| Project/Area Number |
13031047
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| Research Category |
Grant-in-Aid for Scientific Research on Priority Areas
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| Allocation Type | Single-year Grants |
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| Review Section |
Science and Engineering
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| Research Institution | Kyoto University |
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Principal Investigator |
ATOMI Haruyuki Kyoto University, Graduate School of Engineering, Associate Professor, 工学研究科, 助教授 (90243047)
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| Co-Investigator(Kenkyū-buntansha) |
福居 俊昭 京都大学, 工学研究科, 助手 (80271542)
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| Project Period (FY) |
2001 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥28,200,000 (Direct Cost: ¥28,200,000)
Fiscal Year 2003: ¥8,000,000 (Direct Cost: ¥8,000,000)
Fiscal Year 2002: ¥7,800,000 (Direct Cost: ¥7,800,000)
Fiscal Year 2001: ¥12,400,000 (Direct Cost: ¥12,400,000)
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| Keywords | reverse gyrase / DNA topology / positive supercoiling / topoisomerase / hyperthermophiles / archaea / non-covalent bonds / Thermococcus / Rubisco / subunit間相互作用 / イオン結合 / Thermococcus / タンパク質の立体構造 / 耐熱性 / バイオサーファクタント / 界面活性剤 / Bacillus / サーファクチン / リポポリペプチド |
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| Research Abstract |
Reverse gyrase, an enzyme that positively supercoils DNA, is the only enzyme present in all hyperthennophiles, but absent in all mesophiles. The enzyme is composed of two distinct domains, one helicase-like domain and another domain that belongs to the topoisomerase type IA family. It has been pointed out that if reverse gyrase were a prerequisite for hyperthermophilic life, the origin of life could not have been a hyperthermophile, as the two separate domains must have independently evolved in less thermophilic organisms before the occurrence of reverse gyrase. We have disrupted the reverse gyrase gene from a hyperthermophilic archaeon, Thermococcus kodakaraensis KOD1. A single reverse gyrase gene, identified from genome analysis, was replaced with a trpE marker gene via double crossover recombination in a host strain disrupted of trpE. PCR and southern blot analyses were performed to confirm the absence of the reverse gyrase gene on the chromosome of the transformant. Consistent with the absence of the gene, an apparent positive supercoiling activity that was observed in the host strain was not found in the disruptant strain. Growth rates of the disruptant strain were compared with those of the host strain at various temperatures. We found that lack of reverse gyrase led to a retardation in growth, a tendency that became greater at higher temperatures. However, disruption of the reverse gyrase gene did not lead to a lethal phenotype at 90℃. The results provide experimental evidence that reverse gyrase is not a prerequisite for hyperthermophilic life, and that the structure of reverse gyrase and its presence in hyperthermophiles no longer rule out the possibility that hyperthermophiles were the first organisms to evolve. The results also provide evidence that the topology of DNA (the genome) is a key factor for maintaining life at high temperature.
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