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Use of highly efficient procedure in getting revertant cells, for studies of interaction manner between membrane protein and its inhibitor

Research Project

Project/Area Number 17K08274
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeMulti-year Fund
Section一般
Research Field Biological pharmacy
Research InstitutionThe University of Tokushima

Principal Investigator

SHINOHARA Yasuo  徳島大学, 先端酵素学研究所(プロテオ), 教授 (60226157)

Project Period (FY) 2017-04-01 – 2020-03-31
Project Status Completed (Fiscal Year 2019)
Budget Amount *help
¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000)
Fiscal Year 2019: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2018: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
KeywordsADP/ATP輸送体 / ボンクレキン酸 / 分子間相互作用 / 復帰変異株 / タンパク質リガンド相互作用 / ミトコンドリア
Outline of Final Research Achievements

We tried to isolate the revertant yeast cells viable on the glycerol plate containing bongkrekic acid, by screening the library of yeast cells expressing randomly mutated mitochondrial ADP/ATP carrier. As a result, we succeeded in identifying two known mutations of L142S and G298S, and three unknown mutations of I200V, S245P and V300I. Furthermore, these identified amino acids were shown to locate closely to bongkrekic acid, by independently achieved crystallographic analysis of the ADP/ATP carrier complexed with bongkrekic acid.

Academic Significance and Societal Importance of the Research Achievements

膜タンパク質とリガンドの相互作用の様式の理解は、創薬を考える上で極めて大きな課題であるが、これまでは光親和性標識かタンパク質のX線結晶構造解析しか実験法が確立されておらず、新たな研究手法の開発が急務であった。
今回の研究によって、我々が見出した高効率な復帰変異株獲得による膜蛋白質とリガンドの相互作用解析法が実用的なものであることが示され、新たな研究手法になり得るものと期待される。

Report

(4 results)
  • 2019 Annual Research Report   Final Research Report ( PDF )
  • 2018 Research-status Report
  • 2017 Research-status Report
  • Research Products

    (15 results)

All 2019 2018 2017

All Journal Article (6 results) (of which Int'l Joint Research: 1 results,  Peer Reviewed: 6 results,  Open Access: 3 results) Presentation (9 results) (of which Int'l Joint Research: 3 results,  Invited: 2 results)

  • [Journal Article] Pentenediol-type compounds specifically bind to voltage-dependent anion channel 1 in Saccharomyces cerevisiae mitochondria2019

    • Author(s)
      Yufu Unten, Masatoshi Murai, Takenori Yamamoto, Akira Watanabe, Naoya Ichimaru, Shunsuke Aburaya, Wataru Aoki, Yasuo Shinohara and Hideto Miyoshi
    • Journal Title

      Biochemistry

      Volume: 58 Issue: 8 Pages: 1141-1154

    • DOI

      10.1021/acs.biochem.8b01209

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Effects of cold exposure on metabolites in brown adipose tissue of rats2018

    • Author(s)
      Hiroshima Y, Yamamoto T, Watanabe M, Baba Y, Shinohara Y.
    • Journal Title

      Mol Genet Metab Rep

      Volume: 15 Pages: 36-42

    • DOI

      10.1016/j.ymgmr.2018.01.005

    • NAID

      120006980537

    • Related Report
      2018 Research-status Report 2017 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Use of modified U1 small nuclear RNA for rescue from exon 7 skipping caused by 5'-splice site mutation of human cathepsin A gene2018

    • Author(s)
      Yamazaki N, Kanazawa K, Kimura M, Ike H, Shinomiya M, Tanaka S, Shinohara Y, Minakawa N, Itoh K, Takiguchi Y.
    • Journal Title

      Gene

      Volume: 677 Pages: 41-48

    • DOI

      10.1016/j.gene.2018.07.030

    • NAID

      120006706063

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Polyethyleneimine renders mitochondrial membranes permeable by interacting with negatively charged phospholipids in them2018

    • Author(s)
      Yamamoto T, Tsunoda M, Ozono M, Watanabe A, Kotake K, Hiroshima Y, Yamada A, Terada H, Shinohara Y.
    • Journal Title

      Arch Biochem Biophys

      Volume: 652 Pages: 9-17

    • DOI

      10.1016/j.abb.2018.06.003

    • Related Report
      2018 Research-status Report
    • Peer Reviewed
  • [Journal Article] Synthesis and evaluation of simplified functionalized bongkrekic acid analogs2018

    • Author(s)
      Satoshi Fujita, Masaki Suyama, Kenji Matsumoto, Atsushi Yamamoto, Takenori Yamamoto, Yuka Hiroshima, Takayuki Iwata, Arihiro Kano, Yasuo Shinohara, Mitsuru Shindo
    • Journal Title

      Tetrahedron

      Volume: 74 Issue: 9 Pages: 962-969

    • DOI

      10.1016/j.tet.2018.01.018

    • Related Report
      2018 Research-status Report 2017 Research-status Report
    • Peer Reviewed
  • [Journal Article] Advanced glycation end-products and Porphyromonas gingivalis lipopolysaccharide increase calprotectin expression in human gingival epithelial cells.2017

    • Author(s)
      Yuka Hiroshima, Eijiro Sakamoto, Kaya Yoshida, Kaori Abe, Koji Naruishi, Takenori Yamamoto, Yasuo Shinohara, Jun-ichi Kido and Carolyn L Geczy
    • Journal Title

      Journal of Cellular Biochemistry

      Volume: 119 Issue: 2 Pages: 1591-1603

    • DOI

      10.1002/jcb.26319

    • Related Report
      2018 Research-status Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Presentation] ADP/ATP輸送体のボンクレキン酸と相互作用するアミノ酸残基の変異の掛け合わせがヌクレオチド輸送活性に及ぼす影響の解析2019

    • Author(s)
      問山温未、大園瑞音、大和永奈、山越亮平、井戸佑介、山本武範、寺田 弘、篠原康雄
    • Organizer
      第58回 日本薬学会・日本薬剤師会・日本病院薬剤師会 中国四国支部学術大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] Ru360がミトコンドリアのカルシウム取込みを阻害する分子機構の解析2019

    • Author(s)
      前田康輔、山本武範、渡辺 朗、大園瑞音、奈良 篤、橋田芽依、井戸佑介、寺田 弘、篠原康雄
    • Organizer
      第58回 日本薬学会・日本薬剤師会・日本病院薬剤師会 中国四国支部学術大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] マグミット錠の投与がマウスの遺伝子発現に及ぼす影響の解析2019

    • Author(s)
      長尾和輝、山本武範、小泉 諒、井戸佑介、篠原康雄
    • Organizer
      第58回 日本薬学会・日本薬剤師会・日本病院薬剤師会 中国四国支部学術大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] マグネシウム塩を経口投与したマウスの血清マグネシウム濃度の解析2019

    • Author(s)
      小泉 諒、山本武範、長尾和輝、井戸佑介、篠原康雄
    • Organizer
      第58回 日本薬学会・日本薬剤師会・日本病院薬剤師会 中国四国支部学術大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] ミトコンドリアカルシウムイオンチャネルのコアサブユニットの発現量比の決定2019

    • Author(s)
      渡辺 朗、山本武範、前田康輔、井戸佑介、山田安希子、寺田 弘、篠原康雄
    • Organizer
      第58回 日本薬学会・日本薬剤師会・日本病院薬剤師会 中国四国支部学術大会
    • Related Report
      2019 Annual Research Report
  • [Presentation] Two structural features of the 3’ region of the carnitine palmitoyltransferase 1a (CPT1a) gene specifically observed with human but not with rodents2018

    • Author(s)
      Yamazaki N, Ohya T, Kawaguchi Y, Hiroshima Y, Yamamoto T, Shinohara Y, Takiguchi Y.
    • Organizer
      The 43rd FEBS Congress
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research
  • [Presentation] Use of modified U1 small nuclear RNA for improved formation of properly spliced mRNA encoding human cathepsin A from the gene having an IVS7 +3a>g mutation2018

    • Author(s)
      Yamazaki N, Shinomiya M, Ike H, Shinohara Y, Minakawa N, Itoh H, Takiguchi Y.
    • Organizer
      The 43rd FEBS Congress
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research
  • [Presentation] 酵母を用いたタンパク質発現系の魅力 ~ミトコンドリアのタンパク質研究と創薬研究に おける有用性~2017

    • Author(s)
      篠原康雄
    • Organizer
      第15回東京理科大学薬学部DDS研究センターシンポジウム
    • Related Report
      2017 Research-status Report
    • Invited
  • [Presentation] Functional expression of mammalian mitochondrial phosphate carrier in yeast cells.2017

    • Author(s)
      Yasuo Shinohara
    • Organizer
      The First International Conference on Pharmaceutical and Clinical Research
    • Related Report
      2017 Research-status Report
    • Int'l Joint Research / Invited

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Published: 2017-04-28   Modified: 2021-02-19  

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