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Development of novel protein production system using peptide-tag derived from Bacillus thuringiensis Cry toxin.

Research Project

Project/Area Number 23658048
Research Category

Grant-in-Aid for Challenging Exploratory Research

Allocation TypeMulti-year Fund
Research Field Applied entomology
Research InstitutionOkayama University

Principal Investigator

HAYAKAWA Tohru  岡山大学, 自然科学研究科, 助教 (30313555)

Project Period (FY) 2011 – 2013
Project Status Completed (Fiscal Year 2013)
Budget Amount *help
¥3,380,000 (Direct Cost: ¥2,600,000、Indirect Cost: ¥780,000)
Fiscal Year 2013: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2012: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2011: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
KeywordsBacillus thuringiensis / Cry4Aa / 大腸菌 / ペプチドタグ4AaCter / シスタチンC / サソリトキシン / タンパク質生産システム / ペプチドタグ / 4AaCter / サソリ毒 / タンパク質生産 / 凝集体形成 / フェリチン / 発現ベクター / Cryトキシン
Research Abstract

4AaCter and 4AaB7 are peptide-tags derived from mosquitocidal Cry4Aa toxin. These peptide-tags facilitate inclusion formation of fused protein in Escherichia coli. In this study, the ability of 4AaCter and 4AaB7 to from protein inclusion were analyzed. Mutational analyses revealed that the hydrophobic amino acids in 4AaB7 significantly affected the formation of protein inclusion, suggesting some important roles of these hydrophobic amino acid residues. Application of 4AaCter-tag to the production of cystatin C as an endogenous marker of renal dysfunction yielded excellent results, including a dramatic increase in the production level, simplification of the product purification and high immunogenicity. Similarly, scorpion toxins were produced well as alkali-soluble protein inclusion in E. coli, and shown to be a good supplements for Cry toxin-based bioinsecticides.

Report

(4 results)
  • 2013 Annual Research Report   Final Research Report ( PDF )
  • 2012 Research-status Report
  • 2011 Research-status Report
  • Research Products

    (34 results)

All 2014 2013 2012 2011 Other

All Journal Article (5 results) (of which Peer Reviewed: 5 results) Presentation (27 results) Book (2 results)

  • [Journal Article] Potency of insect-specific scorpion toxins on mosquito control using Bacillus thur ingiensis Cry4Aa.2014

    • Author(s)
      Matsumoto R., Shimizu Y., Howlader M. T. H., Namba M., Iwamoto A., Sakai H., Hayakawa T.
    • Journal Title

      Journal of Bioscience and Bioengineering

      Volume: 117 Issue: 6 Pages: 1-4

    • DOI

      10.1016/j.jbiosc.2013.12.004

    • Related Report
      2013 Final Research Report
    • Peer Reviewed
  • [Journal Article] Efficient production of recombinant cystatin C using a peptide-tag, 4AaCter, that facilitates formation of insoluble protein inclusion bodies in Escherichia coli.2013

    • Author(s)
      Hayashi M, Iwamoto S, Sato S, Sudo S, Takagi M, Sakai H, Hayakawa T.
    • Journal Title

      Protein Expression and Purification

      Volume: 88 Issue: 2 Pages: 230-234

    • DOI

      10.1016/j.pep.2013.01.011

    • Related Report
      2013 Annual Research Report 2013 Final Research Report 2012 Research-status Report
    • Peer Reviewed
  • [Journal Article] Parasporin 1Ac2, a novel cytotoxic crystal protein isolated from Bacillus2013

    • Author(s)
      Kuroda S, Begum A, Saga M, Hirao A, Mizuki E, Sakai H, Hayakawa T.
    • Journal Title

      Current microbiology

      Volume: 66 Issue: 5 Pages: 475-480

    • DOI

      10.1007/s00284-013-0301-1

    • Related Report
      2013 Annual Research Report 2013 Final Research Report 2012 Research-status Report
    • Peer Reviewed
  • [Journal Article] Mutational analyses of Cry protein block 7 polypeptides that facilitate the formation of protein inclusion in Escherichia coli2011

    • Author(s)
      Hayakawa T., Shimizu Y., Ishida T., Sakai H
    • Journal Title

      Applied Microbiology and Biotechnology

      Volume: 90 Issue: 6 Pages: 1943-1951

    • DOI

      10.1007/s00253-011-3216-4

    • Related Report
      2013 Final Research Report
    • Peer Reviewed
  • [Journal Article] Mutational analyses of Cry protein block 7 polypeptides that facilitate the formation of protein inclusion in Escherichia coli.2011

    • Author(s)
      Hayakawa T., Shimizu Y., Ishida T., Sakai H.
    • Journal Title

      Applied Microbiology and Biotechnology

      Volume: 90 Pages: 1943-1951

    • Related Report
      2011 Research-status Report
    • Peer Reviewed
  • [Presentation] Bt 菌が産生する殺蚊トキシンCry4Aa の複雑な作用機構について2014

    • Author(s)
      早川徹, Howlader M.T.H, 中尾早織, 井出徹
    • Organizer
      日本蚕糸学会第84回大会
    • Place of Presentation
      神奈川県藤沢
    • Related Report
      2013 Final Research Report
  • [Presentation] Bt菌が産生する殺蚊トキシンCry4Aaの複雑な作用機構について2014

    • Author(s)
      早川徹、Howlader M.T.H.、中尾早織、井出徹
    • Organizer
      平成26年度蚕糸・昆虫機能利用学術講演会
    • Place of Presentation
      神奈川県藤沢
    • Related Report
      2013 Annual Research Report
  • [Presentation] 大腸菌を用いて殺蚊トキシンCry11Aa を効率的に生産する方法2013

    • Author(s)
      早川徹, 佐野乙香, 井出徹
    • Organizer
      第36回分子生物学会年会
    • Place of Presentation
      神戸
    • Related Report
      2013 Final Research Report
  • [Presentation] Interactions between mosquitocidal Cry4Aa and the brush border membrane proteins of Culex pipiens larvae2013

    • Author(s)
      Howlader M.T.H., Nakao S., Sakai H., Hayakawa T
    • Organizer
      46th Annual Meeting of the Society for Invertebrate Pathology
    • Place of Presentation
      Pittsburgh, U.S.A
    • Related Report
      2013 Final Research Report
  • [Presentation] Novel protein production system using a peptide-tag derived from Bacillus thuringiensis mosquitocidal Cry4Aa toxin2013

    • Author(s)
      Hayakawa T., Sato S., Iwamoto S., Sudo S., Howlader M.T.H., Sakai H
    • Organizer
      46th Annual Meeting of the Society for Invertebrate Pathology
    • Place of Presentation
      Pittsburgh, U.S.A
    • Related Report
      2013 Final Research Report
  • [Presentation] Functional structures of Cry4Aa toxin that are responsible for the mosquitocidal activity against Culex pipiens2013

    • Author(s)
      Howlader M.T.H, , Nakao S., Sakai H., Hayakawa T
    • Organizer
      CARES2013 International Conference on Biotechnology
    • Place of Presentation
      Dhaka, Bangladesh
    • Related Report
      2013 Final Research Report
  • [Presentation] 殺蚊トキシンCry4Aa とサソリトキシン(BjαIT 及びAahIT)を利用する新規害虫防除法の可能性2013

    • Author(s)
      早川徹, 松本陸, 清水佳孝, 酒井裕
    • Organizer
      日本蚕糸学会第83回大会
    • Place of Presentation
      筑波
    • Related Report
      2013 Final Research Report
  • [Presentation] Functional structures of Cry4Aa toxin that are responsible for the mosquitocidal activity against Culex pipiens.2013

    • Author(s)
      Howlader¬ M.T.H., Nakao S, Sakai H., Hayakawa T.
    • Organizer
      CARES2013 International Conference on Biotechnology
    • Place of Presentation
      Dhaka, Bangladesh
    • Related Report
      2013 Annual Research Report
  • [Presentation] Novel protein production system using a peptide-tag derived from Bacillus thuringiensis mosquitocidal Cry4Aa toxin.2013

    • Author(s)
      Hayakawa T., Sato S., Iwamoto S., Sudo S., Howlader M.T.H., Sakai H
    • Organizer
      46th Annual Meeting of the Society for Invertebrate
    • Place of Presentation
      Pittsburgh, U.S.A.
    • Related Report
      2013 Annual Research Report
  • [Presentation] Interactions between mosquitocidal Cry4Aa and the brush border membrane proteins of Culex pipiens larvae.2013

    • Author(s)
      Howlader M.T.H., Nakao S., Sakai H., Hayakawa T.
    • Organizer
      46th Annual Meeting of the Society for Invertebrate
    • Place of Presentation
      Pittsburgh, U.S.A.
    • Related Report
      2013 Annual Research Report
  • [Presentation] 大腸菌を用いて殺蚊トキシンCry11Aaを効率的に生産する方法2013

    • Author(s)
      早川徹、佐野乙香、井出徹
    • Organizer
      第36回分子生物学会年会
    • Place of Presentation
      兵庫県神戸
    • Related Report
      2013 Annual Research Report
  • [Presentation] 殺蚊トキシンCry4Aa に由来するペプチドタグの開発と検査薬成分(抗原タンパク質)の効率的生産2012

    • Author(s)
      早川徹, 林雅浩, 高木麻里, 佐藤真也, 岩本繁久, 須藤薫雄, 酒井裕
    • Organizer
      日本蚕糸学会第82回大会
    • Place of Presentation
      福岡
    • Year and Date
      2012-03-18
    • Related Report
      2013 Final Research Report
  • [Presentation] 殺蚊トキシンCry11A の受容体結合部位と考えられるドメインII ループ構造への変異導入2012

    • Author(s)
      金治正泰, 駒田卓也, Howlader M.T.H, 酒井裕, 早川徹
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2013 Final Research Report
  • [Presentation] 不溶性凝集体形成を促す新規ペプチドタグ(4AaCter)を利用したシスタチンC の効率的生産と精製2012

    • Author(s)
      林雅浩, 高木麻里, 佐藤真也, 岩本繁久, 須藤薫雄, 酒井裕, 早川徹
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2013 Final Research Report
  • [Presentation] Detection of domainII loops that are responsible for mosquitocidal activity of Cry4Aa from Bacillus thuringiensis2012

    • Author(s)
      Howlader M.T.H., Nakao S., Sakai H., Hayakawa T
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2013 Final Research Report
  • [Presentation] 殺蚊トキシンCry4Aaに由来するペプチドタグの開発と検査薬成分(抗原タンパク質)の効率的生産2012

    • Author(s)
      早川徹、林雅浩、高木麻里、佐藤真也、岩本繁久、須藤薫雄、酒井裕
    • Organizer
      平成24年度 蚕糸・昆虫機能利用学術講演会-日本蚕糸学会第82回大会
    • Place of Presentation
      福岡
    • Related Report
      2011 Research-status Report
  • [Presentation] BT 菌に由来するポリペプチドタグを利用した検査薬成分(抗原タンパク質)の効率的生産2011

    • Author(s)
      林雅浩, 高木麻里, 山崎貴郁, 山西勇輝, 佐藤真也, 岩本繁久, 須藤薫雄, 早川徹, 酒井裕
    • Organizer
      第34回日本分子生物学会年会
    • Place of Presentation
      横浜
    • Year and Date
      2011-12-13
    • Related Report
      2013 Final Research Report
  • [Presentation] BT 菌由来の新規なペプチドタグは効率的なタンパク質生産を可能にする2011

    • Author(s)
      酒井裕, 林雅浩, 佐藤真也, 岩本繁久, 須藤薫雄, 早川徹
    • Organizer
      第34回日本分子生物学会年会
    • Place of Presentation
      横浜
    • Year and Date
      2011-12-13
    • Related Report
      2013 Final Research Report
  • [Presentation] 昆虫特異的Cry トキシンの新しい利用法-タンパク質を凝集体として発現・蓄積させる新規なペプチドタグの開発-2011

    • Author(s)
      早川徹, 佐藤真也, 岩本繁久, 須藤薫雄, 酒井裕
    • Organizer
      第84回日本生化学会大会
    • Place of Presentation
      京都
    • Year and Date
      2011-09-22
    • Related Report
      2013 Final Research Report
  • [Presentation] 昆虫特異的Cryトキシンの新しい利用法-タンパク質を凝集体として発現・蓄積させる新規なペプチドタグの開発-2011

    • Author(s)
      早川徹、佐藤真也、岩本繁久、須藤薫雄、酒井裕
    • Organizer
      第84回日本生化学会大会(招待講演)
    • Place of Presentation
      京都
    • Related Report
      2011 Research-status Report
  • [Presentation] BT菌由来の新規なペプチドタグは効率的なタンパク質生産を可能にする2011

    • Author(s)
      酒井裕、林雅浩、佐藤真也、岩本繁久、須藤薫雄、早川徹
    • Organizer
      第34回 日本分子生物学会年会
    • Place of Presentation
      横浜
    • Related Report
      2011 Research-status Report
  • [Presentation] BT菌に由来するポリペプチドタグを利用した検査薬成分(抗原タンパク質)の効率的生産2011

    • Author(s)
      林雅浩、高木麻里、山崎貴郁、山西勇輝、佐藤真也、岩本繁久、須藤薫雄、早川徹、酒井裕
    • Organizer
      第34回 日本分子生物学会年会
    • Place of Presentation
      横浜
    • Related Report
      2011 Research-status Report
  • [Presentation] Bt菌B0462株に由来する新規パラスポリン(PS1Ac2)の解析

    • Author(s)
      早川徹、黒田将太、Begum A、佐賀瑞恵、平尾彬奈、水城英一、酒井裕
    • Organizer
      第5回パラスポリン研究会
    • Place of Presentation
      熊本
    • Related Report
      2012 Research-status Report
  • [Presentation] Detection of domainII loops that are responsible for mosquitocidal activity of

    • Author(s)
      Howlader M. T. H., Nakao S., Sakai H., Hayakawa T.
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2012 Research-status Report
  • [Presentation] 不溶性凝集体形成を促す新規ペプチドタグ(4AaCter)を利用したシスタチンCの効率的生産と精製

    • Author(s)
      林雅浩、高木麻里、佐藤真也、岩本繁久、須藤薫雄、酒井裕、早川徹
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2012 Research-status Report
  • [Presentation] 殺蚊トキシンCry11Aの受容体結合部位と考えられるドメインIIループ構造への変異導入

    • Author(s)
      金治正泰、駒田卓也、モハッマド TH ハウラダル、酒井裕、早川徹
    • Organizer
      第35回日本分子生物学会年会
    • Place of Presentation
      福岡
    • Related Report
      2012 Research-status Report
  • [Presentation] 殺蚊トキシンCry4Aaとサソリトキシン(BjαIT及びAahIT)を利用する新規害虫防除法の可能性

    • Author(s)
      早川徹、松本陸、清水佳孝、酒井裕
    • Organizer
      日本蚕糸学会第83回大会
    • Place of Presentation
      筑波
    • Related Report
      2012 Research-status Report
  • [Book] 最新昆虫病理学2014

    • Author(s)
      早川徹, 酒井裕
    • Publisher
      講談社
    • Related Report
      2013 Final Research Report
  • [Book] 最新昆虫病理学、第10章 昆虫病理学の展開、4Bacillus thuringiensisの開発と利用2014

    • Author(s)
      早川徹、酒井裕
    • Total Pages
      251
    • Publisher
      講談社
    • Related Report
      2013 Annual Research Report

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Published: 2011-08-05   Modified: 2019-07-29  

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