QUASIMODO1 is expressed in vascular tissue of Arabidopsis thaliana inflorescence stems, and affects homogalacturonan and xylan biosynthesis.

An insertion in the promoter of the Arabidopsis thaliana QUA1 gene (qua1-1 allele) leads to a dwarf plant phenotype and a reduction in cell adhesion, particularly between epidermal cells in seedlings and young leaves. This coincides with a reduction in the level of homogalacturonan epitopes and the amount of GalA in isolated cell walls (Bouton et al., Plant Cell 14: 2577 2002). The present study was undertaken in order to investigate further the link between QUA1 and cell wall biosynthesis. We have used rapidly elongating inflorescence stems to compare cell wall biosynthesis in wild type and qua1-1 mutant tissue. Relative to the wild type, homogalacturonan alpha-1-4-D-galacturonosyltransferase activity was consistently reduced in qua1-1 stems (by about 23% in microsomal and 33% in detergent-solubilized membrane preparations). Activities of beta-1-4-D-xylan synthase, beta-1-4-D-galactan synthase and beta-glucan synthase II activities were also measured in microsomal membranes. Of these, only beta-1-4-D-xylan synthase was affected, and was reduced by about 40% in qua1-1 stems relative to wild type. The mutant phenotype was apparent in inflorescence stems, and was investigated in detail using microscopy and cell wall composition analyses. Using in situ PCR techniques, QUA1 mRNA was localized to discrete cells of the vascular tissue and subepidermal layers. In mutant stems, the organization of these tissues was disrupted and there was a modest reduction in homogalacturonan (JIM5) epitopes. This study demonstrates a specific role for QUA1 in the development of vascular tissue in rapidly elongating inflorescence stems and supports a role of QUA1 in pectin and hemicellulose cell wall synthesis through affects on alpha-1,4-D-galacturonosyltransferase and beta-1,4-D-xylan synthase activities.

Molecular characterisation of a membrane-bound galactosyltransferase of plant cell wall matrix polysaccharide biosynthesis.

Galactomannan biosynthesis in vitro is catalysed by membrane preparations from developing fenugreek seed endosperms. Two enzymes interact: a GDP-mannose dependent (1-->4)-beta-D-mannan synthase and a UDP-galactose dependent (1-->6)-alpha-D-galactosyltransferase. The statistical distribution of galactosyl substituents along the mannan backbone, and the degree of galactose substitution of the primary product of galactomannan biosynthesis appear to be regulated by the specificity of the galactosyltransferase. We now report the detergent solubilisation of the fenugreek galactosyltransferase with retention of activity, the identification on gels of a putative 51 kDa galactosyltransferase protein, and the isolation, cloning and sequencing of the corresponding cDNA. The solubilised galactosyltransferase has an absolute requirement for added acceptor substrates. Beta-(1-->4)-linked D-manno-oligosaccharides with chain lengths greater than or equal to 5 acted as acceptors, as did galactomannans of low to medium galactose-substitution. The putative galactosyltransferase cDNA encodes a 51282 Da protein, with a single transmembrane alpha helix near the N terminus. We have also confirmed the identity of the galactosyltransferase by inserting the cDNA in frame into the genome of the methylotrophic yeast Pichia pastoris under the control of an AOX promoter and the yeast alpha secretion factor and observing the secretion of galactomannan alpha-galactosyltransferase activity. Particularly high activities were observed when a truncated sequence, lacking the membrane-spanning helix, was expressed.

A new family of oligosaccharides from the xyloglucan of Hymenaea courbaril L. (Leguminosae) cotyledons.

The xyloglucan from cotyledons of Hymenaea courbaril was hydrolysed with endo-(1,4)-beta-D-glucanase (cellulase) and analysed by TLC and HPAEC. The limit digest was different from those obtained from xyloglucans of Tamarindus indica and Copaifera langsdorffii. On treatment with nasturtium beta-galactosidase, two main oligosaccharides were detected by TLC and HPAEC. Using a process of enzymatic sequencing involving alternate treatments with a pure xyloglucan oligosaccharide-specific alpha-xylosidase, and a pure beta-glucosidase, both from nasturtium, their structures were deduced to be XXXG and a new oligosaccharide XXXXG. These structures were confirmed by 1H NMR. The relative proportions of XXXG and XXXXG indicate that approximately half of the subunits in Hymenaea xyloglucan are based on the new oligosaccharides. In the native polymer the XXXXG subunits are likely to carry galactosyl substituents in varying proportions, since cellulase hydrolysates contained many bands which were converted to XXXXG on hydrolysis with nasturtium beta-galactosidase. Although no comparative studies on the physico-chemical properties of Hymenaea courbaril xyloglucan have yet been performed, our results indicate that this polymer is less interactive with iodine when compared with T. indica and C. langsdorffii xyloglucans, suggesting that changes in conformation may occur due to the presence of XXXXG.

A new polysaccharide from a traditional Nigerian plant food: Detarium senegalense Gmelin.

The seed flour of an African leguminous plant, Detarium senegalense Gmelin, is used traditionally in Nigeria as a thickening agent in foods. Recent studies have shown that the detarium seed contains a large amount of water-soluble, non-starch polysaccharide (s-NSP), which suggests it has important nutritional properties. The aims of the present study were to characterise the structure and solution properties of purified s-NSP. The main monosaccharide residues of the extracted s-NSP were glucose, xylose, and galactose in the ratio of 1.39:1.00:0.52, suggesting structural similarity to the xyloglucan group of cell wall storage polysaccharides. This was confirmed by comparing the oligosaccharides released on endo-(1 --> 4)-beta-D-glucanase digestion with those obtained from tamarind seed xyloglucan. The intrinsic viscosity [eta] of a sample of the detarium polysaccharide was found to be 8.9 dl/g, indicating that the sample was of high molecular weight, a result confirmed by light scattering. Histochemical examination of detarium seed using bright field and epifluorescence microscopy showed the presence of xyloglucan in highly thickened cell walls, which were particularly prominent at the cell junctions.

Marginal fit and microleakage of indirect inlay systems.

Three different indirect adhesive inlay systems were investigated for marginal fit and microleakage. MOD cavities were prepared in 67 extracted sound premolar teeth initially with parallel cavity walls. Thirty-four of the premolars had the cavity walls modified to a 10 degrees and 33 to a 15 degrees occlusal divergence. EOS, Isosit and Hi-Ceram porcelain inlays did not fit cavities prepared with parallel walls. All inlays in the Isosit and porcelain groups were able to be seated without modifications in cavities prepared with either a 10 degrees or 15 degrees occlusal divergence. Inlays made by the EOS system required considerable modification of the fitting surface in order for them to fit. Results showed the EOS system to have the largest marginal gaps before cementation (P < 0.001). There was no significant difference in leakage between the three inlay systems after cementation (P < 0.05). The cementum margins leaked more than margins finished on enamel for all inlay systems (P < 0.05). It is concluded from this study that inlay cavities for Isosit and porcelain require at least a 10 degrees occlusal divergence of cavity walls to permit the fitting of inlays without unacceptable modification. The EOS inlays required considerable trimming even when a 15 degrees occlusal divergence of the cavity walls was used.

The treatment of erosion using porcelain veneers.

Two cases reported here demonstrate the use of porcelain veneers to restore lost tooth tissue due to erosion. The veneers have advantages compared with other restorations in terms of aesthetics and resistance to wear. The patients described in these two reports have been seen over periods of two and three years and have been satisfied with the results, with each reporting no further pain.

Comparison of total nutrient admixture stability using two intravenous fat emulsions, Soyacal and Intralipid 20%.

The safe clinical use and physical stability of a total nutrient admixture (TNA) system containing a soybean oil emulsion (Intralipid) has been reported. A study was conducted to compare the physical stability of four admixtures which were divided into two groups based upon the ratios (1:1:1 and 2:1:1) of amino acids (8.5%), dextrose (70%), and fat emulsion (20%). The fat component in each group contained either a new soy bean fat emulsion, Soyacal, or Intralipid. The quantities of all electrolytes, trace elements and vitamin additives were the same. All solutions were stored at 4 degrees C for 28 days and then held at ambient temperature for 5 days for a total 33-day study period. Each admixture was serially analyzed on days 0, 1, 2, 3, 5, 7, 14, 21, 28, 29, 30, 31, 32, and 33. Examination of gross visual appearance and determinations of pH were performed. Osmolality was measured by means of freezing point depression (Advanced Digimatic Osmometer, Advanced Instruments, Inc., Needham Heights, MA). A Brookhaven particle analyzer was used to measure lipid particle size and particle size distribution. Electron and light microscopy were used to verify maximum particle size and distribution on days 0, 7, 14, 21, 28, and 29. The type of lipid emulsion used did not affect the pH or osmolality of the admixtures. Admixtures prepared with the 2:1:1 ratio had slightly higher pH (0.07) and lower osmolality (350 mOsm/kg). The range of mean diameters for the admixtures prepared with Soyacal and Intralipid were 0.283 to 0.310 micron and 0.314 to 0.351 micron, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)