2000 Fiscal Year Final Research Report Summary
Construction of evolutionary reactor for directed evolution of enzymes
| Project/Area Number |
09555253
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
生物・生体工学
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| Research Institution | Osaka University |
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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)
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| 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)
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| Project Period (FY) |
1997 – 2000
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| Keywords | Experimental molecular evolution / Interaction / Coexistence of closely related genes / Plasticity of fitness / Diversification / Glutamine synthetase gene |
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| 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.
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Research Products
(8 results)
