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Time series visualization of DNA replication initiation process by protein X-ray crystallography

Research Project

Project/Area Number 25840027
Research Category

Grant-in-Aid for Young Scientists (B)

Allocation TypeMulti-year Fund
Research Field Structural biochemistry
Research InstitutionNational Institute of Genetics

Principal Investigator

ITOU Hiroshi  国立遺伝学研究所, 細胞遺伝研究系, 特任研究員 (10390626)

Research Collaborator ITOH Tateo  信州大学, 理学部, 特任教授 (40051817)
Project Period (FY) 2013-04-01 – 2015-03-31
Project Status Completed (Fiscal Year 2014)
Budget Amount *help
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2014: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
Fiscal Year 2013: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
KeywordsDNA複製開始因子 / タンパク質DNA複合体 / プラスミド / Rep / PriCT / X線結晶構造解析法 / PriCT領域 / X線結晶構造解析 / タンパク-DNA複合体
Outline of Final Research Achievements

Duplex DNA is generally unwound by protein oligomers prior to replication. The Rep protein of plasmid ColE2 is an essential initiator for plasmid DNA replication. This protein binds the replication origin (Ori) in a sequence-specific manner as a monomer and unwinds DNA. We determined the crystal structure of the DNA-binding domain of Rep (E2Rep-DBD) in complex with Ori. The structure unveils the basis for Ori-specific recognition by the E2Rep-DBD and also reveals that it unwinds DNA by the concerted actions of its three structural modules. The structure also shows that the functionally unknown PriCT domain plays a central role in DNA unwinding. The conservation of the PriCT domain in the C-termini of some archaeo-eukaryotic primases indicates that it probably plays a similar role in these proteins. This is the first report providing the structural basis for the functional importance of the conserved PriCT domain and also reveals a novel mechanism for DNA unwinding by a single protein.

Report

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

    (2 results)

All 2015 2013

All Journal Article (1 results) (of which Peer Reviewed: 1 results,  Acknowledgement Compliant: 1 results) Presentation (1 results)

  • [Journal Article] Structural basis for replication origin unwinding by an initiator primase of plasmid ColE2-P9: Duplex DNA unwinding by a single protein2015

    • Author(s)
      H.Itou, M.Yagura, Y.Shirakihara, T.Itoh
    • Journal Title

      Journal of Biological Chemistry

      Volume: 290 Issue: 6 Pages: 3601-6311

    • DOI

      10.1074/jbc.m114.595645

    • Related Report
      2014 Annual Research Report
    • Peer Reviewed / Acknowledgement Compliant
  • [Presentation] Structural basis of replication origin binding and double-stranded DNA unwinding by an initiator-primase ColE2-Rep protein2013

    • Author(s)
      Hiroshi Itou, Masaru Yagura, Tateo Itoh and Yasuo Shirakihara
    • Organizer
      International Conference on Structural Genomics 2013
    • Place of Presentation
      北海道、札幌市、京王プラザホテル札幌
    • Related Report
      2013 Research-status Report

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

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