Studies on non-covalent interactions between strongly conelated biomolecules
强相关生物分子间非共价相互作用的研究
基本信息
- 批准号:13031047
- 负责人:
- 金额:$ 18.05万
- 依托单位:
- 依托单位国家:日本
- 项目类别:Grant-in-Aid for Scientific Research on Priority Areas
- 财政年份:2001
- 资助国家:日本
- 起止时间:2001 至 2003
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
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.
反向促旋酶,一种正超螺旋DNA的酶,是唯一存在于所有嗜高温生物中的酶,但在所有嗜温生物中不存在。该酶由两个不同的结构域组成,一个解旋酶样结构域和另一个属于拓扑异构酶IA型家族的结构域。有人指出,如果反向旋转酶是极端嗜热生命的先决条件,那么生命的起源就不可能是极端嗜热生物,因为在反向旋转酶出现之前,这两个独立的结构域必须在不太嗜热的生物中独立进化。我们已经破坏了反向旋转酶基因从超嗜热古菌,Thermococcus kodakaraensis KOD1。一个单一的反向旋转酶基因,从基因组分析中确定,被替换为一个trpE标记基因,通过双交换重组在宿主菌株中断的trpE。PCR和Southern印迹分析证实了反向旋转酶基因的缺失。与该基因的缺失相一致,在宿主菌株中观察到的明显的正超螺旋活性在破坏菌株中未发现。在不同温度下,将干扰菌株的生长速率与宿主菌株的生长速率进行比较。我们发现,缺乏反向旋转酶导致生长迟缓,这种趋势在较高的温度下变得更大。然而,反向旋转酶基因的破坏在90℃下不会导致致死表型。这些结果提供了实验证据,证明反向旋转酶不是极端嗜热生命的先决条件,并且反向旋转酶的结构及其在极端嗜热生物中的存在不再排除极端嗜热生物是第一个进化的生物体的可能性。研究结果还提供了证据,证明DNA(基因组)的拓扑结构是在高温下维持生命的关键因素。
项目成果
期刊论文数量(64)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
W.Fukuda, T.Fukui, H.Atomi, T.Imanaka: "The First Characterization of an Archaeal GTP-dependent Phosphoenolpyruvate Carboxykinase from the Hyperthermophilic Archaeon, Thermococcus kodakaraensis KOD1."J.Bacteriol.. (in press). (2004)
W.Fukuda、T.Fukui、H.Atomi、T.Imanaka:“来自超嗜热古菌 Thermococcus kodakaraensis KOD1 的古菌 GTP 依赖性磷酸烯醇丙酮酸羧激酶的首次表征。”J.Bacteriol..(出版中)。
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T.Kanamori, N.Kanou, H.Atomi, T.Imanaka: "Enzymatic characterization of a prokaryotic urea carboxylase."J. Bacteriol.. 186(9). 2532-2539 (2004)
T.Kanamori、N.Kanou、H.Atomi、T.Imanaka:“原核尿素羧化酶的酶学表征。”J。
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N.Rashid, H.Imanaka, T.Kanai, T.Fukui, H.Atomi, T.: "A novel candidate for the true fructose 1,6-bisphosphatase in archaea"J. Biol. Chem.. 277(34). 30649-30655 (2002)
N.Rashid、H.Imanaka、T.Kanai、T.Fukui、H.Atomi、T.:“古细菌中真正的果糖 1,6-双磷酸酶的新候选者”J。
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H.Imanaka, T.Fukui, H.Atomi, T.Imanaka: "Gene cloning and characterization of fructose-1,6-bisphosphate aldolase from the hyperthermophilic archaeon Thermococcus"J. Biosci. Bioeng.. 94(3). 237-243 (2002)
H.Imanaka、T.Fukui、H.Atomi、T.Imanaka:“来自超嗜热古细菌 Thermococcus 的果糖 1,6-二磷酸醛缩酶的基因克隆和表征”J。
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N.Rashid, H.Imanaka, T.Fukui, H.Atomi, T.Imanaka: "Presence of a novel phosphopentomutase and a 2-deoxyribose 5-phosphate aldolase reveals a metabolic link between pentoses and central carbon metabolism in the hvperthermophilic archaeon Thermococcus kodak
N.Rashid、H.Imanaka、T.Fukui、H.Atomi、T.Imanaka:“一种新型磷酸戊糖变位酶和一种 2-脱氧核糖 5-磷酸醛缩酶的存在揭示了嗜热古细菌 Thermococcus 中戊糖和中心碳代谢之间的代谢联系
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ATOMI Haruyuki其他文献
ATOMI Haruyuki的其他文献
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{{ truncateString('ATOMI Haruyuki', 18)}}的其他基金
Exploring the possibilities of genome recombination
探索基因组重组的可能性
- 批准号:
22655053 - 财政年份:2010
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Challenging Exploratory Research
Determining the regulons and their functions in hyperthermophiles
确定超级嗜热菌的调节子及其功能
- 批准号:
21350092 - 财政年份:2009
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Novel metabolic pathway discovery based on contradictions between genome data and biochemical properties
基于基因组数据和生化特性之间矛盾的新代谢途径发现
- 批准号:
19310126 - 财政年份:2007
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Elucidation of gene regulation systems based on genome analysis
基于基因组分析阐明基因调控系统
- 批准号:
17350083 - 财政年份:2005
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Development of a system for enhancing protein thermostability in vivo using hyperthermophiles
利用超嗜热菌开发体内增强蛋白质热稳定性的系统
- 批准号:
15350100 - 财政年份:2003
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Anaerobic carbon dioxide fixation by microorganisms
微生物厌氧固定二氧化碳
- 批准号:
11450314 - 财政年份:1999
- 资助金额:
$ 18.05万 - 项目类别:
Grant-in-Aid for Scientific Research (B).
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