Non-reducing Trisaccharide Research Articles

Structural characterization of the O-polysaccharide of the lipopolysaccharide produced by Salmonella milwaukee O:43 (group U) which possesses human blood group B activity.

The O-polysaccharide of the lipopolysaccharide produced by Salmonella milwaukee O:43 (group U) was shown by composition analysis, methylation, periodate oxidation, and 1H and 13C nuclear magnetic resonance spectroscopic analytical methods to be a polymer of branched pentasaccharide repeating units having the structure: [formula: see text] The blood-group activity of the O-polysaccharide was established by its serological reactivity with a specific monoclonal antibody to human blood group B, using passive hemagglutination and ELISA assays, indicating the common antigenic epitope to be a nonreducing terminal trisaccharide unit composed of L-Fucp and D-Galp (1:2) residues.

Transglycosylation and transfer reaction activities of endo-α-N-acetyl-D-galactosaminidase from Diplococcus (Streptococcus) pneumoniae

Abstract Endo-alpha-N-acetyl-D-galactosaminidase from Diplococcus pneumoniae was shown to have transglycosylation and transfer reaction (reversed hydrolysis) activities. Treatment of asialoglycoproteins having Gal beta 1----3GalNAc alpha 1----Ser/Thr linkages with enzyme preparations containing glycerol resulted in formation of nonreducing trisaccharides. The structure of the main trisaccharide (approximately 80%) was deduced to be Gal beta 1----3GalNAc alpha 1----1(3)-glycerol by analysis of sugar composition and the results of exoglycosidase treatment and periodate oxidation. The ability of the endoglycosidase to catalyze transfer of Gal beta 1----3GalNAc to various acceptors was also demonstrated by incubation of the enzyme with the disaccharide and the test compound. The following were found to show acceptor activity: glycerol, Tris, p-nitrophenol, threonine, serine, D-glucose, D-galactose, D-fucose, and 6-O-methylgalactose. Transfer to the primary hydroxyl groups of glycerol and hexoses appears to be favored since the major glycerol product was 1(3)-substituted and transfer to D-fucose and 6-O-methyl-D-galactose was less than that to D-galactose. In order to avoid spurious results, it is necessary to carry out incubations with this enzyme in the absence of glycerol and other hydroxy compounds. The potential use of this endoglycosidase in the synthesis of glycosides is indicated by our studies.

Structure of the O-polysaccharide chain of lipopolysaccharide from Pseudomonas aeruginosa IID 1001 (ATCC 27577).

Structural studies were carried out on an acidic O-polysaccharide released by mild acid treatment from the lipopolysaccharide of Pseudomonas aeruginosa IID 1001 (ATCC 27577), which is serologically related to the serotypes Habs O3, Lanyi O1, and Wokatsch O13 in other serological classifications of Pseudomonas aeruginosa. The composition and data from structural analyses including H-NMR and C-NMR measurements, methylation, and Smith degradation showed that the structure of the IID 1001 O-polysaccharide was coincident with that of the Habs O3 and Lanyi O1 antigens (or Wokatsch O13). However, whereas solvolysis of the O-antigen of Habs O3 as well as that of Lanyi O1 by hydrogen fluoride has been reported to yield a reducing trisaccharide, GlcNAc(alpha 1----4)GalNAcA(alpha 1----3)Bac2NAc4Nacyl (acyl represents a 3-hydroxybutanoyl group), hydrogen fluoride hydrolysis of IID 1001 O-polysaccharide yielded a nonreducing trisaccharide with the reducing terminal bacillosamine residue linked at C-1 to the hydroxyl group of its N-acyl substituent, 3-hydroxybutanoic acid. These results, in combination with mass spectral data, led to the most likely structure for the tetrasaccharide repeating unit of the IID 1001 O-polysaccharide, (formula; see text) in which the location of N-acyl groups on bacillosamine residues differs from that in the O-antigens of Habs O3 and Lanyi O1 (or Wokatsch O13).

Separation and partial sequence analysis of blood group A-active oligosaccharides by affinity chromatography using monoclonal antibodies

An affinity column containing an anti-blood group A monoclonal antibody coupled to Sepharose beads specifically retards oligosaccharides with the nonreducing trisaccharide sequence GalNAcα1−3(Fucα1−2)Ga1β1-R. Three A-active oligosaccharides, A-tetra, A-penta, and A-hepta, elute as retarded peaks, well-separated from unbound sugars. A-hepta, which contains a difucosylated type 1 (Le b) core structure, elutes much later than A-tetra or A-penta and can be completely separated from the latter oligosaccharides by affinity chromatography. The order of elution of the oligosaccharides agrees with their previously determined specific molar activities as inhibitors of quantitative immune precipitation [H.-T. Chen, and E. A. Kabat, (1985) J. Biol. Chem. 260, 13208–13217]. Treatment of A-hepta with Charonia lampas α-galactosaminidase abolishes its binding by the anti-A affinity column and converts it to a Le b-active oligosaccharide (lacto- N-difucohexaose I) that is specifically retarded on a second affinity column containing an anti-Le b monoclonal antibody.

A new trisaccharide, 3G-.ALPHA.-D-glucosyl-sucrose, synthesized by the transglucosidase action of immobilized buckwheat .ALPHA.-glucosidase.

Enzymatic synthesis of α-glucosyl-sucrose through the transglucosidase action of immobilized buckwheat α-glucosidase was attempted in a reaction system containing maltose and sucrose. Three kinds of nonreducing trisaccharides were produced, and a large portion of the trisaccharide was 4G-α-d-glucosyl-sucrose (erlose). Another was a new trisaccharide, O-α-d-glucopyranosyl-(l → 3)-O-α-d-glucopyranosyl-(l → 2)-O-d-fructofuranoside (3G-α-d-glucosyl-sucrose; [α]d+120°, in water), and the other was 6G-α-d-glucosyl-sucrose (theanderose; [α]d+107°, in water). The new trisaccharide was named “esculose.”

A new trisaccharide, 6f-.ALPHA.-D-glucosyl-sucrose, synthesized by transglucosylation reaction of brewer's yeast .ALPHA.-glucosidase.

Enzymatic synthesis of oligosaccharide through the transglucosylation reaction of brewer’s yeast α-glucosidase was attempted in the reaction system containing only sucrose as substrate. As the transglucosylation product, a new nonreducing trisaccharide was chromato-graphically isolated in crystalline form (monohydrate). giving mp 139~142°C and + 102 (in water). The trisaccharide was confirmed to be 6F-α-d-glucosyl-sucrose, that is, O-α-d-glucopyranosyl-(1 → 6)-O-β-d-fructofuranosyl-(2 → 1)-O-α-d-glucopyranoside, of which undecaacetate gave mp 148°C and (in chloroform), and this sugar was named “isomelezitose.”

Lactulosucrose (4 F-β-galactosylsucrose), a new trisaccharide synthesized by cultures of Leuconostoc mesenteroides strain K (NRRL B-1299)

Leuconostoc mesenteroides strain K has been found to synthesize a new nonreducing trisaccharide of melezitose-planteose type, lactulosucrose. This trisaccharide has been isolated in crystalline form and shown to have the structure, O-β- d-galactopyranosyl-(1 → 4)- O-β- d-fructofuranosyl-(2 → 1)-α- d-glucopyranoside (4 F-β-galactosylsucrose). An experiment with C 14-sucrose established that lactulosucrose is formed by α-glucosyl transfer from sucrose to the anomeric carbon of lactulose (β-fructofuranose form). Discovery of the new saccharide followed an unsuccessful attempt to confirm a recent report that L. mesenteroides K (or B-1299) converts a mixture of sucrose and lactose to lactsucrose (4 G-β-galactosylsucrose). Synthesis of the latter saccharide was verified in the case of the NRRL B-512 strain of L. mesenteroides, and would be due to levansucrase-catalyzed β-fructosyl transfer from sucrose to lactose in this culture.

Trisaccharides of Lolium and Festuca.

IN the course of an investigation into the distribution of water-soluble carbohydrates in seeds of the Gramineae1, it was observed that two species of grass (Lolium perenne L. and Festuca pratensis Huds.) contained in their caryopses substantial amounts of a trisaccharide which was not detected in any of the thirty other wild grasses and cereals examined. This non-reducing trisaccharide (Fig. 1, X) had a specific rotation of + 90.5° (without recrystallization), yielded equal amounts of galactose, glucose and fructose on acid hydrolysis and was readily broken down to fructose and a reducing disaccharide in the presence of invertase or very dilute acid. This ease of hydrolysis suggests that the trisaccharide resembles raffinose in containing a terminal β-fructofuranoside residue. Iodine oxidation2 indicated that the galactose residue was at the non-reducing end of the disaccharide, and electrophoretic mobility of the disaccharide in borate was intermediate between the mobilities of laminari-biose and melibiose. Movement in borate is allied to the mode of linkage of sugar residues3, disaccharides containing 1,2 or 1,4 linkages being of much lower mobility than those containing 1,3 or 1,6 linkages. The disaccharide under consideration would therefore appear to contain either a 1,3 or a 1,6 linkage. Although the electrophoretic behaviour in borate does not distinguish between α and β linkages, the high positive rotation of the trisaccharide suggests that the galactose and glucose residues are α-linked, and, since 6-glucose-α-galactoside is melibiose, it seems probable that the newly isolated disaccharide is 3-glucose-α-galactoside. The trisaccharide from Lolium perenne and Festuca pratensis is therefore tentatively described as α-D-galactopyranosyl-3-α-D-glucopyranosy 1-2- β-D- fructofuranoside.

Structure of the carbohydrates

We investigated the acceptor specificity of a thermostable trehalose phosphorylase from Thermoanaerobacter brockii ATCC 35047 (TbTP) was examined using β-D-glucose-1-phosphate (β-G1P) as a glucosyl donor and oligosaccharides as the acceptor. Oligosaccharides with a reducing-end glucose residue as the C-6 substituent (e.g., isomaltose, gentiobiose, melibiose, isomaltotriose, and isopanose) were found to be successful acceptors. The transfer products of isomaltose, gentiobiose, and melibiose were isolated and characterized as 6-O-α-D-glucopyranosyl trehalose (α-GlcTre), 6-O-β-D-glucopyranosyl trehalose (β-GlcTre), and 6-O-α-D-galactopyranosyl trehalose (α-GalTre), respectively. To produce α-GalTre, a novel nonreducing trisaccharide, the reaction conditions of α-GalTre were examined using trehalose as a glucosyl donor. As a result, the yield of α-GalTre reached 40.5%.