopyranosyl)uronate]-2-deoxy- alpha, beta-glucopyranosyl chloride which was coupled with 8-methoxycarbonyloctyl 4-azido-2-O-benzyl-4,6-dideoxy-alpha-D-galactopyranoside to give 8-methoxycarbonyloctyl O-[benzyl (3,4-di-O-acetyl-2-azido-2-deoxy-beta-D-mannopyranosyl)uronate]- (1----4)-O-(6-O-acetyl-2-azido-3-O-benzyl-2-deoxy-alpha-D-glucopyranosyl )-(1----3)-4-azido-2-O-benzyl-4,6-dideoxy-alpha-D-galactopyranoside. Deblocking gave the spacer-linked repeating unit of the enterobacterial common antigen, beta-D-ManpNAcA-(1----4)-alpha-D-GlcpNAc-(1----3)-alpha- D-Fucp4NACO(CH2)8CO2CH3.
- Paulsen,Lorentzen
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- Synthesis of 4-Methylcoumarin-7-yloxy Tetra-N-acetyl-β-chitotetraoside, a Novel Synthetic Substrate for the Fluorometric Assay of Lysozyme
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4-Methylcoumarin-7-yloxy tetra-N-acetyl-β-chitotetraoside (10) was synthesized from chitin in pure form by a novel procedure.After acetolysis of chitin, chitotetraose tetradecaacetate (4) was isolated by Sephadex LH-20 column chromatography.Compound 4 was chlorinated with dry hydrogen chloride to produce tridecaacetyl chitotetraosyl chloride (6). 7-Hydroxy-4-methylcoumarin sodium salt was condensed with 6 under Koenigs-Knorr reaction conditions, and the final product (10) was obtained by de-O-acetylation of condensation product (8) with sodium methoxide. 4-Methylcoumarin-7-yloxy tri-N-acetyl-β-chitotrioside (9) was also synthesized through chitotriose undecaacetate (3) isolated together with 4 in the same chromatography.Compound 10 was used in a fluorometric assay of lysozyme in comparison with 9.The high sensitivity of fluorometric determination of 7-hydroxy-4-methylcoumarin (12) made it possible to determine lysozyme concentration in the microgram range by using this substrate (10).Unlike the assay using Micrococcus lysodeikticus cell powder, lysozyme assay with this synthetic substrate (10) could be performed directly in biological materials.Keywords - synthesis; lysozyme; enzyme assay; fluorometry; substrate; Sephadex LH-20; 7-hydroxy-4-methylcoumarin; 4-methylcoumarin-7-yloxy tetra-N-acetyl-β-chitotetraoside; 4-methylcoumarin-7-yloxy tri-N-acetyl-β-chitotrioside
- Inaba, Toyoaki,Ohgushi, Tadayasu,Iga, Yoshiro,Hasegawa, Eiichi
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p. 1597 - 1603
(2007/10/02)
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