Journal of applied glycoscience最新文献

Wheat flour-based batter containing 0 to 20 % trehalose was deep-fried, dried and held in various water activity (a _w) conditions. The effects of trehalose content and a _w on oil content, water sorption, isothermal mechanical relaxation, and fracture properties were investigated. For comparison, the fracture properties of freeze-dried porous waxy corn starch solids were also investigated. The 10 % trehalose sample had the lowest oil content, water content, and a _w. A force-reduction value (∆F) of the samples was evaluated as a typical mechanical relaxation parameter. ∆F gradually increased with increasing a _w and sharply increased above a specific a _w presumed to be associated with the glass to rubber transition. Compared to ∆F values among the glassy samples, 10 and 20 % trehalose samples had higher ∆F values (were more rigid) than 0 and 5 % trehalose samples. From the fracture measurements of the glassy samples, the first fracture force increased linearly and the number of fracture peaks decreased linearly with increasing a _w. At each a _w, 10 % trehalose had the lowest first fracture force and the highest the number of fracture peaks. Freeze-dried porous waxy corn starch solids showed similar fracture properties to deep-fried samples. These findings suggest that around 10 % trehalose content is optimal for producing deep-fried foods with a brittle texture.

Pulverization is a potentially powerful solution for the resource management of surplus- and non-standard agricultural products, maintaining their nutritional values for long and ensuring their homogeneity, whereas their original textures could disappear to narrow the application ranges. Therefore, new technologies should be developed for reconstructing the powders to provide them with new physical characteristics. Herein, we developed a novel food material, nata puree (NP), by nata de coco (bacterial cellulose gel) disintegration with a water-soluble polysaccharide using a household blender. The process worked well with (1,3)(1,4)-β-glucan (BGL) as the polysaccharide, which could be substituted with barley extract. Lichenase treatment of the NP dramatically modified its physical properties, suggesting the importance of the BGL polymeric forms. NP exhibited distinct potato powder and starch binding activities, which would be attributed to its interactions with the cell wall components and a physical capture of powders by the NP network, respectively. NP supplementation into the potato paste improved its firmness and enabled its printable range shift for 3D food printing to a lower powder-concentration. NP also promoted the dispersion of powders in its suspension, and designed gelation could also be successfully performed by the laser irradiation of an NP suspension containing dispersed curdlan and turmeric powders. Therefore, NP could be applied as a powder modifier to a wide range of products in both conventional cooking, food manufacturing, and next generation processes such as 3D food printing.

Carbohydrate materials that produce lower postprandial blood glucose increase are required for diabetic patients. To develop slowly digestible carbohydrates, the effect of degree of polymerization (DP) of α-1,6 glucan on its digestibility was investigated in vitro and in vivo. We prepared four fractions of α-1,6 glucan composed primarily of DP 3-9, DP 10-30, DP 31-150, and DP 151+ by fractionating a dextran hydrolysate. An in vitro experiment using digestive enzymes showed that the glucose productions of DP 3-9, DP 10-30, DP 31-150, and DP 151+ were 70.3, 53.4, 28.2, and 19.2 % in 2 h, and 92.1, 83.9, 39.6, and 33.3 % in 24 h relative to dextrin, respectively. An in vivo glycemic response showed that the incremental area under the curve (iAUC) of blood glucose levels of α-1,6 glucan with DP 3-9, DP 10-30, DP 31-150, and DP 151+ were 99.5, 84.3, 65.4, and 40.1 % relative to dextrin, respectively. These results indicated that α-1,6 glucan with higher DP had stronger resistance to digestion and produced a smaller blood glucose response. DP 10-30 showed significantly lower maximum blood glucose levels than dextrin; however, no significant difference was observed in iAUC, indicating that DP 10-30 was slowly digestible. In addition, α-1,6 glucan was also produced using an enzymatic reaction with dextrin dextranase (DDase). This produced similar results to DP 10-30. The DDase product can be synthesized from dextrin at low cost. This glucan is expected to be useful as a slowly digestible carbohydrate source.

Erianthus arundinaceus (ER) is greatly appreciated among domestic energy crops in Japan for the production of fermentable sugars from lignocellulosic polysaccharides. In this study, we developed an efficient Ca(OH)₂-based pretreatment of both stems and leaves of ER at ambient temperature with the addition of a washing step for enzymatic saccharification. The recoveries of glucans and xylans in the pretreated ER after four countercurrent washing cycles were 91 and 76 %, respectively, the former being considerably higher than that of rice straw (RS) (72 %). Their saccharification ratios in the washed sample under the pressure of 1 atm CO₂ were 80 and 92.5 %, respectively. The application of this simple sugar production process from ER would further support the domestic bioprocess development. ER is also foreseen to provide the additional feedstock favorable for harvesting from winter to spring in Japan, preventing a risk for feedstock shortage generated by single harvesting such as RS.

We recently characterized a 3-O-α-D-galactosyl-α-L-arabinofuranosidase (GAfase) for the release of α-D-Gal-(1→3)-L-Ara from gum arabic arabinogalactan protein (AGP) in Bifidobacterium longum subsp. longum JCM7052. In the present study, we cloned and characterized a neighboring α-galactosidase gene (BLGA_00330; blAga3). It contained an Open Reading Frame of 2151-bp nucleotides encoding 716 amino acids with an estimated molecular mass of 79,587 Da. Recombinant BlAga3 released galactose from α-D-Gal-(1→3)-L-Ara, but not from intact gum arabic AGP, and a little from the related oligosaccharides. The enzyme also showed the activity toward blood group B liner trisaccharide. The specific activity for α-D-Gal-(1→3)-L-Ara was 4.27- and 2.10-fold higher than those for melibiose and raffinose, respectively. The optimal pH and temperature were 6.0 and 50 °C, respectively. BlAga3 is an intracellular α-galactosidase that cleaves α-D-Gal-(1→3)-L-Ara produced by GAfase; it is also responsible for a series of gum arabic AGP degradation in B. longum JCM7052.

Glycogen is a highly branched storage polysaccharide found mainly in the liver and the muscles. Glycogen is also present in the skin, but its functional role is poorly understood. Recently, it has been reported that glycogen plays an important role in intracellular signal transduction. In the epidermis of the skin, keratinocytes are the predominant cells that produce ceramide. Ceramides are lipids composed of sphingosine, and prevent water loss, as well as protecting the skin against environmental stressors. In this study, we investigated the effects of glycogen on ceramide production in cultured keratinocytes. Thin-layer chromatography revealed that incubation of keratinocytes with 2 % glycogen enhanced the cellular amount of ceramide NS (ceramide 2) by 3.4-fold compared to the control. We also found that glycogen regulated the mRNA expression levels of signaling molecules of the sphingomyelin-ceramide pathway by quantitative real-time PCR. The activity of sphingomyelinase was also significantly enhanced by 2.5-fold in cultures with 1 % glycogen compared to the control. Moreover, glycogen increased the ATP production by 1.5-fold compared to the control, while glucose did not affect the production. Western blotting showed that phosphorylation of Akt, a cellular signaling molecule, was inhibited in the presence of glycogen in cultured keratinocytes. This study shows that glycogen upregulates the ceramide production pathway from sphingomyelin in epidermal keratinocytes, and provides new insights into the role of glycogen in cellular signal transduction.

Human urinary trypsin inhibitor (UTI) is a proteoglycan composed of one core protein covalently linked to one glycosaminoglycan, which is a low sulfated chondroitin 4-sulfate. It is used as an anti-inflammatory medicine based on the protease inhibitory activity of the core protein. However, functions of the chondroitin sulfate have not been clarified. Recently, we succeeded in remodeling the UTI chondroitin sulfate to hyaluronan to create hyaluronan hybrid UTI, without changing the core protein. Here, we investigated the effect of the remodeled chondroitin sulfate on the activities of serine proteases. Native UTI showed stronger protease inhibitory activity than hyaluronan hybrid UTI or hydrolyzed glycosaminoglycan UTI. Chondroitin 4-sulfate chains with a small peptide derived from the native UTI did not show any protease inhibitory activity. These results suggest that the chondroitin sulfate chain linked covalently to core protein enhances protease inhibitor activity of UTI although the chondroitin sulfate chain itself does not.

Starch biosynthetic enzymes form multi-protein complexes consisting of starch synthase (SS) I, SSIIa, and starch branching enzyme (BE) IIb, which synthesize amylopectin clusters. This study analyzed the starch properties in two double mutant rice lines lacking SSIIa and BEIIb, one of which expressed an inactive BEIIb protein. The ss2a be2b lines showed similar or greater seed weight than the be2b lines, and plant growth was not affected. The ss2a line showed increased short amylopectin chains resulting in a lower gelatinization temperature. Starch granule morphology and A-type crystallinity were similar between the ss2a line and the wild type, except for a mild chalky seed phenotype in the ss2a line. However, the starch phenotype of the ss2a be2b lines, which was similar to that of be2b but not ss2a, was characterized by increased long amylopectin chains, abnormal starch granules, and B-type crystallinity. The similarity in phenotype between the ss2a be2b and be2b lines may be attributed to the inability of the be2b mutants to generate short amylopectin branches, which serve as primers for SSIIa. Therefore, the presence or absence of SSIIa hardly affected the amylopectin structure under the be2b background. The amylose content was significantly higher in the ss2a be2b lines than in the be2b lines. Starch crystallinity was greater in ss2a be2b lines than in be2b lines, despite the fact that starch crystallinity is generally negatively correlated with amylose content. This suggests that the formation of a double helix between long amylopectin chains and amylose affects starch crystallinity in the ss2a be2b mutants.

Cellobiohydrolase I from Trichoderma reesei ( Tr Cel7A) is one of the best-studied cellulases, exhibiting high activity towards crystalline cellulose. Tryptophan residues at subsites -7 and -4 (Trp40 and Trp38 respectively) are located at the entrance and middle of the tunnel-like active site of Tr Cel7A, and are conserved among the GH family 7 cellobiohydrolases. Trp40 of Tr Cel7A is important for the recruitment of cellulose chain ends on the substrate surface, but the role of Trp38 is less clear. Comparison of the effects of W38A and W40A mutations on the binding energies of sugar units at the two subsites indicated that the contribution of Trp38 to the binding was greater than that of Trp40. In addition, the smooth gradient of binding energy was broken in W38A mutant. To clarify the importance of Trp38, the activities of Tr Cel7A WT and W38A towards crystalline cellulose and amorphous cellulose were compared. W38A was more active than WT towards amorphous cellulose, whereas its activity towards crystalline cellulose was only one-tenth of that of WT. To quantify the effect of mutation at subsite -4, we measured kinetic parameters of Tr Cel7A WT, W40A and W38A towards cello-oligosaccharides. All combinations of enzymes and substrates showed substrate inhibition, and comparison of the inhibition constants showed that the Trp38 residue increases the velocity of substrate intake ( k _on for forming productive complex) from the minus side of the subsites. These results indicate a key role of Trp38 residue in processively loading the reducing-end of cellulose chain into the catalytic tunnel.

Most functional biopharmaceuticals such as antibodies are glycoproteins carrying N-linked oligosaccharides (N-glycans). In animal cells, these glycans are generally expressed as heterogeneous glycoforms that are difficult to separate into a pure form. The structure of these glycans directly affects several biological aspects of the glycoproteins, especially binding affinity. Therefore, the preparation of glycoproteins with well-defined and homogeneous glycoforms is necessary for functional studies and improved efficacy, particularly for biopharmaceuticals. This review describes the recent remarkable progress in the development and production of biopharmaceutical glycan-modified antibodies, through the use of glycan remodeling using microbial endoglycosidases and sophisticated glycoengineering techniques utilizing microbial enzymatic reaction mechanisms.