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Elucidation of the catalytic mechanism of bacterial cell-surface-expressed chitinase ChiW and chitin oligosaccharide production by using the high transglycosylation activity of ChiW.

Research Project

Project/Area Number 25850234
Research Category

Grant-in-Aid for Young Scientists (B)

Allocation TypeMulti-year Fund
Research Field Environmental agriculture(including landscape science)
Research InstitutionFukui Prefectural University

Principal Investigator

Itoh Takafumi  福井県立大学, 生物資源学部, 准教授 (10402827)

Research Collaborator HIBI Takao  福井県立大学, 生物資源学部, 教授 (00285181)
KIMOTO Hisashi  福井県立大学, 生物資源学部, 教授 (70283166)
FUJII Yutaka  福井大学, 医学部, 教授 (80211522)
Project Period (FY) 2013-04-01 – 2016-03-31
Project Status Completed (Fiscal Year 2015)
Budget Amount *help
¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000)
Fiscal Year 2015: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2014: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2013: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
KeywordsChitinase / Paenibacillus / Structural biology / chitin oligosaccharides / chitinase / structural biology / chitin / oligosaccharides
Outline of Final Research Achievements

The purposes of this study are to elucidate the degradation mechanism of the novel multi-modular surface-expressed chitinase, ChiW, of Paenibacillus sp. str. FPU-7 and to enhance the transglycosylation activity of the enzyme.
(1) The degradation mechanism of ChiW. The crystal structure of ChiW was determined at 2.0 angstrom resolution. The structural and functional analyses reveal that the enzyme is composed of six structural domains for efficient chitin degradation. (2) Enhancement of the transglycosylation activity of ChiW. The important residues for substrate binding were selected as the mutation sites. The mutant enzymes were prepared by site-specific mutagenesis. As the result, two mutant enzymes with higher transglycosylation activities (120-130%) were obtained. (3) Identification of P. FPU-7 proteins for chitin degradation. Proteomic analysis was carried on P. FPU-7 extracellular proteins. A number of chitinase and chitin-related proteins were identified thorough the analysis.

Report

(4 results)
  • 2015 Annual Research Report   Final Research Report ( PDF )
  • 2014 Research-status Report
  • 2013 Research-status Report
  • Research Products

    (17 results)

All 2015 2014 2013 Other

All Journal Article (6 results) (of which Peer Reviewed: 6 results,  Open Access: 2 results,  Acknowledgement Compliant: 2 results) Presentation (8 results) (of which Invited: 4 results) Remarks (3 results)

  • [Journal Article] Colorimetric Determination of Fructose for the High-Throughput Microtiter Plate Assay of Glucose Isomerase2015

    • Author(s)
      H. Katano, M. Takakuwa, T. Itoh, T. Hibi
    • Journal Title

      Bioscience, Biotechnology, and Biochemistry.

      Volume: 79 Issue: 7 Pages: 1057-1060

    • DOI

      10.1080/09168451.2015.1010477

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Open Access / Acknowledgement Compliant
  • [Journal Article] Microplate Assay of α-Glucosidase and Its Inhibitors Based on the Direct Reduction of Molybdosilicate by Glucose2015

    • Author(s)
      H. Katano, M. Takakuwa, T. Itoh, T. Hibi
    • Journal Title

      Analytical Sciences

      Volume: 31 Issue: 12 Pages: 1291-1295

    • DOI

      10.2116/analsci.31.1291

    • NAID

      130005129734

    • ISSN
      0910-6340, 1348-2246
    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Open Access / Acknowledgement Compliant
  • [Journal Article] Paenibacillus属細菌のキチン分解機構2014

    • Author(s)
      木元 久, 伊藤 貴文, 日び 隆雄, 藤井 豊, 草桶 秀雄
    • Journal Title

      応用糖質化学

      Volume: 4 Pages: 113-120

    • Related Report
      2014 Research-status Report
    • Peer Reviewed
  • [Journal Article] Crystallization and preliminary X-ray analysis of the catalytic domains of Paenibacillus sp. strain FPU-7 cell-surface-expressed chitinase ChiW.2014

    • Author(s)
      Itoh, T., Hibi, T., Sugimoto, I., Suzuki, F., Fujii, Y., Taketo, A. and Kimoto, H.
    • Journal Title

      Acta Crystallogr. F Struct. Biol. Commun.

      Volume: 70 Issue: 3 Pages: 350-353

    • DOI

      10.1107/s2053230x14002325

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Overexpression, purification, and characterization of Paenibacillus cell surface-expressed chitinase ChiW with two catalytic domains2014

    • Author(s)
      T. Itoh, I. Sugimoto, T. Hibi, F. Suzuki, K. Matsuo, Y. Fujii, A. Taketo, H. Kimoto
    • Journal Title

      Biosci. Biotech. Biochem.

      Volume: 78 Issue: 4 Pages: 624-634

    • DOI

      10.1080/09168451.2014.891935

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Cooperative degradation of chitin by extracellular and cell surface-expressed chitinases from Paenibacillus sp. strain FPU-7.2013

    • Author(s)
      Itoh, T., Hibi, T., Fujii, Y., Sugimoto, I., Fujiwara, A., Suzuki, F., Iwasaki, Y., Kim, J-K., Taketo, A. and Kimoto, H.
    • Journal Title

      Appl. Environ. Microbiol.

      Volume: 79 Issue: 23 Pages: 7482-7490

    • DOI

      10.1128/aem.02483-13

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Presentation] 固い細胞壁を細菌はどのようにして破壊するのか-細胞表層結合型キチナーゼの立体構造解析-2015

    • Author(s)
      伊藤 貴文, 日び 隆雄, 木元 久
    • Organizer
      第8回北陸合同バイオシンポジウム2015
    • Place of Presentation
      山中節と温泉の館 山中座 (石川県・加賀市)
    • Year and Date
      2015-10-31
    • Related Report
      2015 Annual Research Report
    • Invited
  • [Presentation] 立体構造に基づく糖質関連酵素の反応機構の解明とポストゲノミクスへの新展開2015

    • Author(s)
      伊藤 貴文
    • Organizer
      日本農芸化学会、中部支部第173回例会
    • Place of Presentation
      名古屋大学農学部 (愛知県・名古屋市)
    • Year and Date
      2015-06-20
    • Related Report
      2015 Annual Research Report
    • Invited
  • [Presentation] Paenibacillus属細菌由来細胞表層キチナーゼChiWの組換えタンパク質発現と細胞表層結合ドメインの応用2015

    • Author(s)
      伊藤 貴文, 日ビ 隆雄, 横内 佳奈, 高宮 杏奈, 藤井 豊, 武藤 明, 木元 久
    • Organizer
      日本農芸化学会2015年度大会
    • Place of Presentation
      岡山大学、岡山市
    • Year and Date
      2015-03-27
    • Related Report
      2014 Research-status Report
  • [Presentation] 立体構造に基づく糖質関連酵素の反応機構の解明とポストゲノミクスへの新展開2015

    • Author(s)
      伊藤 貴文
    • Organizer
      日本農芸化学会2015年度大会 奨励賞受賞講演
    • Place of Presentation
      ホテルグランヴィア岡山、岡山市
    • Year and Date
      2015-03-26
    • Related Report
      2014 Research-status Report
    • Invited
  • [Presentation] “越前がに”のブランドイメージを活用した次世代型農業資材2014

    • Author(s)
      木元 久, 笠原 康一, 加藤 久晴, 伊藤 貴文, 日び 隆雄
    • Organizer
      2014北陸合同バイオシンポジウム
    • Place of Presentation
      八尾ゆめの森 ゆうゆう館、富山市
    • Year and Date
      2014-11-28 – 2014-11-29
    • Related Report
      2014 Research-status Report
  • [Presentation] Paenibacillus属細菌由来キチン分解酵素ChiWのN末端領域の解析2014

    • Author(s)
      伊藤 貴文, 日ビ 隆雄, 高宮 杏奈, 鈴木 史子, 杉本 郁美, 藤井 豊, 武藤 明, 木元 久
    • Organizer
      日本農芸化学会2014年度大会
    • Place of Presentation
      川崎市, 明治大学
    • Related Report
      2013 Research-status Report
  • [Presentation] Chitinases secreted by Paenibacillus sp. strain IK-5 form a huge complex.2013

    • Author(s)
      Kimoto, H., Itoh, T., Hibi, T., Fujiwara, A., Fujii, Y., Tanaka, Y., Suzuki, F., Taketo, A., Kusaoke, H.
    • Organizer
      10th Asia-Pacific Chitin & Chitosan Symposium
    • Place of Presentation
      米子市, 米子コンベンションセンター BIG SHIP
    • Related Report
      2013 Research-status Report
    • Invited
  • [Presentation] Paenibacillus str. FPU-7由来キチナーゼ ChiWのN末端領域の機能解析2013

    • Author(s)
      高宮 杏奈, 伊藤 貴文, 日ビ 隆雄, 大門 結花, 髙木 良樹, 仲上 真由, 鈴木 史子, 杉本 郁美, 藤井 豊, 武藤 明, 木元 久
    • Organizer
      2013北陸合同バイオシンポジウム
    • Place of Presentation
      七尾市, 国民宿舎能登小牧台 コンベンションホール「万葉」
    • Related Report
      2013 Research-status Report
  • [Remarks] 福井県立大学 教員情報 生物資源学部 伊藤 貴文

    • URL

      http://fpuinfo.fpu.ac.jp/fpu/system/php/faculties_open/showone.php?id=ito-t

    • Related Report
      2015 Annual Research Report
  • [Remarks] 応用生化学領域ホームページ

    • URL

      http://biotech.fpu.ac.jp/3f.html

    • Related Report
      2013 Research-status Report
  • [Remarks] 教員情報

    • URL

      http://fpuinfo.fpu.ac.jp/fpu/system/php/faculties_open/showone.php?id=ito-t

    • Related Report
      2013 Research-status Report

URL: 

Published: 2014-07-25   Modified: 2019-07-29  

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