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Chemical Creation of Artificial Restriction Enzyme and Its Application to Genomic Analysis

Research Project

Project/Area Number 05557111
Research Category

Grant-in-Aid for Developmental Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field 医薬分子機能学
Research InstitutionKYOTO UNIVERSITY

Principal Investigator

SUGIURA Yukio  Kyoto Univ.Professor, 化学研究所, 教授 (40025698)

Co-Investigator(Kenkyū-buntansha) OTSUKA Masami  Kyoto Univ.Associate Professor, 化学研究所, 助教授 (40126008)
Project Period (FY) 1993 – 1995
Project Status Completed (Fiscal Year 1995)
Budget Amount *help
¥15,200,000 (Direct Cost: ¥15,200,000)
Fiscal Year 1995: ¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 1994: ¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 1993: ¥6,700,000 (Direct Cost: ¥6,700,000)
KeywordsArtificial Restriction Enzymo / Bleomycin / DNA Cutting / Zinc Finger / Nuclease / Triple Stranded DNA / Human Geome / ヒトゲノム / 塩基配列特異性 / エンジイン / DNA結合蛋白質 / 人工リプレッサー / 転写因子 / 活性酸素 / ニッケル錯体 / グリシル・グリシル・ヒスチジン
Research Abstract

The molecular mechanism of bleomycin antibiotics facilitated design and synthesis of srtificial DNA cleaving molecules. Transformation of a sequence-specific DNA-binding molecule into a sequence-specific DNA-cleaving molecule allows identification of the preferred binding locations of these molecules on restriction fragments analyzed on sequencing gels. Various types of DNA-cleaving molecules have been demonstrated. Sequence-specific DNA-cleaving peptides and oligonucleotides equipped with a DNA-cleaving moiety have also been designed. Zinc finge motif is a novel DNA binding motif characterized by the unique role of zinc ; the protein folding and the DNA binding ability are goverened by the coordination of a zinc ion. In C_2H_2-type zinc finger, two cysteines and two histidines contribute to form a globular domain through zinc coordination. The zinc finger of C_2H_2-type gives promise of recognition for any DNA sequences because of its recognition mode. A C_2H_2-type zinc finger protein Sp1 has been converted into artificial sitespecific nuclease with an attached Ni-based DNA cleavage unit (Gly-Gly-His). This protein cleaved DNA at a single site near the Sp1 recognition sequence. The zinc finger-based nuclease is applicable to chromosome mapping and sequencing.

Report

(4 results)
  • 1995 Annual Research Report   Final Research Report Summary
  • 1994 Annual Research Report
  • 1993 Annual Research Report
  • Research Products

    (21 results)

All Other

All Publications (21 results)

  • [Publications] 永岡 真: "ジンクフィンガータンパク質の機能変換" 日本臨床. 53. 2589-2596 (1955)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 桑原 淳: "Znフィンガー型DNA切断分子の分子設計" 蛋白質核酸酵素. 40. 1457-1464 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] J. Kuwahara: "Binding of transcription factor Sp1 to GC box revealed by footprinting analysis" Biochemistry. 32. 5994-6001 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] A. Yonezawa: "DNA binding mode of class-IIS restriction endonuclease FokI revealed by DNA footprinting analysis" Biochim. Biophys. Acta. 1219. 369-379 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] M. Nagaoka: "A novel zinc finge-based DNA cutter" J. Am. Chem. Soc.116. 4085-4086 (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T. Kusakabe: "Sequence preference for strand cleavage of gapped duplexes by dynemicin A" Biochemistry. 34. 9944-9950 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] M.Nagaoka: "Conversion of zinc finger protein to artificial site-specific nuclease" Japanese J.Clinical Med.53. 2589-2596 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] J.Kuwahara: "Molecular design of a zinc fingertype DNA cutter" Protein, Nucleic Acids, and Enzyme. 40. 1457-1464 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] T.Kusakabe: "Sequence preference for strand cleavage of gapped duplexes by dynemic in A" Biochemistry. 34. 9944-9950 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] M.Nagaoka: "A novel zinc finger-based DNA cutter" J.Am.Chem.Soc.116. 4085-4086 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] A.Yonezawa: "DNA binding mode of class-IIS restriction endonuclease FokI revealed by DNA footprinting analysis" Biochim.Biophys.Acta. 1219. 369-379 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] J.Kuwahara: "Binding of transcription factor Sp1 to GC box revealed by footprinting analysis" Biochemistry. 32. 5994-6001 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1995 Final Research Report Summary
  • [Publications] 永岡真: "ジンクフィンガータンパ質の機能変換" 日本臨床. 53. 2589-2596 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] 桑原淳: "Znフィンガー型DNA切断分子の分子設計" 蛋白質核酸酵素. 40. 1457-1464 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] T. Kusakabe: "Sequence preference for strand cleavage of gapped duplexes by dynemicin A" Biochemistry. 34. 9944-9950 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] M.Nagaoka: "A novel zinc finger-based DNA cutter:Biosynthetic design and highly selective DNA cleavage" J.Am.Chem.Soc.116. 4085-4086 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] A.Yonezawa: "DNA binding mode of class-IIS restriction endonuclease FokI revealed by DNA footprinting analysis" Biochim.Biophys.Acta. 1219. 369-379 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] T.Matsumoto: "Alteration of binding mode and cutting site by G→I replacement in 5′-AGG of chromoprotein C-1027" Biochim.Biophys.Res.Commun.205. 1533-1538 (1994)

    • Related Report
      1994 Annual Research Report
  • [Publications] 杉浦幸雄: "長鎖DNAの化学切断" 蛋白質 核酸 酵素. 38. 516-523 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] J.Kuwahara: "Binding of transcription factor Sp1 to GC box DNA revealed by footprinting analysis" Biochemistry. 32. 5994-6001 (1993)

    • Related Report
      1993 Annual Research Report
  • [Publications] M.Nagaoka: "A novel zinc finger-based DNA cutter" J.Am.Chem.Soc.116(印刷中). (1994)

    • Related Report
      1993 Annual Research Report

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Published: 1993-04-01   Modified: 2016-04-21  

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