著者

Rikako Tsukida Makoto Yoshida Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

pp.jag.JAG-2022_0009, (Released:2022-12-26)

被引用文献数

3

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Woody biomass is anticipated to be a resource for a decarbonized society, but the difficulty of isolating woody components is a significant challenge. Brown-rot fungi, a type of wood rotting fungi, decompose hemicellulose particularly efficiently. However, there are few reports on the hemicellulases from brown-rot fungi. An α-L-arabinofuranosidase belonging to glycoside hydrolase family 51 (GH51) from the brown-rot fungus Gloeophyllum trabeum (GtAbf51A) was cloned and characterized in the present study. Analyses of the phylogeny of GH51 enzymes in wood rotting fungi revealed the existence of two groups, intercellular and extracellular enzymes. After deglycosylation, the recombinant GtAbf51A produced by Pichia pastoris appeared on SDS-PAGE as approximately 71,777 daltons, which is the expected molecular weight based on the amino acid sequence of GtAbf51A. Maximum enzyme activity occurred between pH 2.2 and 4.0 and at 50°C, while it was stable between pH 2.2 and 10.0 and up to 40°C. Due to the presence of a signal peptide, GtAbf51A was thought to hydrolyze polysaccharide containing arabinose. However, the hydrolysis rate of arabinosyl linkages in polysaccharides was only 3%–4% for arabinoxylan and 17% for arabinan. GtAbf51A, in contrast, efficiently hydrolyzed arabinoxylooligosaccharides, particularly O-α-L-arabinofuranosyl-(1→3)-O-β-D-xylopyranosyl-(1→4)-β-Dxylopyranose, which is the principal product of GH10 β-xylanase. These data suggest that GtAbf51A cooperates with other xylan-degrading enzymes, such as β-xylanase, to degrade xylan in nature.

著者

Kentaro Suzuki Mari Michikawa Haruna Sato Masahiro Yuki Kei Kamino Wataru Ogasawara Shinya Fushinobu Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

vol.65, no.2, pp.13-21, 2018-05-20 (Released:2018-05-20)

参考文献数

40

被引用文献数

9

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Highly thermostable β-mannanase, belonging to glycoside hydrolase family 5 subfamily 7, was purified from the culture supernatant of Talaromyces trachyspermus B168 and the cDNA of its transcript was cloned. The recombinant enzyme showed maximal activity at pH 4.5 and 85 °C. It retained more than 90 % of its activity below 60 °C. Obtaining the crystal structure of the enzyme helped us to understand the mechanism of its thermostability. An antiparallel β-sheet, salt-bridges, hydrophobic packing, proline residues in the loops, and loop shortening are considered to be related to the thermostability of the enzyme. The enzyme hydrolyzed mannans such as locust bean gum, carob galactomannan, guar gum, konjac glucomannan, and ivory nut mannan. It hydrolyzed 50.7 % of the total mannans from coffee waste, producing mannooligosaccharides. The enzyme has the highest optimum temperature among the known fungal β-mannanases and has potential for use in industrial applications.

著者

Sosyu Tsutsui Tomohiro Hatano Ryo Funada Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

pp.jag.JAG-2022_0008, (Released:2022-10-08)

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In recent years, the importance of biomass utilization has increased, but it has not been effectively exploited. In particular, it is difficult to use hemicellulose, the second most abundant biopolymer of biomass. Therefore, in order to promote the utilization of hemicellulose, we screened for microorganisms capable of producing polysaccharides from xylose. The following four strains were selected from samples collected from various regions of Okinawa Prefecture: Kosakonia sp. (SO_001), Papiliotrema terrestris (SO_005), Pseudoarthrobacter sp. (SO_006), and Williamsia sp. (SO_009). Observation with a scanning electron microscope (SEM) confirmed that each bacterium produced polysaccharides with different shapes. In addition, the molecular weight and sugar composition of the polysaccharides produced by each bacterium were distinct. The selected microorganisms include closely related species known to promote plant growth and known to suppress postharvest pathogens. Since these microorganisms may be used not only in known fields but also in new fields, the results of this research are expected to greatly expand the uses of hemicellulose.

著者

Kiyohiko Igarashi Satoshi Kaneko Motomitsu Kitaoka Masahiro Samejima

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

vol.67, no.2, pp.51-57, 2020-05-20 (Released:2020-05-20)

参考文献数

40

被引用文献数

2

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Cellobiose dehydrogenase (CDH) is a flavocytochrome catalyzing oxidation of the reducing end of cellobiose and cellooligosaccharides, and has a key role in the degradation of cellulosic biomass by filamentous fungi. Here, we use a lineup of glucose/xylose-mixed β-1,4-linked disaccharides and trisaccharides, enzymatically synthesized by means of the reverse reaction of cellobiose phosphorylase and cellodextrin phosphorylase, to investigate the substrate recognition of CDH. We found that CDH utilizes β-D-xylopyranosyl-(1→4)-D-glucopyranose (Xyl-Glc) as an electron donor with similar Km and kcat values to cellobiose. β-D-Glucopyranosyl-(1→4)-D-xylopyranose (Glc-Xyl) shows a higher Km value, while xylobiose does not serve as a substrate. Trisaccharides show similar behavior; i.e., trisaccharides with cellobiose and Xyl-Glc units at the reducing end show similar kinetics, while the enzyme was less active towards those with Glc-Xyl, and inactive towards those with xylobiose. We also use docking simulation to evaluate substrate recognition of the disaccharides, and we discuss possible molecular mechanisms of substrate recognition by CDH.

著者

Sosyu Tsutsui Kiyoshi Sakuragi Kiyohiko Igarashi Masahiro Samejima Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

vol.67, no.1, pp.17-22, 2020-02-20 (Released:2020-02-20)

参考文献数

29

被引用文献数

4

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Sugarcane bagasse is a useful biomass resource. In the present study, we examined the efficacy of ammonia pretreatment for selective release of hemicellulose from bagasse. Pretreatment of bagasse with aqueous ammonia resulted in significant loss of xylan. In contrast, pretreatment of bagasse with anhydrous ammonia resulted in almost no xylan loss. Aqueous ammonia or anhydrous ammonia-pretreated bagasse was then subjected to enzymatic digestion with a xylanase from the glycoside hydrolase (GH) family 10 or a xylanase from the GH family 11. The hydrolysis rate of xylan in bagasse pretreated with aqueous ammonia was approximately 50 %. In contrast, in the anhydrous ammonia-treated bagasse, xylan hydrolysis was > 80 %. These results suggested that anhydrous ammonia pretreatment would be an effective method for preparation of sugarcane bagasse for enzymatic hydrolysis to recover xylooligosaccharides.

著者

Haruka Yagi Ryo Takehara Aika Tamaki Koji Teramoto Sosyu Tsutsui Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

pp.jag.JAG-2018_0008, (Released:2018-11-05)

被引用文献数

14

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We functionally characterized the GH10 xylanase (SoXyn10A) and the GH11 xylanase (SoXyn11B) derived from the actinomycete Streptomyces olivaceoviridis E-86. Each enzyme exhibited differences in the produced reducing power upon degradation of xylan substrates. SoXyn10A produced higher reducing power than SoXyn11B. Gel filtration of the hydrolysates generated by both enzymes revealed that the original substrate was completely decomposed. Enzyme mixtures of SoXyn10A and SoXyn11B produced the same level of reducing power as SoXyn10A alone. These observations were in good agreement with the composition of the hydrolysis products. The hydrolysis products derived from the incubation of soluble birchwood xylan with a mixture of SoXyn10A and SoXyn11B produced the same products as SoXyn10A alone with similar compositions. Furthermore, the addition of SoXyn10A following SoXyn11B-mediated digestion of xylan produced the same products as SoXyn10A alone with similar compositions. Thus, it was hypothesized that SoXyn10A could degrade xylans to a smaller size than SoXyn11B. In contrast to the soluble xylans as the substrate, the produced reducing power generated by both enzymes was not significantly different when pretreated milled bagasses were used as substrates. Quantification of the pentose content in the milled bagasse residues after the enzyme digestions revealed that SoXyn11B hydrolyzed xylans in pretreated milled bagasses much more efficiently than SoXyn10A. These data suggested that the GH10 xylanases can degrade soluble xylans smaller than the GH11 xylanases. However, the GH11 xylanases may be more efficient at catalyzing xylan degradation in natural environments (e.g. biomass) where xylans interact with celluloses and lignins.

著者

Hitomi Ichinose Kentaro Suzuki Mari Michikawa Haruna Sato Masahiro Yuki Kei Kamino Wataru Ogasawara Shinya Fushinobu Satoshi Kaneko

出版者

The Japanese Society of Applied Glycoscience

雑誌

Journal of Applied Glycoscience (ISSN:13447882)

巻号頁・発行日

pp.jag.JAG-2017_018, (Released:2017-12-29)

被引用文献数

9

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Highly thermostable β-mannanase, belonging to glycoside hydrolase family 5 subfamily 7, was purified from the culture supernatant of Talaromyces trachyspermus B168 and the cDNA of its transcript was cloned. The recombinant enzyme showed maximal activity at pH 4.5 and 85 °C. It retained more than 90 % of its activity below 60 °C. Obtaining the crystal structure of the enzyme helped us to understand the mechanism of its thermostability. An antiparallel β-sheet, salt-bridges, hydrophobic packing, proline residues in the loops, and loop shortening are considered to be related to the thermostability of the enzyme. The enzyme hydrolyzed mannans such as locust bean gum, carob galactomannan, guar gum, konjac glucomannan, and ivory nut mannan. It hydrolyzed 50.7 % of the total mannans from coffee waste, producing mannooligosaccharides. The enzyme has the highest optimum temperature among the known fungal β-mannanases and has potential for use in industrial applications.

著者

Maki Ishiguro Tomonobu Hori Takuya Ishida Makoto Yoshida Koji Takabatake Satoshi Kaneko Kiyohiko Igarashi Masahiro Samejima

出版者

Japanese Society for Plant Biotechnology

雑誌

Plant Biotechnology (ISSN:13424580)

巻号頁・発行日

vol.27, no.3, pp.273-281, 2010-06-25 (Released:2010-07-15)

参考文献数

26

被引用文献数

5 4

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The basidiomycete Flammulina velutipes is one of the most popular edible mushrooms in Japan, and has the ability to grow on cellulosic biomass as a carbon source. In this study, we have isolated two enzymes belonging to glycoside hydrolase (GH) family 7 (FvCel7A and FvCel7B) from the cellulose-grown culture of the fungus, and cloned cDNAs encoding these enzymes by utilizing a transcriptomic database of this fungus. Although both enzymes contain a catalytic domain belonging to GH family 7, only FvCel7A has the family 1 carbohydrate-binding module at the C-terminal. Sequence comparison indicated that FvCel7A and FvCel7B have a similar pattern of disulfide bonds and similar active site architecture to other fungal GH family 7 enzymes, but show small differences at loop regions covering the active site, which may affect the reactivity of cellulosic substrates.