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Construction of evolutionary reactor for directed evolution of enzymes

Research Project

Project/Area Number 09555253
Research Category

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

Allocation TypeSingle-year Grants
Section展開研究
Research Field 生物・生体工学
Research InstitutionOsaka University

Principal Investigator

URABE Itaru  Department of Biotechnology Graduate School of Engineering, Osaka University, Professor, 大学院・工学研究科, 教授 (60029246)

Co-Investigator(Kenkyū-buntansha) NEGORO Seiji  The Faculty of Engineering, Himeji Inetitute of Technology, Professor, 工学部, 教授 (90156159)
SHIMA Yasufumi  Department of Biotechnology Graduate School of Engineering, Osaka University, assistant Professor, 大学院・工学研究科, 助手 (50187423)
YOMO Tetsuya  Department of Biotechnology Graduate School of Engineering, Osaka University, associate Professor, 大学院・工学研究科, 助教授 (00222399)
Project Period (FY) 1997 – 2000
Project Status Completed (Fiscal Year 2000)
Budget Amount *help
¥13,000,000 (Direct Cost: ¥13,000,000)
Fiscal Year 2000: ¥2,500,000 (Direct Cost: ¥2,500,000)
Fiscal Year 1999: ¥2,500,000 (Direct Cost: ¥2,500,000)
Fiscal Year 1998: ¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1997: ¥4,900,000 (Direct Cost: ¥4,900,000)
KeywordsExperimental molecular evolution / Interaction / Coexistence of closely related genes / Plasticity of fitness / Diversification / Glutamine synthetase gene / 自己複製系 / 適応度地形 / DNA合成酵素
Research Abstract

A simplified experimental evolution encompassing the essence of natural one was designed in an attempt to understand the involved mechanism. In our system, molecular evolution was observed through three serial cycles of consecutive random mutagenesis of the glutamine synthetase gene and chemostat culture of the transformed Escherichia coli cells containing the mutated genes. Selection pressure was imposed solely on the glutamine synthetase gene when varieties of mutant genes compete in an unstructured environment of the chemostat. The molecular phylogeny and population dynamics were deduced from the nucleotide sequences of the genes isolated from each of the chemostat runs. An initial mutant population in each cycle, comprised of diversified closely-related genes, ended up with several varieties of mutants in a state of coexistence. Competition between two mutant genes in the final population of the first cycle ascertained that the observed coexisting state is not an incidental event and that cellular interaction via environmental nutrients is a possible mechanism of coexistence. In addition, the mutant gene once extinct in the previous passage was found to have the capacity to reinvade and constitute the gene pool of the later cycle of molecular evolution. These results, including the kinetic characteristics of the purified wild-type and mutant glutamine synthetases in the phylogenetic tree, revealed that the enzyme activity had diverged, rather than optimized, to a fittest value during the course of evolution. Here, we proposed that the plasticity of the fitness of a gene in consequence of cellular interaction via the environment is an essential mechanism governing molecular evolution.

Report

(5 results)
  • 2000 Annual Research Report   Final Research Report Summary
  • 1999 Annual Research Report
  • 1998 Annual Research Report
  • 1997 Annual Research Report
  • Research Products

    (12 results)

All Other

All Publications (12 results)

  • [Publications] Tetsuya Yomo: "In vitro self-replication system as a minimum set of life."Artificial Life. 5. 402-405 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Tetsuya Yomo: "Kinetic equation for template-dependent polymerase reactions and its application to thermus thermophilus DNA polymerase."Journal of Fermentation and bioengineering. 86(4). 379-384 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Akiko Kashiwagi: "How small can the difference among competitors by for coexistence to occur"Researches on Population Ecology. 40(2). 223-226 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Akiko Kashiwagi: "Plasticity of fitness and diversification process during an experimental molecular evolution."J.Mol.Evol., in press.

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Tetsuya Yomo et al.: "In vitro self-replication system as a minimum set of life."Artificial Life. 5. 402-405 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Tetsuya Yomo et al.: "Kinetic equation for template-dependent polymerase reactions and its application to thermus thermophilus DNA polymerase."Journal of Fermentation and bioengineering. 86(4). 379-384 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Akiko Kashiwagi et al.: "How small can the difference among competitors be for coexistence to occur."Researches on Population Ecology. 40(2). 223-226 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Akiko Kashiwagi et al.: "Plasticity of fitness and diversification process during an experimental molecular evolution."Journal of Molecular Evolution. (in press).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Kashiwagi Akiko: "Flexibility of fitness and diversification process during an exper imental molecular evolution."J.Mol.Evol.,. (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] T.Yomo et al.: "Kinetic equation for template-dependent polymerase reactions and its application to Thermus thermophilus DNA polymerase" J.Ferment.Bioeng.86(4). 379-384 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] A.Kashiwagi et al.: "How small can the difference among competitors be for coexistence to occur" Res.Popul.Ecol.40(2). 223-226 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] T,Yomo et al.: "In vitro self-replication system as a minimum set of life" Arti ficial Life. 5. 402-405 (1997)

    • Related Report
      1997 Annual Research Report

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Published: 1997-04-01   Modified: 2016-04-21  

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