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At least 19 recordsLinked to original sources
PubMed2001

Enzymic feruloylation of arabinoxylan-trisaccharide by feruloyl-CoA:arabinoxylan-trisaccharide O-hydroxycinnamoyl transferase from Oryza sativa.

Feruloyl-CoA:arabinoxylan-trisaccharide O-hydroxycinnamoyl transferase, which catalyzes the transfer of ferulic acid from Fer-CoA to arabinoxylan-trisaccharide in the formation of feruloyl arabinoxylan-trisaccharide (Fer-AXX), has been found in an ionically bound fraction and a cytosol fraction of suspension-cultured rice (Oriza sativa L. cv. Nipponbare) cells. Analysis of reaction products by high-performance liquid chromatography showed the formation of product A, which is one of the transfer products having the same retention time as authentic Fer-AXX. Product A was purified by reverse-phase chromatographies to characterize its structure. The isolated product A showed the same ultraviolet spectrum and molecular weight on fast atom bombardment mass spectrometric analysis as those of authentic Fer-AXX. Alakaline saponification of product A released ferulic acid and oligosaccharide. The released oligosaccharide consisted of arabinose and xylose in a molar ratio of 1:2. These results support the identity of product A as feruloylated arabinoxylan-trisaccharide and show the existence of a feruloyltransferase catalyzing the feruloylation of a hemicellulosic fragment.

Acylation↗
PubMed1994

Solution structure of a trisaccharide-antibody complex: comparison of NMR measurements with a crystal structure.

NMR and crystallography have been used to study antigen conformational changes that occur in a trisaccharide-Fab complex in solution and in the solid state. NOE buildup rates from transferred NOE experiments show that the antigenic determinant of a Salmonella lipopolysaccharide, represented by the trisaccharide methyl glycoside alpha-D-Galp(1-->2 [alpha-D-Abep(1-->3)]- alpha-D-Manp1-->OMe (1), undergoes a protein-induced conformational shift about the Gal-->Man glycosidic linkage when it is bound by a monoclonal antibody in aqueous solution. The same trisaccharide was crystallized with Fab, and a solved structure at 2.1-A resolution revealed that the conformation of the trisaccharide ligand was similar to that seen in a dodesaccharide-Fab complex [Cygler et al. (1991) Science 253, 442-445), where the Gal-Man linkage also experienced a similar conformational shift. Distance constraints derived from the TRNOE buildup curves are consistent with two bound trisaccharide conformations, one of which correlates with the ligand conformation of the crystalline Fab-trisaccharide complex. In this bound conformation, short interatomic distances between Abe O-2 and Gal O-2 permit an oligosaccharide intramolecular hydrogen bond. Despite its relatively low energy, a preponderance of this conformer could not be detected in aqueous or DMSO solutions of free trisaccharide by either 1H or 13C NMR experiments. In DMSO, a different intramolecular hydrogen bond between Abe O-2 and Man O-4 was observed due to a solvent-induced shift in the conformational equilibria (relative to aqueous solution). Molecular modeling of the trisaccharide in the binding site and as the free ligand suggested that the protein imposes an induced fit on the antigen, primarily resulting in a shift of the Gal-Man phi torsional angle. This reduces the interproton separation between Abe H-3 and Gal H-1 with a marked increase in the intensity of the previously weak NOEs between the protons of the noncovalently linked galactose and abequose residues. The impact of the conformational shift on gross trisaccharide topology is sufficiently small that binding modes inferred from functional group replacements are not impaired.

Antibodies, Monoclonal↗
PubMed2005

Oligosaccharide preferences of beta1,4-galactosyltransferase-I: crystal structures of Met340His mutant of human beta1,4-galactosyltransferase-I with a pentasaccharide and trisaccharides of the N-glycan moiety.

beta-1,4-Galactosyltransferase-I (beta4Gal-T1) transfers galactose from UDP-galactose to N-acetylglucosamine (GlcNAc) residues of the branched N-linked oligosaccharide chains of glycoproteins. In an N-linked biantennary oligosaccharide chain, one antenna is attached to the 3-hydroxyl-(1,3-arm), and the other to the 6-hydroxyl-(1,6-arm) group of mannose, which is beta-1,4-linked to an N-linked chitobiose, attached to the aspargine residue of a protein. For a better understanding of the branch specificity of beta4Gal-T1 towards the GlcNAc residues of N-glycans, we have carried out kinetic and crystallographic studies with the wild-type human beta4Gal-T1 (h-beta4Gal-T1) and the mutant Met340His-beta4Gal-T1 (h-M340H-beta4Gal-T1) in complex with a GlcNAc-containing pentasaccharide and several GlcNAc-containing trisaccharides present in N-glycans. The oligosaccharides used were: pentasaccharide GlcNAcbeta1,2-Manalpha1,6 (GlcNAcbeta1,2-Manalpha1,3)Man; the 1,6-arm trisaccharide, GlcNAcbeta1,2-Manalpha1,6-Manbeta-OR (1,2-1,6-arm); the 1,3-arm trisaccharides, GlcNAcbeta1,2-Manalpha1,3-Manbeta-OR (1,2-1,3-arm) and GlcNAcbeta1,4-Manalpha1,3-Manbeta-OR (1,4-1,3-arm); and the trisaccharide GlcNAcbeta1,4-GlcNAcbeta1,4-GlcNAc (chitotriose). With the wild-type h-beta4Gal-T1, the K(m) of 1,2-1,6-arm is approximately tenfold lower than for 1,2-1,3-arm and 1,4-1,3-arm, and 22-fold lower than for chitotriose. Crystal structures of h-M340H-beta4Gal-T1 in complex with the pentasaccharide and various trisaccharides at 1.9-2.0A resolution showed that beta4Gal-T1 is in a closed conformation with the oligosaccharide bound to the enzyme, and the 1,2-1,6-arm trisaccharide makes the maximum number of interactions with the enzyme, which is in concurrence with the lowest K(m) for the trisaccharide. Present studies suggest that beta4Gal-T1 interacts preferentially with the 1,2-1,6-arm trisaccharide rather than with the 1,2-1,3-arm or 1,4-1,3-arm of a bi- or tri-antennary oligosaccharide chain of N-glycan.

Carbohydrate Conformation↗
PubMed1993

[Development of hybridomas secreting monoclonal anti-idiotypic antibodies that bear the internal image of the terminal trisaccharide of PGL-I].

Two hybridomas designated as F7B7 and F7B9 secreting monoclonal anti-idiotypic antibodies against terminal trisaccharide of PGL-I were developed by fusion of SP2/0 cells and spleen cells of BALB/c mouse immunized with mouse monoclonal anti-trisaccharide of PGL-I (MAb1-E10F1). To characterize the F7B7, the following results were obtained. First, F7B7 reacted with MAb1-E10F1 specifically. Secondly, the cross ELISA neutralizing tests gave positive results. The binding of anti-trisaccharide positive serum with trisaccharide (contained in semi-synthetic antigen, NT-O-BSA) was inhibited F7B7 and the degree of inhibition showed dose-dependent manner. The binding of anti-trisaccharide positive serum with F7B7 was inhibited by NT-O-BSA and the degree of inhibition also showed dose-dependent manner. It was concluded that the hybridoma F7B7 is able to secrete monoclonal anti-idiotypic antibodies that bear the internal image of trisaccharide of PGL-I. The potentials and advantages of monoclonal anti-idiotypic antibody F7B7 as surrogate antigen in the serodiagnosis of leprosy have been discussed.

Animals↗
PubMed1996

Conformational analysis of blood group A trisaccharide in solution and in the binding site of Dolichos biflorus lectin using transient and transferred nuclear Overhauser enhancement (NOE) and rotating-frame NOE experiments.

The present study is concerned with the elucidation of the conformation of the blood group A trisaccharide (alpha-D-GalNAc(1-3)[alpha-L-Fuc(1-->2)] beta-D-Gal-O-R) in the combining site of Dolichos biflorus seed lectin by use of 400-MHz and 600-MHz NMR spectroscopy. D. biflorus lectin displays a unique specificity for GalNAc residues. It occurs in solution as a tetrameric assembly having a molecular mass of 110 kDa, with two carbohydrate-binding sites per molecule. First, NOE build-up curves were obtained for the free blood group A trisaccharide from one-dimensional transient NOE experiments. Simulated NOE build-up curves were constructed from an ensemble of low-energy conformers derived from previous investigations. The comparison of theoretical and experimental data indicates that an equilibrium between two families of low-energy conformers most likely reflects the solution behavior of the trisaccharide in solution. Two-dimensional transferred NOE and rotating-frame enhancements (ROE) were subsequently measured for the trisaccharide complexed with the D. biflorus seed lectin. In addition to the NOEs observed for the free trisaccharide, the transferred NOESY spectrum showed several new NOEs that were identified as spin diffusion using a rotating-frame NOESY (ROESY) experiment. Experimental interglycosidic transferred nuclear Overhauser effect (TRNOE) build-up curves were compared to theoretical curves calculated for both low-energy conformers located in the D. biflorus lectin-binding site. Calculations of theoretical TRNOE were performed using a combination of the full relaxation matrix and the protein-ligand exchange matrix. Comparison between experimental and simulated TRNOE volumes leads to the conclusion that one conformation of blood group A trisaccharide is selected upon binding by D. biflorus lectin.

ABO Blood-Group System↗
PubMed1992

Linkage position determination in a novel set of permethylated neutral trisaccharides by collisional-induced dissociation and tandem mass spectrometry.

A set of neutral permethylated trisaccharides identical in the non-reducing (A----B) disaccharide and linkage isomeric in the reducing terminal (B----C) disaccharide has been synthesized. Collision-activated tandem mass spectrometry was used for analysis of the B----C linkage position. The trisaccharides, gal(beta 1----4)glc(beta 1----X)glc, where X = 3, 4 and 6, were synthesized and examined using fast atom bombardment collision-induced dissociation tandem mass spectrometry. Results were rationalized using molecular modeling. We have previously reported results for determination of the A----B linkage position with isomeric sets of synthetic trisaccharides containing internal amino sugars. The neutral trisaccharides were synthesized to isolate electronic effects of the amino group. An approach of relating daughter ion to parent ion ratios and collision energy offset was used to generate slopes that predict linkage position in glc beta 1----X glc reducing end glycoside form of terminal trisaccharides.

Magnetic Resonance Spectroscopy↗
PubMed2005

An improvement in the bending ability of a hinged trisaccharide with the assistance of a sugar-sugar interaction.

Hinged di- and trisaccharides incorporating 2,4-diamino-beta-D-xylopyranoside as a hinge unit (Hin) were synthesized. Bridging of the diamino group of Hin by carbonylation or chelation to a metal ion results in a conformational change from (4)C1 to (1)C4, which in turn causes a bending of the oligosaccharides. In this study, the bending abilities of the hinged oligosaccharides were compared, in terms of the reactivities toward carbonylation and chelation. Di- or trisaccharides containing a 6-O-glycosylated mannopyranoside or galactopyranoside at their reducing ends had bending abilities similar to that of the Hin monosaccharide, probably because there were neither attractive nor repulsive interactions between the reducing and nonreducing ends. However, when Hin was attached at O2 of methyl mannopyranoside (Man alphaMe), the bending ability was dependent on the nonreducing sugar and the reaction conditions. Typically, a disaccharide--Hin beta(1,2)Man alphaMe--was difficult to bend under all the tested reaction conditions, and the bent population in the presence of Zn(II) was only 4%. On the other hand, a trisaccharide--Man alpha(1,3)Hin beta(1,2)Man alphaMe--was bent immediately after the addition of Zn(II) or Hg(II), and the bent population reached 75%, much larger than those of all the other hinged trisaccharides ever tested (<40%). This excellent bending ability suggests an attractive interaction between the reducing and nonreducing ends. The extended conformation was recovered by the addition of triethylenetetramine, a metal ion chelator. Reversible, quick, and efficient bending of the hinged trisaccharide was thus achieved.

Carbohydrate Conformation↗
PubMed1993

Monoclonal antibodies that recognize the trisaccharide epitope Gal alpha 1-3Gal beta 1-4GlcNAc present on Ehrlich tumor cell membrane glycoproteins.

Monoclonal antibodies were prepared against the trisaccharide Gal alpha 1-3Gal beta 1-4GlcNAc, a sequence which occurs on the surface of Ehrlich ascites tumor cells as well as in thyroglobulin, laminin and a variety of other proteins. This was accomplished by immunizing BALB/c mice with the fraction of Ehrlich cell membrane glycoproteins obtained by affinity chromatography on a Griffonia simplicifolia I (GS I) column which selectively binds alpha-D-galactosyl-terminated structures. Detection of Gal alpha 1-3Gal beta 1-4GlcNAc-specific antibodies was accomplished by employing glycoproteins containing the trisaccharide sequence; fusion with spleen cells from an immunized mouse was accomplished in the presence of polyethylene glycol (PEG1500). An enzyme-linked immunosorbent assay (ELISA) system was used to identify two clones (2.10G and 6.8E), which recognized the desired trisaccharide conjugate. These clones also recognized a thyroglobulin fraction isolated by GS I affinity chromatography and murine laminin, both of which possess the Gal alpha 1-3Gal beta 1-4GlcNAc sequence. Inhibition of antibody-trisaccharide reactivity, examined employing an ELISA assay, revealed that two trisaccharides, Gal alpha 1-3Gal beta 1-4GlcNAc/Glc, were the best inhibitory haptens; Gal beta 1-4GlcNAc (LacNAc), Gal alpha 1-3Gal and Gal beta 1-4Glc (lactose) were poor inhibitors. Indirect immunofluorescence staining of unfixed Ehrlich cells using the monoclonal antibody at 4 degrees C revealed fluorescence over the entire cell surface. Indirect immunogold labeling of semithin and ultrathin sections of aldehyde fixed and Lowicryl K4M-embedded Ehrlich cells resulted in specific labeling of the cell surface and internal structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Synthesis of O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1----3)-O-beta-D- galactopyranosyl-(1----4)-2-acetamido-2-deoxy-D-glucopyranose and two related trisaccharides containing the "lacto-N-biose II" unit.

Condensation of 3-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D- glucopyranosyl)-2,4,6-tri-O-acetyl-alpha-D-galactopyranosyl bromide (1) with benzyl 2-acetamido-3,6-di-O-benzyl-2-deoxy-alpha-D-glucopyranoside in boiling benzene and in the presence of mercuric cyanide afforded a trisaccharide that was O-deacetylated to give benzyl O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-(1----3)-O-beta-D- galactopyranosyl-(1----4)-3,6-di-O-benzyl-2-deoxy-alpha-D-glucopyranosid e, the benzyl groups of which were cleaved by catalytic hydrogenolysis to furnish the title trisaccharide. Alternatively, bromide 1 was allowed to react with 2-acetamido-1,6-anhydro-3-O-benzyl-2-deoxy-beta-D-glucopyranose in 1:1 benzene-nitromethane in the presence of mercuric cyanide, followed by O-deacetylation, column chromatography, and reacetylation to give O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D- glucopyranosyl)-(1----3)-O-(2,4,6-tri-O-acetyl-beta-D-galactopyranosyl)- (1----4)-2-acetamido-1,6-anhydro-3-O-benzyl-2-deoxy-beta-D-glucopyranose . Acetolysis, followed by catalytic hydrogenation and subsequent O-deacetylation furnished the title trisaccharide. A similar condensation of bromide 1 with 4-nitrophenyl 2-acetamido-2-deoxy-4,6-O-(4-methoxybenzylidene)-beta-D-glucopyran oside and 2-nitrophenyl 2-acetamido-2-deoxy-4,6-O-(4-methoxybenzylidene)-alpha-D-galactopy ranoside produced two trisaccharide derivatives, the acetal groups of which were cleaved in hot, 60% aqueous acetic acid, and the resulting diol intermediates O-deacetylated to furnish the desired trisaccharides 4-nitrophenyl O-(2-acetamido-2-deoxy-beta-D- glucopyranosyl)-(1----3)-O-beta-D-galactopyranosyl)-(1----3)-2-acetamido -2- deoxy-beta-D-glucopyranoside and 2-nitrophenyl O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl-(1----3)-O-beta- D-galactopyranosyl-(1----3)-2-acetamido-2-deoxy-beta-D-galactopyranoside .

Carbohydrate Conformation↗
PubMed2006

Targeted gene delivery with trisaccharide-substituted chitosan oligomers in vitro and after lung administration in vivo.

The aim of this study was to improve the gene delivery efficacy of chitosan oligomer polyplexes by introducing a trisaccharide branch that targets cell-surface lectins. For this purpose, chitosan oligomers were substituted by a trisaccharide with the N-acetylglucosamine residue at the free end, and the ability of the trisaccharide-substituted chitosan oligomers (TCO) polyplexes to transfect various cell lines in vitro and lung tissue after in vivo administration to mice was investigated. Live-cell confocal microscopy showed improved cellular uptake in HEK 293 cells (11-fold, p<0.001) for the TCO polyplexes compared with the linear chitosan oligomers. Colloidal stability was also enhanced with the substituted form, which suggests that the trisaccharide branch stabilised the polyplexes by means of a steric stabilisation mechanism. Interestingly, gene expression levels in the human liver hepatocyte (HepG2) cells were 10-fold higher with the TCO polyplexes than those mediated by polyethyleneimine. A similar improvement was obtained in a human bronchial epithelial cell line (16HBE14o-). Transfection with the TCO was significantly inhibited (by 30-80%), for all the cell lines tested, in the presence of the free trisaccharide branch, confirming lectin-mediated uptake. Finally, in vivo studies showed that, 24 h after lung administration to mice, luciferase gene expression was 4-fold higher with the TCO than with the corresponding linear chitosan oligomers.

Animals↗
PubMed2002

Characterization of the recognition of blood group B trisaccharide derivatives by the lectin from Marasmius oreades using frontal affinity chromatography-mass spectrometry.

A novel lectin from the mushroom Marasmius oreades (MOA) has been shown to bind to human blood group B oligosaccharides [1]. In the present work we examine the binding of a series of analogues of the blood group B-trisaccharide, alphaGal(1-3)[alphaFuc(1-2)]betaGal-OR (1, R = (CH2)8COOMe). MOA was biotinylated and immobilized on a micro column (9.8 microL) for evaluation by Frontal Affinity Chromatography-Mass Spectrometry (FAC-MS) [2]. The trisaccharide 1 was found to be the epitope needed for maximum recognition (Kd = 3.6 microM). A series of synthetic deoxygenated and O-methylated analogues of the B-trisaccharide (R = OMe) were then screened against the lectin, and the key structural elements for binding were determined. OH-4 of the beta-Gal residue and OH-2 of the alpha-Gal residue were found to be critical for recognition. The FAC-MS technique also proved powerful in evaluating mixtures of compounds. Since the solution NMR structure and crystal structure of the B-trisaccharide are known [3], we propose the specific surface of the trisaccharide that is recognized by the lectin.

ABO Blood-Group System↗
PubMed1987

Chemical synthesis of the trisaccharide unit of the species-specific phenolic glycolipid from Mycobacterium leprae.

O-(3,6-Di-O-methyl-beta-D-glucopyranosyl)-(1----4)-O-(2,3-di-O-methyl- alpha-L-rhamnopyranosyl)-(1----2)-3-O-methyl-L-rhamnopyranose, the haptenic trisaccharide of the Mycobacterium leprae-specific phenolic glycolipid I (PGL-I) antigen, and related trisaccharides, were synthesized by allylation of O-2 of benzyl 4-O-benzyl-alpha-L-rhamnopyranoside using phase-transfer catalysis, methylation of the product, deallylation, and coupling to O-(2,4-di-O-acetyl-3,6-di-O-methyl-beta-D-glucopyranosyl)-(1----4)-2,3- di-O-methyl-L-rhamnopyranosyl bromide or related disaccharides. Anomeric mixtures of the trisaccharide derivatives were separated by preparative t.l.c., deacetylated, and hydrogenolyzed, to give the pure trisaccharides. It had already been demonstrated that only those trisaccharides containing an intact, terminal 3,6-di-O-methyl-beta-D-glucopyranosyl unit are effective in inhibiting the specific binding between PGL-I and anti-PGL-I immunoglobulin M antibodies in human lepromatous leprosy sera.

Antigen-Antibody Complex↗
PubMed1998

Synthesis of Tethered Trisaccharides To Probe Inter-Saccharide Flexibility in Carbohydrate-Protein Interactions.

Two crystal structures of the trisaccharide epitope alpha-D-Galp(1-->2)[alpha-D-Abep(1-->3)alpha-D-Manp1-->OCH(3) 1 bound to antibody Fab and single-chain Fv fragments have been used to guide the design of an intramolecular tether that constrains the trisaccharide ligand to conformations close to those of the bound state. Preorganization of the ligand should overcome entropic penalties to binding and provide enhanced affinity. Three tethers [O(CH(2))(n)()O, n = 6, 7, and 8] were used to anchor the solvent-exposed C6 atoms of the mannose and galactose residues. Two synthetic schemes were employed. The first utilized an ethyl 1-thiogalactoside bearing the tether protected as a trityl ether 6-8. Glycosylation of the mannopyranoside 5 that contains a methanesulfonate at C6 by any of the donors 6-8 afforded disaccharides 37-39. Subsequent removal of the trityl group allowed an alkoxide to be generated on the omega-hydroxyl group of the tether for displacement of the sulfonate in 38% yield. By comparison when a omega-methanesulfonyloxy tether was incorporated as the mannoside 9, the disaccharide product 52 after reaction with the galactosyl donor 10 was converted to the macrocycle 4 in 61% yield. The 3,6-dideoxy-xylo-hexopyranose residue was introduced by thioglycoside chemistry to yield the protected, tethered trisaccharide target molecules 49-51, which were deprotected in a single hydrogenation step. Solid-phase enzyme immunoassays showed the tethered trisaccharides 2-4 to be weaker inhibitors than trisaccharide 1 (+0.2-0.3 kcal mol(-)(1)). This suggests that preorganization of oligosaccharides provides negligible gains in binding affinity.

Journal Article↗
PubMed2003

Application of NMR, molecular simulation, and hydrodynamics to conformational analysis of trisaccharides.

The preferred conformations and conformational flexibilities of the trisaccharides alpha-D-Glcp-(1-->2)-beta-D-Glcp-(1-->3)-alpha-D-Glcp-OMe (I) and alpha-D-Glcp-(1-->3)[beta-D-Glcp-(1-->4)]-alpha-D-Glcp-OMe (II) in aqueous solution were determined using nuclear magnetic resonance (NMR) spectroscopy, molecular dynamics (MD) and Langevin dynamics (LD) simulations, and hydrodynamics calculations. Both trisaccharides have a vicinal substitution pattern in which long range (nonsequential) interactions may play an important role. LD simulation at 600 K indicated that the all-syn conformation predominated, though other conformations were apparent. NOE data and MD and LD simulations at 298 K all indicated that trisaccharide I is a single all-syn conformer in solution. Given that previous studies showed evidence of anti-conformers in beta-D-Glcp-(1-->2)-beta-D-Glcp-(1-->3)-alpha-D-Glcp-OMe, this result provides an example of how changing the anomeric configuration of one residue from beta to alpha can make an oligosaccharide more rigid. Discrepancies in inter-ring distances obtained by experiment and by simulation of the all-syn conformer suggest the presence of an anti-psi conformation at the beta-(1-->4)-linkage for II. A combined analysis of measured and calculated translational diffusion constants and (13)C T(1) relaxation times yield order parameters of 0.9 for each trisaccharide. This implies that any interconversion among conformations is significantly slower than tumbling. Anisotropies of approximately 1.6 and 1.3 calculated for I and II, respectively, are consistent with the observed relatively flat T(1) profiles because the tumbling is not in the motional narrowing regime.

Carbohydrate Conformation↗
PubMed2003

Synthesis of a core trisaccharide as a versatile building block for N-glycans and glycoconjugates.

N-linked oligosaccharides from glycoproteins (N-glycans) can be conveniently assembled with a building block approach. A protected form of the core trisaccharide (beta-mannosyl chitobiose) was identified as a key building block. The chitobiose part of the core trisaccharide was built from a glycosyl fluoride, which served as a precursor for the reducing GlcNAc azide and the inner GlcNAc moiety. Beta-mannosylation was accomplished at the trisaccharide stage by intramolecular inversion of a beta-glucosyl chitobiose. The benzylidene protection of the beta-mannoside and the azido group at the reducing end of the core trisaccharide allow modular synthesis of N-glycans and their glycoconjugates.

Carbohydrate Conformation↗
PubMed1988

Synthesis and immunoreactivity of neoglycoproteins containing the trisaccharide unit of phenolic glycolipid I of Mycobacterium leprae.

The trisaccharide segment, O-(3,6-di-O-methyl-beta-D-glucopyranosyl)-(1----4)-O-(2,3-di-O-methyl- alpha-L-rhamnopyranosyl)-(1----2)-3-O-methyl-L-rhamnopyranose, of the Mycobacterium leprae-specific phenolic glycolipid I has been synthesized as its 8-(methoxycarbonyl)octyl glycoside and coupled to a carrier protein, to produce a leprosy-specific neoglycoprotein, the so-called natural trisaccharide-octyl-bovine serum albumin (NT-O-BSA). Special features of the synthetic strategy were the use of silver trifluoromethanesulfonate (triflate) to promote glycosylation, resulting in the rhamnobiose in high yield and absolute stereospecificity. The terminal 3,6-di-O-methyl-D-glucopyranosyl group was introduced after O-deallylation of the rhamnobiose. Removal of protecting groups yielded the trisaccharide hapten suitable for coupling to carrier protein. Poly(acrylamide)-gel electrophoresis of the neoglycoprotein demonstrated its purity, and subsequent immunoblotting with a monoclonal antibody directed to the terminal 3,6-di-O-methyl-beta-D-glucopyranosyl epitope of the native glycolipid demonstrated its antigenicity. Comparative serological testing in enzyme-linked immunosorbent assays of NT-O-BSA, the corresponding disaccharide-containing products, and another trisaccharide-containing neoglycoprotein, O-(3,6-di-O-methyl-beta-D-glucopyranosyl)-(1----4)-O-(2,3-di-O- methyl-alpha-L-rhamnopyranosyl)-(1----2)-(3-O-methyl-alpha-L-rhamnopy ran osyl)- (1----4')-oxy-(3-phenylpropanoyl)-BSA (NT-P-BSA) [Fujiwara et al., Agric. Biol. Chem., 51 (1987) 2539-2547] against sera from leprosy patients and control populations showed concordance; the presence of the innermost sugar did not contribute significantly to sensitivity or specificity. The di- and tri-saccharide-containing neoantigens, on account of ready availability and solubility, provide greater flexibility than the native glycolipid for the serodiagnosis of leprosy.

Antigens, Bacterial↗
PubMed1995

Simulation of the 13C nuclear magnetic resonance spectra of trisaccharides using multiple linear regression analysis and neural networks.

Predictive models are developed for the 13C NMR chemical shifts of the carbon atoms comprising the central rings of 46 trisaccharide compounds. Thirty-nine trisaccharides are used as a training set for development of models using regression analysis and computational neural networks, and seven compounds are used as an external prediction set. The descriptors used in the models are developed directly from the molecular structures of the trisaccharides. Three different methods of descriptor selection are compared. The dependence of the models on the geometries of the trisaccharides is explored. The models developed with geometric descriptors are better than those developed without geometric descriptors, although the latter models are still of a comparable quality. Overall, the best model found is a neural network based on descriptors selected by multiple linear regression.

Algorithms↗
PubMed2004

Synthesis of an alpha-linked dimer of the trisaccharide repeating unit of the exopolysaccharide produced by Pediococcus damnosus 2.6.

A hexasaccharide, beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->2)]-alpha-D-Glcp-(1-->3)-beta-D-Glcp-(1-->3)-[beta-D-Glcp-(1-->2)]-D-Glcp, the alpha-linked dimer of the trisaccharide repeating unit of the exopolysaccharide produced by Pediococcus damnosus 2.6, was synthesized as its methyl glycoside. Condensation of fully benzoylated alpha-D-glucopyranosyl trichloroacetimidate (1) with isopropyl 4,6-O-benzylidene-1-thio-beta-D-glucopyranoside (2) selectively furnished (1-->3)-linked disaccharide 3, and subsequent 2-O-acetylation, desulfation, and trichloroacetimidate formation afforded the disaccharide donor 6. Meanwhile, selective 3-O-coupling of methyl 4,6-O-benzylidene-alpha-d-glucopyranoside (8) with 3-O-allyl-2,4,6-tri-O-benzoyl-alpha-D-glucopyranosyl trichloroacetimidate (7), followed by coupling with 1 gave the trisaccharide 10. Removal of the benzylidene group of 10, benzoylation, and deallylation produced the trisaccharide acceptor 12. Condensation of 12 with 6 yielded a pentasaccharide mixture 13 with beta and alpha isomers in a ratio of 2:1. Removal of the benzylidene group of 13, followed by benzoylation gave the pentasaccharide mixture 14. Selective 2'''-deacetylation of the isolated beta-linked 14beta with MeCOCl/MeOH/CH2Cl2 did not give the expected pentasaccharide acceptor, and serious decomposition occurred, indicating a large steric hindrance at C-2'''. Alternatively, 2,3-di-O-glycosylation of allyl 4,6-O-benzylidene-beta-D-glucopyranoside (21) with 1 gave 22, then deallylation and trichloroacetimidate formation afforded the trisaccharide donor 24. Condensation of 12 with 24 furnished only the alpha-linked hexasaccharide 25, and its deprotection gave the free hexaoside 27.

Carbohydrate Conformation↗