Journal of Applied Glycoscience
Online ISSN : 1880-7291
Print ISSN : 1344-7882
ISSN-L : 1344-7882
69 巻, 3 号
選択された号の論文の3件中1~3を表示しています
Regular Paper
  • Yuitsu Otsuka, Koki Sato, Shigekazu Yano, Haruki Kanno, Wasana Suyotha ...
    2022 年 69 巻 3 号 p. 49-56
    発行日: 2022/08/22
    公開日: 2022/09/29
    [早期公開] 公開日: 2022/04/14
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    電子付録

    The GH-16 type β-1,3-glucanase (BgluC16MK) gene of Lysobacter sp. MK9-1 was cloned to study its antifungal activities. BgluC16MK displays amino acid sequence similarity with GluC from L. enzymogenes strain N4-7. BgluC16MK includes a signal sequence, a catalytic domain and carbohydrate-binding module family 6-type β-glucan binding domain (B-GBD). The expression of the BgluC16MK gene in Escherichia coli without the signal sequence resulted in antifungal activity at a dose of 0.6-0.8 nmol/disk. However, BgluC16MK displayed antifungal activity at a dose of 0.025 nmol/disk in combination with Chi19MK. Substrate-specific assay revealed that purified BgluC16MK hydrolyzed insoluble curdlan more readily than the soluble substrate. Furthermore, to explore the binding selectivity of B-GBD of BgluC16MK, we constructed a fusion protein (B-GBD-GFP) using the B-GBD and green fluorescent protein. The activity of the fusion protein against various substrates indicates that B-GBD was selective for glucans with β-1,3-linkages. An additional study demonstrated the binding ability of B-GBD-GFP to the cell-wall of living fungi, such as T. reesei and Aspergillus oryzae. These findings suggest that BgluC16MK can be utilized to generate antifungal enzyme preparations and that the fusion protein B-GBD-GFP can be used to identify the fungal cell surface structure using β-glucans.

  • Eri Kokubo, Hirofumi Sonoki, Kenta Aizawa, Hiroki Takagi, Masayasu Tak ...
    2022 年 69 巻 3 号 p. 57-63
    発行日: 2022/08/22
    公開日: 2022/09/29
    [早期公開] 公開日: 2022/04/19
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    Slowly digestible carbohydrates are needed for nutritional support in diabetic patients with malnutrition. They are a good source of energy and have the advantage that their consumption produces a low postprandial peak in blood glucose levels because they are slowly and completely digested in the small intestine. A high-amount isomaltomegalosaccharide containing carbohydrate (H-IMS), made from starch by dextrin dextranase, is a mixture of glucose polymers which has a continuous linear structure of α-1,6-glucosidic bonds and a small number of α-1,4-glucosidic bonds at the reducing ends. It has a broad degree of polymerization (DP) distribution with glucans of DP 1030 as the major component. In our previous study, H-IMS has been shown to exhibit slow digestibility in vitro and not to raise postprandial blood glucose to such levels as that raised by dextrin in vivo. This marks it out as a potentially useful slowly digestible carbohydrate, and this study aimed to evaluate its in vivo digestibility. The amount of breath hydrogen emitted following oral administration of H-IMS was measured to determine whether any indigestible fraction passed through to and was fermented in the large intestine. Total carbohydrate in the feces was also measured. H-IMS, like glucose and dextrin, did not result in breath hydrogen excretion. Carbohydrate excretion with dietary H-IMS was no different from that of glucose or water. These results show that the H-IMS is completely digested and absorbed in the small intestine, indicating its potential as a slowly digestible carbohydrate in the diet of diabetic patients.

  • Daichi Ito, Emiri Nakano, Shuichi Karita, Midori Umekawa, Khanok Ratan ...
    2022 年 69 巻 3 号 p. 65-71
    発行日: 2022/08/22
    公開日: 2022/09/29
    [早期公開] 公開日: 2022/04/20
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    Paenibacillus xylaniclasticus strain TW1, a gram-positive facultative anaerobic bacterium, was isolated as a xylanolytic microorganism from the wastes of a pineapple processing factory. A gene encoding one of its xylanolytic enzymes, a β-xylosidase, was cloned and sequenced. Sequence analysis revealed that this β-xylosidase, named PxXyl43A, was composed of a glycoside hydrolase (GH) family 43 subfamily 12 catalytic module and an unknown function module (UM). The full-length PxXyl43A (PxXyl43A) was heterologously expressed in Escherichia coli and purified. Recombinant PxXyl43A exhibited hydrolysis activity against both p-nitrophenyl-β-D-xylopyranoside (pNPX) and p-nitrophenyl-α-L-arabinofuranoside at specific activities of 250 and 310 mU/mg, respectively. The optimal reaction pH and temperature for pNPX hydrolysis were 7.1 and 54 ˚C, respectively. At pH 7.0 and 54 ˚C, the Km and kcat for pNPX were 1.2 mM and 2.8 ± 0.15 s-1, respectively. It was also discovered that the recombinant unknown function module of PxXyl43A (PxXyl43A-UM) could bind to insoluble xylans like birchwood xylan and oat spelt xylan, whereas it did not bind to cellulosic substrates such as ball-milled cellulose, carboxymethyl cellulose or lichenan. The PxXyl43A-UM's binding constant value Ka for oat spelt xylan was 2.0 × 10-5 M-1. These results suggest that PxXyl43A possesses a novel carbohydrate-binding module, named as CBM91, specific for xylan-containing polysaccharides.

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