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ランチビオティックnukacin ISK-1のペプチド工学と膜結合モチーフの解析

Research Project

Project/Area Number 10F00099
Research Category

Grant-in-Aid for JSPS Fellows

Allocation TypeSingle-year Grants
Section外国
Research Field Applied microbiology
Research InstitutionKyushu University

Principal Investigator

園元 謙二  九州大学, 大学院・農学研究院, 教授

Co-Investigator(Kenkyū-buntansha) ISLAM M.R.  九州大学, 大学院・農学研究院, 外国人特別研究員
Project Period (FY) 2010 – 2011
Project Status Completed (Fiscal Year 2011)
Budget Amount *help
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 2011: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2010: ¥1,000,000 (Direct Cost: ¥1,000,000)
Keywordsランチビオティック / nukacin ISK-1 / 異常アミノ酸 / 等温滴定型熱量計 / ランダム変異導入法 / 抗菌ペプチド / ペプチドデザイン / 分子設計 / 翻訳後修飾 / MALDI-TOF/MS / ランチビオティックの分子設計
Research Abstract

Nukacin ISK-1は、抗菌活性や構造安定性に重要である3つのランチオニン環(Ring A,B,C)を持っている。我々はnukacin ISK-1構造遺伝子のランダム変異導入により変異体ライブラリーを構築し、その中で抗菌活性が増強、低下または消失する変異体の取得に成功している(Islam et al.,Mol.Microbiol.,72:1438,2009)。本研究では、nukacin ISK-1中のring A領域に着目し、この環構造が抗菌活性および細胞壁前駆体であるlipid IIへの結合に及ぼす影響を解析することを目的とした。
Nukacin ISK-1 (WT)およびringA変異体(D13E、高活性;D13A、低活性;C14S(ring Aを破壊した変異体)、活性なし)を作用させたS.aureus SG511細胞内の細胞壁前駆体蓄積を解析した結果、WTとD13Eでは顕著にUDP-N-acetylmuramyl pentapeptideが蓄積していた。また、細胞壁前駆体lipid Iおよびlipid IIはnukacin ISK-1の抗菌活性を阻害した。さらに等温滴定型熱量計を用いてnukacin ISK-1やその変異体とlipid IIとの結合を定量的に解析した。WTとD13Eは同程度の親和性を示した(WT、K_D=0.17μM;D13E、K_D=0.19μM)。一方、D13Aは低い親和性を示し(K_D=3-4μM)、C14Sは結合が見られなかった。以上の結果から、nukacin ISK-1はring Aを介してlipid IIと結合し細胞壁合成を阻害することで抗菌活性を示すことが明らかになった。本研究は、nukacin ISK-1を含むランチビオティックのサブタイプにおいて初めてlipid II結合モチーフを見出したものである。

Report

(2 results)
  • 2011 Annual Research Report
  • 2010 Annual Research Report
  • Research Products

    (18 results)

All 2012 2011 2010 Other

All Journal Article (8 results) (of which Peer Reviewed: 8 results) Presentation (6 results) Book (2 results) Remarks (2 results)

  • [Journal Article] Enhanced production of nukacin D13E in Lactococcus lactis NZ9000 by the additional expression of immunity genes2012

    • Author(s)
      Tijo Varghese Puramattathu, et al
    • Journal Title

      Applied Microbiology and Biotechnoly

      Volume: 93 Issue: 2 Pages: 671-678

    • DOI

      10.1007/s00253-011-3563-1

    • Related Report
      2011 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Ring A of nukacin ISK-1 : a lipid II-binding motif for type-A (II) lantibiotic2012

    • Author(s)
      ISLAM M.R., et al
    • Journal Title

      Journal of the American Chemical Society

      Volume: 134 Issue: 8 Pages: 3687-3690

    • DOI

      10.1021/ja300007h

    • Related Report
      2011 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Lantibiotic transporter requires cooperative functioning of the peptidase domain and the ATP binding domain2011

    • Author(s)
      Mami Nishie, et al
    • Journal Title

      Journal of Biological Chemistry

      Volume: 286 Issue: 13 Pages: 11163-11169

    • DOI

      10.1074/jbc.m110.212704

    • Related Report
      2011 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Methodologies and strategies for the bioengineering of lantibiotics2011

    • Author(s)
      Jun-ichi Nagao, et al
    • Journal Title

      Current Pharmaceutical Biotechnology

      Volume: 12 Pages: 1221-1230

    • Related Report
      2011 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Structural and functional diversity of lantibiotic immunity proteins2011

    • Author(s)
      Ken-ichi Okuda, et al
    • Journal Title

      Current Pharmaceutical Biotechnology

      Volume: 12 Pages: 1231-1239

    • Related Report
      2011 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Lantibiotic transporter requires cooperative functioning of the peptidase domain and the ATP binding domain2011

    • Author(s)
      Mami Nishie, et al.
    • Journal Title

      J.Biol.Chem.

      Volume: in press

    • Related Report
      2010 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Characterization of modification enzyme NukM and engineering of a novel thioet her bridge in lantibiotic nukacin ISK-12010

    • Author(s)
      Kouki Shioya, et al.
    • Journal Title

      Appl.Microbiol.Biotechnol.

      Volume: 86 Pages: 891-899

    • Related Report
      2010 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Functional significance of the E-loop, a novel motif conserved in the lantibiotic-immunity ABC transport systems2010

    • Author(s)
      Ken-ichi Okuda, et al.
    • Journal Title

      J.Bacteriol.

      Volume: 192 Pages: 2801-2808

    • Related Report
      2010 Annual Research Report
    • Peer Reviewed
  • [Presentation] Ring A of nukacin ISK-1 : a lipid II binding motif for type-A (II) antibiotic2012

    • Author(s)
      ISLAM M.R.
    • Organizer
      日本農芸化学会2012年度大会
    • Place of Presentation
      京都女子大学(京都)
    • Year and Date
      2012-03-23
    • Related Report
      2011 Annual Research Report
  • [Presentation] Lantibiotic transporter, NukT, requires cooperative functioning of the peptidase domain and the ATP binding domain2012

    • Author(s)
      Kenji Sonomoto
    • Organizer
      ATP-Binding Cassette (ABC) Proteins : From Multidrug Resistance to Genetic Diseases
    • Place of Presentation
      Innsbruck, Austria(招待講演)
    • Year and Date
      2012-03-08
    • Related Report
      2011 Annual Research Report
  • [Presentation] New Era of Lactic Acid Bacteria to Open the Future2012

    • Author(s)
      Kenji Sonomoto
    • Organizer
      The Young Scientist Seminar 2011 (Bioresources : Their Potentials & Applications)
    • Place of Presentation
      Bangkok, Thailand(招待講演)
    • Year and Date
      2012-01-31
    • Related Report
      2011 Annual Research Report
  • [Presentation] New Insight of Lantibiotic Engineering : Characterization of the enzymes, design and mode of action of lantibiotics, and immunity proteins2011

    • Author(s)
      Kenji Sonomoto
    • Organizer
      Enzyme Engineering XXI
    • Place of Presentation
      Vail, CO, USA
    • Year and Date
      2011-09-20
    • Related Report
      2011 Annual Research Report
  • [Presentation] Ring A of nukacin ISK-1 : a lipid II binding motif for type-A (II) antibiotic2011

    • Author(s)
      ISLAM M.R.
    • Organizer
      IUMS国際会議
    • Place of Presentation
      札幌コンベンションセンター(札幌)
    • Year and Date
      2011-09-10
    • Related Report
      2011 Annual Research Report
  • [Presentation] New Insight of Lantibiotic Engineering : Characterization of the enzymes, design and mode of action of lantibiotics, and immunity proteins2010

    • Author(s)
      Kenji Sonomoto, et al.
    • Organizer
      11th China-Japan-Korean Joint Symposium on Enzyme Engineering
    • Place of Presentation
      Green Land Hotel (中国・成都)
    • Year and Date
      2010-11-05
    • Related Report
      2010 Annual Research Report
  • [Book] Lactic Acid Bacteria and Bifidobacteria : Current Progress in Advanced Research2011

    • Author(s)
      Kenji Sonomoto
    • Total Pages
      286
    • Publisher
      Caister Academic Press
    • Related Report
      2011 Annual Research Report
  • [Book] Heterologus Gene Expression in E.coli (Methods in Molecular Biology 705)2010

    • Author(s)
      Jun-ichi Nagao, et al.
    • Total Pages
      12
    • Publisher
      The Humana Press INC
    • Related Report
      2010 Annual Research Report
  • [Remarks]

    • URL

      http://www.agr.kyushu-u.ac.jp/~microbt/html/

    • Related Report
      2011 Annual Research Report
  • [Remarks]

    • URL

      http://www.agr.kyushu-u.ac.jp/biosci-biotech/biseibutu/

    • Related Report
      2010 Annual Research Report

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Published: 2010-12-03   Modified: 2024-03-26  

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