Arabinofuranosyl Residues Research Articles

A water-soluble polysaccharide from the leaves of Litsea gardneri (Lauraceae)

A neutral polysaccharide isolated from the water extract of the leaves of Litsea gardneri was composed mainly of xylose (70%) and small, approximately equimolar amounts of arabinose, mannose, galactose and glucose. Methylation analysis combined with GLC-MS, 1[H] and 13[C]NMR spectroscopy of the native and degraded polysaccharides indicated the presence of a 1 → 4 linked β- d-xylopyranosyl backbone. It is possible that some arabinofuranosyl residues were incorporated in the backbone of the polymer. Side chains were attached to O-2 of some of the xylopyranosyl residues in the polymer chain. The available evidence suggests that the side chains may be either single units of terminal arabinofuranosyl/xylopyranosyl residues or multiple unit side chains containing both arabinosyl and xylosyl residues.

Structure identification of feruloylated oligosaccharides from sugar-beet pulp by NMR spectroscopy

1D NMR ( 1H and 13C) and 2D NMR spectroscopy have been used to determine the structure of feruloylated oligosaccharides obtained by enzymic degradation or mild acid hydrolysis of sugar-beet pulp. Feruloylated oligosaccharides derived from pectic neutral side-chains containing arabinose or galactose residues were identified. In the feruloylated arabinose oligosaccharides, feruloyl groups were linked to 0-2 of l,-Ara f residues. The structure of the feruloylated arabinose disaccharide was identified as 0-[2- 0-( trans-feruloyl)-α- l-Ara f]-(1 → 5)- l-Ara f and that of the feruloylated arabinose trisaccharide as O-α- l-Ara f-(1 → 3)- O-[2- 0-( trans-feruloyl)-α- l-Ara f]-(1 → 5)- l-Ara f. The structure of the feruloylated galactose disaccharide was identified as 0-[6- 0-( trans-feruloyl)-β- d-Gal p]-(1 → 4)- d-Gal p. From our results, we suggest that the feruloyl groups present in sugar-beet pulp are linked to the arabinofuranosyl residues of the main core of α-(1 → 5)-linked arabinan chains and to the galactopyranosyl residues of the main core of β-(1 → 4)-linked type I galactan chains.

Mode of action, kinetic properties and physicochemical characterization of two different domains of a bifunctional (1-->4)-beta-D-xylanase from Ruminococcus flavefaciens expressed separately in Escherichia coli.

Two catalytic domains, A and C, of xylanase A (XYLA) from Ruminococcus flavefaciens were expressed separately as truncated gene products from lacZ fusions in Escherichia coli. The fusion products, referred to respectively as XYLA-A1 and XYLA-C2, were purified to homogeneity by anion-exchange chromatography and chromatofocusing. XYLA-A1 was isoelectric at pH 5.0 and had a molecular mass of 30 kDa, whereas XYLA-C2 had a pI of 5.4 and a molecular mass of 44 kDa. The catalytic activity shown by both domains was optimal at 50 degrees C, but XYLA-A1 was more sensitive than XYLA-C2 to temperatures higher than the optimum. XYLA-A1 showed a higher sensitivity to pH than XYLA-C2. The enzyme activity of both domains was completely inactivated in the presence of copper or silver ions and partially inactivated by iron or zinc ions. Neither domain was active on xylo-oligosaccharides shorter than xylopentaose: the rate of degradation of longer xylo-oligosaccharides (degree of polymerization 5-10) increased as the chain length increased. Analysis of the products of hydrolysis of xylo-oligosaccharides and xylan (arabinoxylan) polysaccharide showed that the two domains differed in their modes of action: xylobiose was the shortest product of the hydrolysis. With oat spelt xylan as substrate, XYLA-A1 activity was apparently restricted to regions where xylopyranosyl residues did not carry arabinofuranosyl substituents, whereas XYLA-C2 was able to release hetero-oligosaccharides carrying arabinofuranosyl residues. Neither domain was able to release arabinose from oat spelt xylan.

Highly selective cleavage of furanosyl linkages in liquid hydrogen fluoride

Earlier experiments indicated that anhydrous liquid I-IF at Dry Ice-acetone temperatures was nearly specific for cleavage of arabinofuranosyl linkages in plant cell-walls’. Proof of this indication became necessary because of our need for a method to remove arabinofuranosyl residues from the plant cell-wall protein “ Extensin2 “, in order to render the protein susceptible to proteolysis by trypsin, without cleavage of any other sugar linkages associated with the protein. Early experiments showed incomplete specificity for furanose linkages. However, recently we have found that our norma procedures for deactivating HF, forming a relatively unreactive complex with ether, is highly exothermic3. We now report that, if precautions are taken to ensure that during the reaction the HF is maintained at temperatures below -70°C and that during quenching of the reaction equally low temperatures are maintained, only furanosyl linkages are cleaved. We have tested the lability of three different furanosides in several different compounds by comparing the ‘H and/or 13C NMR spectra of the compounds before and after such an HF treatment. Fructofiranosidtx-Sucrose and related oligomers are readily available examples containing fructofuranosyl linkages. These oligomers contain the cr-p-glucopyranosyl-(1 + 2)+?-D-fructofuranoside linkage, which we presumed would be labile at the very low temperature conditions being tested. The higher oligomers raffinose, stachyose, and melezitose contain additional less-labile linkages which could illustrate the specificity of HF at such a low temperature. Table I summarizes our results with four oligomers of this type. Fig. 1 shows the ‘H NMR spectrum of one of them, raffinose, before and after the HF treatment. In all cases

Water-unextractable cell wall material from wheat flour. 1. Extraction of polymers with alkali

Water-unextractable cell wall material (WUS) prepared from wheat flour was extracted sequentially with saturated Ba(OH) 2 , 1 M KOH and 4 M NaOH. The resultant extracts, which together contained 96% of the total WUS in the flour, were enriched in arabinoxylans, (1 → 3,1 → 4)-β-glucans and glucomannans, respectively. The remaining 4% of the wheat flour WUS was mainly cellulose. Methylation analysis of the arabinoxylans showed that the xylopyranosyl residues were substituted at 0-3 or 0-2,3 with mainly terminal arabinofuranosyl residues. Only minor amounts of 0-2 substituted xylose were found. The degree of substitution of xylopyranosyl residues was highest in the NaOH extract and lowest in the first Ba(OH) 2 extract, with that in the second Ba(OH) 2 extract and in the KOH extract being intermediate. Only minor differences in protein content and amino acid composition were observed among the extracts. Alkali-extractable arabinoxylans had slightly higher apparent molecular weights than water-extractable arabinoxylans. The alkali-extractable (1 → 3,1 → 4)-β-glucans and (gluco)mannans had substantially lower apparent molecular weights than the arabinoxylans.

Location of the mycolyl ester substituents in the cell walls of mycobacteria

The question of the precise location of mycolic acids, the single most distinctive cell wall entity of members of the Mycobacterium genus, has now been addressed. The free hydroxyl functions of the arabinogalactan component of the mycobacterial cell wall were O-methylated under conditions in which the mycolyl esters were not cleaved. Subsequent replacement of the mycolyl functions with O-ethyl groups resulted in an acid- and base-stable differentially O-alkylated surrogate polysaccharide, more amenable to analysis. Complete hydrolysis, reduction, acetylation, and gas chromatography/mass spectrometry revealed the unexpected finding that the mycolyl substituents were selectively and equally distributed on the 5-hydroxyl functions of terminal- and 2-linked arabinofuranosyl (Araf) residues. Further analysis of the O-alkylated cell wall through partial acid hydrolysis, NaB[2H]4 reduction, pentadeuterioethylation, and gas chromatography/mass spectrometry demonstrated that the mycolyl units are clustered in groups of four on the previously recognized nonreducing terminal pentaarabinosyl unit [beta-Araf-(1----2)-alpha-Araf)2-3, 5-alpha-Araf. However, only about two-thirds of the available pentasaccharide units are so substituted. Thus, the antigenicity of the arabinan component of mycobacterial cell walls may be explained by the fact that about one-third of the pentaarabinosyl units are not mycolyated and are available for interaction with the immune system. On the other hand, the extreme hydrophobicity and impenetrability of the mycobacterial cell may be explained by the same motif also acting as the fulerum for massive esterified paraffin residues. New fundamental information on the structure of mycobacterial cell walls will aid in our comprehension of its impenetrability to antibiotics and role in immunopathogenesis and persistence of disease.

Structural studies of an arabinoxylan isolated from Litsea glutinosa (Lauraceae)

A water-soluble arabinoxylan ( d-xylose and l-arabinose in the molar ratio 1.0:3.4) was isolated from the mucilaginous bark of Litsea glutinosa (Lauraceae). The results of methylation analysis, partial hydrolysis, and 1H- and 13C-n.m.r. spectroscopy indicated a backbone of (1→4)-linked β- d-xylopranosyl residues substituted at both positions 2 and 3 with side chains composed of either single or (1→3)-linked arabinofuranosyl residues. Both α- l- and β- l-arabinofuranosyl residues were present. It is possible that side chains composed of two β- l-arabinofuranosyl residues are attached mainly at O-2 of some xylosyl residues.

Composition and structural features of cell wall polysaccharides from tomato fruits

Cell wall material was isolated from the pericarp of unripe and ripe tomatoes, free from intracellular compounds and active wall degrading enzymes. The wall preparations were sequentially extracted with cyclohexane-trans-1,2-diaminetetra-acetate (CDTA) at 20°, 0.05 M Na 2CO 3 at 1°, 0.05 M Na 2CO 3 at 20°, and 0.5, 1 and 4 M KOH at 20° to leave the α-cellulose residue, which contained a significant amount of pectic material. The polysaccharides isolated from the extracts were fractionated by anion-exchange chromatography and selected fractions were subjected to methylation analysis. The CDTA-soluble pectic polysaccharides had slightly-branched rhamnogalacturonan back-bones compared with the sodium carbonate-soluble pectic polysaccharides. The side chains of the pectic polysaccharides were mainly composed of β-(1→4)-Iinked galactopyranosyl and α-(1→5)-Iinked arabinofuranosyl residues, and the evidence for this was obtained by both methylation analysis and 13C NMR spectroscopy. The major hemicellulosic polysaccharide was a xyloglucomannan and there was evidence for the occurrence of a small amount of a xylan-pectic complex. The ripe fruit contained much less pectic galactans compared with the unripe fruit, and there was a significant decrease in the content of galactan side-chains of the ripe fruit. The hemicelluloses of the unripe and ripe fruit, however, showed negligible difference in composition.

Glycoprotein allergens in pollen of timothy. IV. Structural studies of a basic glycoprotein allergen.

Heat treatment of the glycoprotein allergen with alkaline tritiated borohydride released carbohydrate material in the molecular weight (MW) range 500-2,000 daltons. The radioactivity of the highest MW carbohydrate fraction was low, indicating the presence of alkali-stable hydroxyproline-arabinose linkages. N-acetylglucosaminitol containing 3H was detected in the hydrolysate of the low-MW carbohydrate, indicating that some of the carbohydrate is bound to the protein chain through a N-glycosidic linkage between N-acetylglucosamine and asparagine. The presence of this linkage was also suggested by fast atom bombardment mass spectrometry of pronase-digested allergen. The glycoprotein seems to contain only a few carbohydrate chains made up of a total of 25 monosaccharide units. Methylation analysis indicated that the carbohydrate moiety consists of galactopyranosyl units as branching points linked through 1,3,6-positions or as terminal galactopyranosyl residues, mannopyranosyl groups as part of the chain either as 1,3- or 1,4-linked units, and arabinofuranosyl groups as branching points linked through 1,3,5-positions or as part of the chain, either as 1,3- or 1,5-linked units or as terminal arabinofuranosyl residues.

Structural features of an arabinan from cowpea ( Vigna sinensis)

The purified arabinan, isolated by 10% trichloroacetic acid extraction of cowpea endosperm, consisted of 1,5-linked l- arabinofuranosyl residues in the main chain, some of which were further involved in branching through O-2 and/or O-3.

Structural features of cell-wall polysaccharides of the carrot Daucus carota

Cell-wall material was isolated from the alcohol-insoluble residue of carrot by treatment with Pronase, phenol—acetic acid—water, and aqueous 90% methyl sulphoxide. Some pectic material was solubilised, but the major component was a highly esterified, acidic arabinogalactan. The purified cell-wall material, which contained ∼1% of protein, was sequentially extracted with water at 80°, ammonium oxalate at 80°, and m and 4 m KOH at 20°, to leave a residue of α-cellulose, which contained some pectic material. From the hot-water-soluble fraction, a major pectic polymer was isolated by anion-exchange chromatography. Methylation analysis showed that it was a rhamnogalacturonan, probably having highly branched arabinan and slightly branched galactan side-chains linked to O-4 of rhamnopyranosyl residues. An unusual feature of this pectic polymer is that it contained a small but significant proportion of 1,4-linked xylopyranosyl residues. From the alkali-soluble fractions, a range of pectic polymers associated with various amounts of xylans and possibly xyloglucans was isolated. The main linkages present in these complexes were 1,4-linked galactopyranosyluronic acid, 1,4-linked galactopyranosyl, and 1,5-linked arabinofuranosyl residues, terminal arabinofuranosyl and galactopyranosyl groups, and, in some fractions, 1,4-linked xylopyranosyl residues. The possible association of some of these polymers with proteins and phenolics is discussed.

The Mucilage of Opuntia aurantiaca

The mucilage isolated from Opuntia aurantiaca has been shown by methylation and partial hydrolysis studies to possess a highly branched structure. The mucilage is composed of 4-linked galactosyluronic acid, 2-linked rhamnosyl, and 6-linked galactosyl residues. Most of the last-named residues carry branches at C-3 or C-4, or both C-3 and C-4. The branches are composed mainly of (1→3)-linked, (1→5)-linked, and end-group arabinofuranosyl residues, and end-group xylopyranosyl residues. The mucilage possesses both acid-stable and acid-labile galactosyluronic acid residues. The ratio of galacturonic to rhamnose in the degraded polysaccharide is 1:2.

Structural and immunochemical characterization of the acidic arabinomannan of mycobacterium smegmatis

A serologically active, acidic arabinomannan has been isolated from Mycobacterium smegmatis. The polysaccharide contains approximately 56 arabinosyl and 11 mannosyl residues, and 2 phosphate, 6 monoesterified succinate, and 4 ether-linked lactate groups. After saponification to remove succinyl groups, the polysaccharide can be separated into phosphorylated (55%) and nonphosphorylated (45%) forms, the former containing a little more arabinose and a little less mannose than the latter. The structures of these polysaccharides were investigated by 1 H- and 13C-n.m.r. spectroscopy and methylation analysis, before and after selective cleavage of furanosyl linkages. The phosphorylated and nonphosphorylated forms of the polysaccharide were found to have similar, if not identical, structures. The main structural feature of the polysaccharides is the presence of chains of contiguous arabinofuranosyl residues linked α-(1→5). These chains are attached at O-4 of arabinopyranosyl residues that are present in a core region of the polysaccharide that also contains mannopyranosyl residues. Immunochemical studies demonstrated that the polysaccharide is an effective, precipitating antigen with antisera from rabbits immunized with cell walls or heat-killed cells of M. smegmatis. The polysaccharide is, however, more effective as a precipitating antigen after removal of the succinate groups, and completely ineffective after removal of arabinofuranosyl residues. The polysaccharide therefore contains an important antigen in common with the arabinogalactan lipopolysaccharide of the cell wall of the bacterium, i.e., chains of contiguous α-(1→5)-linked arabinofuranosyl residues.

A Carbohydrate-Binding Arabinogalactan-Protein From Liquid Suspension Cultures of Endosperm From Lolium multiflorum

Water-soluble arabinogalactan-proteins have been isolated from tissue-cultured rye grass endosperm cells and their culture medium by precipitation with the β-glucosyl Yariv artificial antigen. The intracellular and extracellular polymers are similar in composition and molecular size (apparent mol. wt 2.2 — 2.8 × 105). The protein portion represents up to 7% of the molecule and is rich in hydroxyproline, alanine and serine. The carbohydrate portion (84%) consists solely of arabinose (36%) and galactose (64%) and methylation analysis shows it to be a branched 1,3 : 1,6-galactan substituted by arabinofuranosyl residues. The carbohydrate compositions and methylation analyses for polymers precipitated by the β-glucosyl Yariv antigen from lima beans, cashew, tomato, silver beet and asparagus and an arabinogalactan-peptide from wheat endosperm are compared. As a group, they show homologies both in their peptide and polysaccharide portions. A histochemical method, based on precipitation with the β-glucosyl Yariv antigen, showed the binding polymers to be localized in discrete irregular vesicles in the tissue-cultured rye grass endosperm cells. Native rye grass endosperm cells show staining between starch granules, and staining is also seen in the aleurone cells of barley, wheat and rye grass.

Hydroxyproline arabinosides from suspension-cultured tobacco cell wall.

Hydroxyproline tri- and tetra-arabinosides were isolated from suspension-cultured tobacco cell wall preparation by Ba(OH)2 treatment followed by gel filtration and ion-exchange chromatography. Methylation analysis showed that they consisted of terminal, 1,2-linked and 1,3-linked arabinofuranosyl residues in molar ratios of 1: 2: 0 and 1:2: 1, respectively. A part of trans-4-hydroxy-l-proline in the arabinosides was revealed to have been isomerized into cis-4-hydroxy-d-proline during the alkaline hydrolysis.

The Structure of Plant Cell Walls

The walls of barley (Hordeum vulgare var. Himalaya) aleurone cells are composed of two major polysaccharides, arabinoxylan (85%) and cellulose (8%). The cell wall preparations contain 6% protein, but this protein does not contain detectable amounts of hydroxyproline. The arabinoxylan has a linear 1,4-xylan backbone; 33% of the xylosyl residues are substituted at the 2 and/or 3 position with single arabinofuranosyl residues. The results of in vitro cellulose binding experiments support the hypothesis that noncovalent bonds between the arabinoxylan chains and cellulose fibers play a part in maintaining wall structure. It is suggested that bonding between the arabinoxylan chains themselves is also utilized in forming the walls.