SANS measurements of semiflexible xyloglucan polysaccharide chains in water reveal their self-avoiding statistics.

We explored the behavior and the characteristics of xyloglucan polysaccharide chains extracted from tamarind seeds in aqueous media. The initial solubilization is achieved by using a 0.01 M NaOH solution. The absence of compact aggregates in the solution and the average molecular mass of the individual chains were unambiguously demonstrated by size exclusion chromatography with multi-angle light scattering detection. The composition and the stability of the solution were quantitatively checked over weeks by using liquid state nuclear magnetic resonance with DMSO as internal standard. The conformational characteristics of the chains were measured using nondestructive small-angle neutron scattering (SANS). The unambiguous determination of the Flory exponent (ν = 0.588) by SANS enabled us to directly prove that xyloglucan chains in water behave like semiflexible worm-like chains with excluded volume statistics (good solvent), contrary to most of the neutral water-soluble polymer chains that rather exhibit Gaussian statistics (θ-solvent). In addition to the Flory exponent, the persistence length l(p) and the cross section of the chains were also determined by SANS with utmost precision, with values of 80 and of 7 Å, respectively, which provides a complete description of the conformational characteristics of XG chains at all relevant length scales.

Enthalpic studies of xyloglucan-cellulose interactions.

We report a study of xyloglucan (XG)-cellulose interactions made possible by the preparation of various well-defined cellulosic and xyloglucosidic substrates. Bacterial microcrystalline cellulose (BMCC) as well as cellulose whiskers (CellWhisk) were used as cellulosic substrates. Xyloglucosidic substrates were obtained from Rubus cells and Tamarindus indica seeds. Different primary structure characteristics of XGs such as the backbone length and the nature of the side chains, as well as their repartition, were considered in order to examine the influence of the primary structure on their interaction capacity. Two complementary approaches were carried out: first, the determination of adsorption isotherms and its associated models, and second, an enthalpic study using isothermal titration calorimetry (ITC). This study highlighted that an increase of XG interaction capacity occurred with increasing XG molecular weight. Furthermore, we determined that a minimum of 12 glucosyl residues on the backbone is required to observe significant interactions. Moreover, both the presence of trisaccharidic side chains with fucosyl residues and an increase of unsubstituted glucosyl residues enhanced XG-cellulose interactions. The evolution of adsorption isotherms with temperature and ITC measurements showed that two different processes were occurring, one exothermic and one endothermic, respectively. Although the presence of an exothermic interaction mechanism has long been established, the presence of an endothermic interaction mechanism has never been reported.

Non-electrostatic building of biomimetic cellulose-xyloglucan multilayers.

Layer-by-layer assembly was used to build thin films, consisting of multiple layers alternating cellulose nanocrystals and xyloglucan, benefiting from the strong non-electrostatic cellulose-xyloglucan interaction. Data from atomic force microscopy and neutron reflectivity showed that these well-defined films exhibited a thickness increasing linearly with the number of layers, without increase in surface roughness. These "green" nanocomposite films, reminiscent of plant cell wall, are composed of a regular stack of single layers of cellulose nanocrystals separated by very thin xyloglucan spacers. Such architecture differs from the one formed by cellulose/polycations multilayers, where the cellulose phase itself consists of a double layer.

Contributions of hemicellulose, cellulose and lignin to the mass and the porous properties of chars and steam activated carbons from various lignocellulosic precursors.

In this study, contributions of hemicellulose, cellulose and lignin to the mass and the porous properties of chars and activated carbons from various lignocellulosic materials were studied. A predictive calculation was established using the experimental results obtained for the three components separately to evaluate the carbonization and activation yields and their respective contributions to the chars and to the subsequent activated carbons of various precursors in term of weight fraction. These equations were validated. The results showed that lignin can be considering as being the major contributor of all chars and activated carbons. Besides, the evolution of the mean pore size versus the specific porous volume showed that each component contributes to the porosity of chars and activated carbons whatever is its weight contribution.

Identification of an Arabidopsis gene encoding a GH95 alpha1,2-fucosidase active on xyloglucan oligo- and polysaccharides.

alpha1,2-linked fucose can be found on xyloglucans which are the main hemicellulose compounds of dicotyledons. The fucosylated nonasaccharide XXFG derived from xyloglucans plays a role in cell signaling and is active at nanomolar concentrations. The plant enzyme acting on this alpha1,2-linked fucose residues has been previously called fucosidase II; here we report on the molecular identification of a gene from Arabidopsis thaliana (At4g34260 hereby designed AtFuc95A) encoding this enzyme. Analysis of the predicted protein composed of 843 amino acids shows that the enzyme belongs to the glycoside hydrolase family 95 and has homologous sequences in different monocotyledons and dicotyledons. The enzyme was expressed recombinantly in Nicotiana bentamiana, a band was visible by Coomassie blue staining and its identity with the alpha1,2-fucosidase was assessed by an antibody raised against a peptide from this enzyme as well as by peptide-mass mapping. The recombinant AtFuc95A is active towards 2-fucosyllactose with a Km of 0.65 mM, a specific activity of 110 mU/mg and a pH optimum of 5 but does not cleave alpha1,3, alpha1,4 or alpha1,6-fucose containing oligosaccharides and p-nitrophenyl-fucose. The recombinant enzyme is able to convert the xyloglucan fragment XXFG to XXLG, and is also active against xyloglucan polymers with a Km value for fucose residues of 1.5mM and a specific activity of 36 mU/mg. It is proposed that the AtFuc95A gene has a role in xyloglucan metabolism.

Structural characterization of mesquite (Prosopis velutina) gum and its fractions.

Structural and physicochemical characteristics of mesquite gum (from Prosopis velutina) were investigated using FT-IR spectroscopic, mass spectrometric and chromatographic methods. Four fractions (F-I, F-IIa, F-IIb and F-III) were isolated by hydrophobic interaction chromatography. The samples were characterized and analyzed for their monosaccharide and oligomers composition by high performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). L-Arabinose (L-Ara) and D-galactose (D-Gal) were found as the main carbohydrate constituent residues in the polysaccharides from mesquite gum and their ratio (L-Ara/D-Gal) varied within the range 2.54 to 3.06 among the various fractions. Small amounts of D-glucose (D-Glc), D-mannose (D-Man) and D-xylose (D-Xyl) were also detected, particularly in Fractions IIa, IIb and III. Infrared spectroscopy identified polysaccharides and protein in all the samples. Data from mass spectrometry (MALDI-TOF MS) was consistent with the idea that the structure corresponding to the periphereal chains of Fraction I is predominantly a chain of pentoses attached to uronic acid.

Characterization of gtf, a glucosyltransferase gene in the genomes of Pediococcus parvulus and Oenococcus oeni, two bacterial species commonly found in wine.

"Ropiness" is a bacterial alteration in wines, beers, and ciders, caused by beta-glucan-synthesizing pediococci. A single glucosyltransferase, Gtf, controls ropy polysaccharide synthesis. In this study, we show that the corresponding gtf gene is also present on the chromosomes of several strains of Oenococcus oeni isolated from nonropy wines. gtf is surrounded by mobile elements that may be implicated in its integration into the chromosome of O. oeni. gtf is expressed in all the gtf(+) strains, and beta-glucan is detected in the majority of these strains. Part of this beta-glucan accumulates around the cells forming a capsule, while the other part is liberated into the medium together with heteropolysaccharides. Most of the time, this polymer excretion does not lead to ropiness in a model medium. In addition, we show that wild or recombinant bacterial strains harboring a functional gtf gene (gtf(+)) are more resistant to several stresses occurring in wine (alcohol, pH, and SO(2)) and exhibit increased adhesion capacities compared to their gtf mutant variants.

Changes in red wine soluble polysaccharide composition induced by malolactic fermentation.

The polysaccharide content of wine is generally assumed to originate from grapes and yeasts, independent of bacterial metabolism, except for the action of certain spoilage species. This study shows that malolactic fermentation (MLF) significantly modifies the soluble polysaccharide (SP) concentration of various red Bordeaux wines. Wines with the highest initial SP concentration go on to present decreased SP concentration, whereas those with the lowest initial SP concentration rather go on to have a higher SP concentration after MLF. These tendencies were observed whatever the Oenococcus oeni strain (indigenous or starter) used for MLF. Neutral and charged SPs were affected, but to a degree that depended on the microorganisms driving the MLF. The SP modifications were directly linked to bacterial development, because non MLF controls did not present any significant change of SP concentration.

The fucose-binding lectin from Ralstonia solanacearum. A new type of beta-propeller architecture formed by oligomerization and interacting with fucoside, fucosyllactose, and plant xyloglucan.

Plant pathogens, like animal ones, use protein-carbohydrate interactions in their strategy for host recognition, attachment, and invasion. The bacterium Ralstonia solanacearum, which is distributed worldwide and causes lethal wilt in many agricultural crops, was shown to produce a potent L-fucose-binding lectin, R. solanacearum lectin, a small protein of 90 amino acids with a tandem repeat in its amino acid sequence. In the present study, surface plasmon resonance experiments conducted on a series of oligosaccharides show a preference for binding to alphaFuc1-2Gal and alphaFuc1-6Gal epitopes. Titration microcalorimetry demonstrates the presence of two binding sites per monomer and an unusually high affinity of the lectin for alphaFuc1-2Gal-containing oligosaccharides (KD = 2.5 x 10(-7) M for 2-fucosyllactose). R. solanacearum lectin has been crystallized with a methyl derivative of fucose and with the highest affinity ligand, 2-fucosyllactose. X-ray crystal structures, the one with alpha-methyl-fucoside being at ultrahigh resolution, reveal that each monomer consists of two small four-stranded anti-parallel beta-sheets. Trimerization through a 3-fold or pseudo-3-fold axis generates a six-bladed beta-propeller architecture, very similar to that previously described for the fungal lectin of Aleuria aurantia. This is the first report of a beta-propeller formed by oligomerization and not by sequential domains. Each monomer presents two fucose binding sites, resulting in six symmetrically arranged sugar binding sites for the beta-propeller. Crystals were also obtained for a mutated lectin complexed with a fragment of xyloglucan, a fucosylated polysaccharide from the primary cell wall of plants, which may be the biological target of the lectin.

Impact and efficiency of GH10 and GH11 thermostable endoxylanases on wheat bran and alkali-extractable arabinoxylans.

The results of a comparative study of two thermostable (1-->4)-beta-xylan endoxylanases using a multi-technical approach indicate that a GH11 xylanase is more useful than a GH10 xylanase for the upgrading of wheat bran into soluble oligosaccharides. Both enzymes liberated complex mixtures of xylooligosaccharides. 13C NMR analysis provided evidence that xylanases cause the co-solubilisation of beta-glucan, which is a result of cell-wall disassembly. The simultaneous use of both xylanases did not result in a synergistic action on wheat bran arabinoxylans, but instead led to the production of a product mixture whose profile resembled that produced by the action of the GH10 xylanase alone. Upon treatment with either xylanase, the diferulic acid levels in residual bran were unaltered, whereas content in ferulic and p-coumaric acids were unequally decreased. With regard to the major differences between the enzymes, the products resulting from the action of the GH10 xylanase were smaller in size than those produced by the GH11 xylanase, indicating a higher proportion of cleavage sites for the GH10 xylanase. The comparison of the kinetic parameters of each xylanase using various alkali-extractable arabinoxylans indicated that the GH10 xylanase was most active on soluble arabinoxylans. In contrast, probably because GH11 xylanase can better penetrate the cell-wall network, this enzyme was more efficient than the GH10 xylanase in the hydrolysis of wheat bran. Indeed the former enzyme displayed a nearly 2-fold higher affinity and a 6.8-fold higher turnover rate in the presence of this important by-product of the milling industry.