A teichoic acid of Nocardioides albus VKM Ac-805T cell walls

A teichoic acid of Nocardioides albus VKM Ac-805T cell walls, a typical species of the genus Nocardioides, contains a poly(glycosylglycerol phosphate). The repeating unit of the polymer has the structure: [figure]. These units are in phosphodiester linkage at C-3 of glycerol and C-3 of beta-D-galactopyranose. beta-D-Galactopyranosyl residues are substituted at C-4 by beta-D-glucopyranose carrying a 4,6-pyruvate ketal group in S-configuration. The presence of pyruvic acid in the majority of repeating units increases the anionic properties of the polymer in comparison with most other common teichoic acids. This is the first report of the occurrence of a beta-D-galactofuranosyl residue in teichoic acids; it probably acts as a terminator of an extending chain of the polymer. The ratio of beta-D-galactopyranosyl to beta-D-galactofuranosyl units is 7:1. The polymer structure was determined by NMR spectroscopy. This type of teichoic acid structure has not been reported previously.     

An acetylated galactoglucomannan from Picea abies L. Karst

A water-soluble galactoglucomannan composed of D-galactose, D-glucose, and D-mannose in 1:3:17 mole proportion has been isolated from the secondary cell walls of Picea abies L. Karst. About 33% of the polysaccharide units were substituted by acetyl groups. Structural studies of the polymer indicated a beta-(1-->4)-linked glucomannopyranosyl backbone with a low content of branch points at O-6 of mannosyl and glucosyl residues. A preference for mannosyl groups indicates the presence of a single D-galactosyl unit side-chain. About half of the mannose residues were O-acetylated at C-2 and C-3 in 1.7:1 mole proportion.     

Purification and mode of action of two different arabinoxylan arabinofuranohydrolases from Bifidobacterium adolescentis DSM 20083

Two novel arabinofuranohydrolases (AXH-d3 and AXH-m23) were purified from Bifidobacterium adolescentis DSM 20083. Both enzymes were induced upon growth of Bi. adolescentis on xylose and arabinoxylan-derived oligosaccharides. They were only active with arabinoxylans and therefore denoted as arabinoxylan arabinofuranohydrolases. Their optimal activity was at pH 6 and 30–40 °C. They were very specific in their mode of action and were clearly different from AXH-m from Aspergillus awamori. AXH-m23 released only arabinosyl groups, which were linked to the C-2 or C-3 position of singly substituted xylose residues in arabinoxylan oligomers. AXH-d3 hydrolysed C-3-linked arabinofuranosyl residues of doubly substituted xylopyranosyl residues of arabinoxylans and arab- inoxylan-derived oligosaccharides. No activity was observed with C-2-linked arabinofuranosyl residues of these doubly substituted xylopyranosyl residues, or against C-2- and C-3-linked arabinofuranosyl residues of singly substituted xylopyranosyl residues. The combination of AXH-d3 and AXH-m showed low debranching activity with highly substituted glucurono-arabinoxylans. However, arabinoxylan from wheat flour was debranched almost completely. Received: 12 January 1999 / Accepted: 17 January 1999     

Characterization of structural component of cell walls of alkalophilic strain of Bacillus sp. C-125. Preparation of poly(gamma-L-glutamate) from cell wall component.

The cell wall of an alkalophilic strain of Bacillus sp. C-125 is composed of A1 gamma-peptidoglycan, a teichuronic acid and an unknown acidic polymer composed of glutamic acid and glucuronic acid, of which the molar ratio is approx. 4-5:1. Poly(gamma-L-glutamate) was prepared from the acidic polymer by removal of almost all of the glucuronic residues with trifluoromethanesulphonic acid treatment and purified chromatographically. The Mr of the polyglutamate preparation was estimated to be 14,000 by gel chromatography, or 43,000 on the basis of the content of N-terminal acid residues. The acidic polymer found in the cell wall of the organism was concluded to be a polyglutamate substituted with (oligo)glucuronic acid residues or a complex composed of two kinds of polymers (polyglutamate and polyglucuronate).     

Structural analysis of arabinoxylans isolated from native and malted finger millet (Eleusine coracana, ragi)

Structural elucidation of purified arabinoxylans isolated from finger millet and its malt by methylation, GLC-MS, periodate oxidation, Smith degradation, NMR, IR, optical rotation, and oligosaccharide analysis indicated that the backbone was a 1,4-beta-D-xylan, with the majority of the residues substituted at C-3. The major oligosaccharide generated by endo xylanase treatment was homogeneous with a molecular weight of 1865 Da corresponding to 14 pentose residues as determined by MALDI-TOF-MS and gel filtration on Biogel P-2. The structural analysis of this oligosaccharide showed that it contained 8 xylose and 6 arabinose residues, substituted at C-3 (monosubstituted) and at both C-2 and C-3 (disubstituted).     

Studies of polysaccharide fractions from the lipopolysaccharide of Pseudomonas aeruginosa N.C.T.C. 1999.

Two polymeric water-soluble fractions were isolated by gel filtration after mild acid hydrolysis of the lipopolysaccharide from Pseudomonas aeruginosa N.C.T.C. 1999. The fraction of higher molecular weight retained the O-antigenic specificity of the lipopolysaccharide and may be 'side-chain' material. This fraction was rich in N (about 10%) and gave several basic amino compounds on acid hydrolysis; fucosamine (at least 2.8% w/w) was the only specifc component identified. The fraction of lower molecular weight was a phosphorylated polysaccharide apparently corresponding to 'core' material. The major components of this fraction and their approximate molar proportions were: glucose (3-4); rhamnose (1); heptose (2); 3-deoxy-2-octulonic acid (1); galactosamine (1); alanine (1-1.5); phosphorus (6-7). In the intact lipopolysaccharide this fraction was probably linked to lipid A via a second residue of 3-deoxy-2-octulonic acid, and probably also contained additional phosphate residues and ethanolamine. The residues of 3-deoxy-2-octulonic acid were apparently substituted in the C-4 or C-5 position, and the phosphorylated heptose residues in the C-3 position. The rhamnose was mainly 2-substituted, though a little 3-substitution was detected. The glucose residues were either unsubstituted or 6-substituted. Four neutral oligosaccharides were produced by partial acid hydrolysis and were characterized by chemical, enzymic, chromatographic and mass-spectrometric methods of analysis. The structures assigned were: Glcpalpha1-6Glc; Glcpbeta1-2Rha; Rhapalpha1-6Glc; Glcpbeta1-2Rhapalpha1-6Glc. The galactosamine was substituted in the C-3 or C-4 position, the attachment of alanine was indicated, and evidence that the amino sugar linked the glucose-rhamnose region to the 'inner core' was obtained.     

The ribitol-phosphate-containing lipopolysaccharide from Proteus mirabilis, strain D52. Investigations on the structure of O-specific chains.

A soluble hydrophilic lipopolysaccharide, termed lipopolysaccharide II, isolated from Proteus mirabilis, strain D52 contained N-acetylglucosamine, glucose, galactose, ribitol phosphate and ethanolamine phosphate as constituents of the O-specific polysaccharide. Periodate oxidation studies were carried out on the polymer before and after dephosphorylation with hydrofluoric acid and on oligosaccharides derived from the polymer by partial acid hydrolysis. The results obtained indicate that the polysaccharide chain consists of the chemical repeating unit Gal-1,3(4)-GlcNAc-1,3-Glc-1,3-GlcNAc-, where GlcNAc stands for N-acetylglucosamine. Whereas the galactose residue is substituted at C-3 by ribitol phosphate, the glucose is substituted by ethanolamine phosphate at C-6.     

Some structural features of the teichuronic acid of Bacillus licheniformis N.C.T.C. 6346 cell walls

A teichuronic acid, containing glucuronic acid and N-acetylgalactosamine, was purified from acid extracts of Bacillus licheniformis 6346 cell walls as described by Janczura, Perkins & Rogers (1961). After reduction of the carboxyl function of glucuronic acid residues in the polysaccharide the reduced polymer contains equimolar amounts of N-acetylgalactosamine and glucose. Methylation of the reduced polysaccharide by the Hakamori (1964) technique showed the glucose residues to be substituted on C-4. A disaccharide, 3-O-glucuronosylgalactosamine, was isolated from partial acid hydrolysates of teichuronic acid. After N-acetylation the disaccharide produces chromogen readily on heating at pH7, in agreement with C-3 substitution of the reducing N-acetylamino sugar. Teichuronic acid also produces chromogen under the same conditions, with concurrent elimination of a modified polysaccharide from C-3 of reducing terminal N-acetylgalactosamine residues of the teichuronic acid chains. The number-average chain lengths of several preparations of teichuronic acid were estimated from the amounts of chromogen produced in comparison with the N-acetylated disaccharide. The values obtained are in good agreement with the weight-average molecular weight determined by ultracentrifugal analysis. The reducing terminals of teichuronic acid are shown to be exclusively N-acetylgalactosamine by reduction with sodium boro[(3)H]hydride. The number-average chain lengths of the teichuronic acid preparations were estimated by the extent of in corporation of tritium and are in agreement with values obtained by the other methods.     

The 5-O-beta-D-galactofuranosyl-containing peptidophosphogalactomannan of Penicillium charlesii. Characterization of the mannan by 13C NMR spectroscopy

Penicillium charlesii secretes a galactofuranosyl and phosphodiester-containing peptidophosphogalactomannan (pPGM). A linear mannan was prepared from pPGM by treatment with 48% aqueous HF which selectively cleaves galactofuranosyl and phosphodiesters; treatment with alkaline borohydride releases the mannan from the polypeptide. Mannan from P. charlesii cultured in D-[1,2-13C2]glucose contained mannopyranosyl residues which were enriched in 13C at both C-1 and C-2 and, to a lesser extent, at C-5 and C-6. The mannan was examined with a combination of 13C NMR INADEQUATE pulse sequence and selective 13C saturation to assign the resonance frequency of anomeric carbons directly coupled to specific C-2 signals. Three species of mannosyl residues, each substituted with a glycosidic linkage at C-2, and a fourth species substituted at C-6 and not substituted at C-2 were observed. Mannan obtained from P. charlesii cultured in D-[6-13C]glucose contained mannopyranosyl residues which were enriched in 13C primarily in C-6. The mannan was examined by DEPT 13C NMR to determine the number of species which were substituted at C-6. Mannan, treated as described above, contained a 1----6-linked mannopyranosyl species. pPGM contains minor quantities of at least four other substances attached to hydroxymethyl groups of the hexosyl residues.     

Structure of anionic carbohydrate-containing cell wall polymers in several representatives of the order actinomycetales

A new teichoic acid was identified in the cell walls of Streptomyces griseoviridis VKM Ac-622T, Streptomyces sp. VKM Ac-2091, and Actinoplanes campanulata VKM Ac-1319T. The polymer is poly(glycosylglycerol phosphate). The repeating units of the polymer, alpha-galactopyranosyl-(1-->3)-2-acetamido-2-deoxy-beta-galactopyran+ ++ osyl-(1-->1)-glycerols, are in phosphodiester linkage at C-3 of glycerol and C-6 of galactose. The structures of cell wall teichoic acids in the strains Streptomyces chryseus VKM Ac-200T and "Streptomyces subflavus" VKM Ac-484 similar in morphology and growth characteristics are also identical: 1,5-poly(ribitol phosphate) substituted at C-4(2) by 2-acetamido-2-deoxy-beta-glucopyranosyl residues and 1,3-poly(glycerol phosphate). The taxonomic aspects of these results are discussed.     

Structures of cell wall teichoic acids of <Emphasis Type="Italic">Brevibacterium iodinum</Emphasis> VKM Ac-2106<Superscript>T</Superscript>

Structures of two cell wall teichoic acids of Brevibacterium iodinum VKM Ac-2106^T were studied. The structure of mannitol teichoic acid described earlier was mainly confirmed. This polymer is 1,6-poly(mannitol phosphate) bearing -D-glucopyranosyl residues at the C-2 of mannitol and pyruvic acid residues at the C-4 and C-5. The absolute configurations of D-mannitol and S-pyruvic acid were found. The following distinctions from the earlier described structure were found: unsubstituted 1,6-poly(mannitol phosphate) residues and residues substituted only by -D-glucopyranosyl at the C-2 of mannitol but unsubstituted by pyruvic acid are present in the chain. The structure of glycerol teichoic acid present in the cell wall as a minor component (7%) is also described. This acid is identified as 1,3-poly(glycerol phosphate) substituted at the C-2 of glycerol by 2-acetamido-2-deoxy--D-galactopyranosyl residues bearing R-pyruvic acid residues at the C-4 and C-6 of galactose. This polymer is for the first time described in the cell wall of Gram-positive bacteria.Translated from Biokhimiya, Vol. 69, No. 12, 2004, pp. 1659–1666.Original Russian Text Copyright © 2004 by Potekhina, Evtushenko, Senchenkova, Shashkov, Naumova.     

Chemical structure of the lipid A component of Plesiomonas shigelloides and its taxonomical significance

Abstract The chemical structure of the lipid A moiety of the lipopolysaccharide of the type strain of Plesiomonas shigelloides was elucidated. It consists of a β-(1 → 6)-linked glucosamine disaccharide carrying phosphate groups at C-1 of the reducing and at C-4' of the non-reducing glucosamine. It contains a total of 6 residues of fatty acids, 2 amide-linked and 4 ester-linked. The amino groups of the backbone disaccharide are N -acylated by substituted 3-hydroxyacyl residues: at the reducing glucosamine by 3-O-(14:0)14:0; and at the non-reducing glucosamine by 3-O-(12:0)14:0.
Two residues of 3-hydroxytetradecanoic acid are linked to C-3 and C-3' of the glucosamine residues; the hydroxy groups of these ester-linked 3-hydroxytetradecanoic acids are unsubstituted. In free lipid A, the hydroxyl groups at C-4 and C-6' are unsubstituted, indicating that the 2-keto-3-deoxyoctonic acid (KDO) is linked to C-6' of the non-reducing glucosamine, as was shown with enterobacterial lipid A. The taxonomical significance of these structural details is discussed.     

The biofilm matrix of Pseudomonas sp. OX1 grown on phenol is mainly constituted by alginate oligosaccharides

The structure of the major constituent of the biofilm matrix produced by Pseudomonas sp. OX1, when grown on phenol as the sole carbon source is described. This investigation, carried out by chemical analysis, NMR spectroscopy and MALDI-TOF MS spectrometry, showed the presence of an oligosaccharide blend with the typical alginate structure, namely (1-->4) substituted beta-D-mannuronic (ManA) and alpha-L-guluronic acid (GulA). GulA residues were non-acetylated whereas ManA was always O-acetylated at C-2 or C-3.     

Use of 13C-n.m.r. spectroscopy for the quantitative estimation of 3-O- and 3,6-di-O-substituted D-glucopyranosyl residues in alpha-D-glucans formed by the D-glucosyltransferases of Streptococcus sobrinus

The 13C-n.m.r. spectra of the three alpha-D-glucans from Streptococcus sobrinus and the dextran from Leuconostoc mesenteroides, which differ widely in the ratios of omega (terminal, nonreducing) D-glucopyranosyl groups: 3-:6-:3,6-linked D-glucopyranosyl (Glc) residues, were measured in 0.5M NaOH at 22 degrees. The C-1 signals of 3-O-substituted Glc in a linear sequence, 6-O-substituted Glc in a linear sequence, 3,6-di-O-substituted Glc in a (1----6)-linked sequence, and Glc attached to O-3 of 3,6-di-O-substituted Glc were distinguished from each other. The C-3 signal of 3,6-linked Glc appeared downfield by 0.6 to 1.0 p.p.m. compared to the C-3 signal of 3-linked Glc in a linear sequence. The C-6 signals of omega-terminal, 3-linked, 6-linked, and 3,6-linked Glc were also assigned. The C-2 signal of 3-linked Glc in a linear sequence appeared separately, at 73.76 p.p.m. Based on these assignments, the various D-glucopyranosyl residues of the S. sobrinus alpha-D-glucans were quantitatively estimated from the signal areas of the C-2 atom of 3-linked Glc, the C-3 atom of 3-linked and 3,6-linked Glc, the C-6 atom of 6-linked and 3,6-linked Glc, and the C-6 atom of the omega-Glc groups and 3-linked Glc residues. The figures thus derived for the linkage ratios were close to those obtained by methylation analysis.     

Structure of a teichoic acid from Nocardioides luteus VKM Ac-1246T cell wall

A teichoic acid from the cell walls of Nocardioides luteus VKM Ac-1246T, a validly described species of the Nocardioides genus, is a 1,5-poly(ribitol phosphate) completely substituted at C-4 by alpha-D-galactopyranosyl residues carrying a 4,6-pyruvate ketal group in R-configuration. The structure of the repeating unit of the polymer is as follows: [figure]. The chain consists of approximately 18 repeating units and six beta-D-galactofuranosyl residues linked in the oligomer by 1,6-glycosidic bonds. The oligomer probably terminates the growing end of the teichoic acid. The structure of the polymer was determined by chemical methods and NMR spectroscopy. This teichoic acid has not been described so far.     

Analysis of bromine-oxidised dextran by 13C-n.m.r. spectroscopy

Dextran T 10, elaborated by Leuconostoc mesenteroides NRRL B-512, was oxidised with aqueous bromine at pH 7.0. The resulting oxodextran and its methoximated derivative were analysed by ¹³C-n.m.r. spectroscopy. The total amount of keto groups and their positions were established. Assignments of the ¹³C signals were made by referring to spectra of the corresponding methyl glucosiduloses and an oxodextran having most of the carbonyl groups at position 3 of the glycopyranosyl residues. In accordance with the mechanism for bromine oxidation of mono- and di-saccharides, the glucopyranosyl residues of dextran were oxidised mainly at C-2 and C-4. Over-oxidation resulted in a small proportion of acidic, ring-cleavage products.     

Oxidized saccharides as inhibitors of alpha-glucan synthesis by Streptococcus mutans glucosyltransferase

Specific inhibition by periodate-oxidized dextrans of the synthesis of alpha-glucan by S. mutans glucosyltransferase prompted a search for structurally related inhibitors that might be effective as anticaries agents. Clinical dextran derivatives in which from 5 to 50% of the D-glucose units were oxidized acted as potent and specific enzyme-inhibitors, as did 10%-oxidized derivatives of dextran fractions ranging in mol. wt. from 10(4) to 2 X 10(6). Within these limits, differences in oxidation or molecular weight did not significantly affect the high inhibitory potency of the derivatives. In contrast, periodate oxidation of (1 leads to 6)-alpha, (1 leads to 3)-alpha-, and (1 leads to 4)-alpha-linked oligosaccharides containing less than approximately 15 D-glucose units, and of sucrose and structurally related trisaccharides, yielded derivatives that were poor inhibitors. Enzymic hydrolysis of oxidized dextrans caused a loss of their inhibitory power and indicated that, to act as specific inhibitors, oxidized molecules must contain at least 16 to 20 D-glucosyl residues. The similar, minimum size required in order that unoxidized oligosaccharides may act as efficient acceptors in the glucosyltransferase reaction suggests that the inhibitory potencies of oxidized derivatives may reflect their relative abilities to bind at the acceptor site of the enzyme.     

Anionic carbohydrate-containing polymers of cell walls in two streptoverticille genospecies.

The cell walls of two streptoverticille genospecies which belong to a historically isolated group of the genus Streptomyces contain anionic polymers of different structure. Streptomyces hachijoensis VKM Ac-191T and Streptomyces cinnamoneus subsp. azacoluta VKM Ac-606T assigned to one genospecies on the basis of DNA--DNA hybridization [5] contain 37% of an identical sugar-1-phosphate polymer. The repeating disaccharide units of the polymer, 2-amino-2-deoxy-alpha-D-glucopyranosyl-(1-->6)-2-acetamido-2-deoxy-al pha-D-glucopyranose, are linked at C-1 and C-6' by phosphodiester bonds. The cell walls of Streptomyces biverticillatus VKM Ac-891T and Streptomyces baldaccii VKM Ac-821T, members of another genospecies, contain about 30% 1,3-poly(glycerol phosphate) completely substituted by 2-amino-2-deoxy-alpha-D-glucopyranosyl residues at C-2. Due to the presence of an amino sugar with a free amino group in the repeating unit, the polymers exhibit neutral properties. Polymer structures were determined by chemical methods and NMR spectroscopy. The data indicate taxonomic specificity of anionic polymers in streptoverticille cell walls.     

Isolation and partial characterization of fucan sulfates from the body wall of sea cucumber Stichopus japonicus and their ability to inhibit osteoclastogenesis

Two types of fucan sulfate were isolated from chloroform/methanol extract of the body wall of the sea cucumber Stichopus japonicus. One type (type A) contained 3.41 mmol fucose/g and 2.35 mmol sulfate/g, and the molecular mass was determined to be 9 kDa by gel permeation chromatography (GPC). Structural analysis suggested that type A consists of a backbone of (1-->3)-linked fucosyl residues that are substituted at C-4 with fucosyl residues, and that fucosyl residues are sulfated at C-2 and/or C-4. Another type (type B) contained 3.90 mmol fucose/g and 3.07 mmol sulfate/g, and the molecular mass was determined to be 32kDa by GPC. Structural analysis showed that type B is largely composed of unbranched (1-->3)-linked fucosyl residues, and that sulfate substitution(s) occur at C-2 and/or C-4. The potential of both types to inhibit osteoclastogenesis was examined by an in vitro assay system, showing that both types of fucan sulfate inhibit osteoclastogenesis more than 95% at 50 microg/mL concentration. These results suggest that types A and B fucan sulfate from sea cucumber are potent inhibitors of osteoclastogenesis.     

Anti-herpetic activity of a sulfated xylomannan from Scinaia hatei

Many viruses display affinity for cell surface heparan sulfate proteoglycans with biological relevance in virus entry. This raises the possibility of the application of sulfated polysaccharides in antiviral therapy. In this study we have analyzed polysaccharide fractions isolated from Scinaia hatei. The crude water extract (ShWE) as well as one fraction (F1) obtained by size exclusion chromatography had potent anti-HSV activity. Their inhibitory concentration 50% (IC50) values ranging from 0.5 to 4.6 microg/ml were much lower than the cytotoxic concentration 50% (CC50) values (1000 microg/ml). These fractions had very low anticoagulant activity. Furthermore, they had a weak inactivating effect on virions in a virucidal assay at concentrations in the range of 60-100 microg/ml. Chemical, chromatographic and spectroscopic methods showed that the major polysaccharide, which had 0.4 sulfate group per monomer unit and an apparent molecular mass of 160 kDa, contained a backbone of alpha-(1-->3)-linked D-mannopyranosyl residues substituted at C-6, C-4 and C-2 with single stub of beta-d-xylopyranosyl residues. Sulfate groups, when present, are located at C-4 of alpha-(1-->3)-linked D-mannopyranosyl units, and appeared to be very important for the anti-herpetic activity of this polymer.