2000 Fiscal Year Final Research Report Summary
Repair Mechanisms of DNA Double Strand Breaks
| Project/Area Number |
10044206
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| Research Category |
Grant-in-Aid for Scientific Research (A).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Molecular biology
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| Research Institution | Osaka University |
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Principal Investigator |
SHINAGAWA Hideo Research Institute for Microbial Diseases, Osaka Univ. Professor, 微生物病研究所, 教授 (40029799)
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| Co-Investigator(Kenkyū-buntansha) |
IWASAKI Hiroshi Research Institute for Microbial Diseases, Research Assiate, 微生物病研究所, 助手 (60232659)
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| Project Period (FY) |
1998 – 2000
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| Keywords | Double Stand breaks / DNA recombination / Hjc protein / Holliday junction / Escherichia coli / Thermophilic archaea / Thermophilic bacteria / Yeast |
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| Research Abstract |
DNA double strand breaks occur quite frequently during DNA replication not only by external genotoxic agents but also by genotoxic agents internally produced as a result of normal cellular metabolism. To elucidate the repair mechanisms of DNA double strand breaks, we employed Escherichia coli and a thermophilic bacterium as model prokaryotes, Saccharomyces cerevisiaea and Schizosaccharomyces pombe as model eukaryotes, and Pyrococcus furiosus as a model archaea.
Holliday junctions are formed as recombination intermediates and during DNA replication when replication fork is blocked. RuvABC proteins are involved in processing the Holliday junctions in prokaryotes. To gain insights into molecular mechanisms of Holliday junction resolution by RuvABC, we combined mutational analysis of these proteins with ctystallographic studies. A large number of RuvABC mutant proteins have been analyzed in vivo and in vitro. The three dimensional structure of RuvA-Holliday junction complex was revealed by
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crystallographic analysis and the three domains of RuvA were assigned to differerest functions. The crystal structure of monomeric RuvB protein from thermophilic bacteria was elucidated and the hexameric model structure of RuvB was proposed based on the similarity with the crystal structures of AAA^+ ATPases recently revealed. Based on mutational studies, two basic residues and a phenylalanine of RuvC critically important for the interactions with Holliday junction were revealed.
We identified 4 novel genes involved in repair of DNA double strand breaks in S.pombe. They are also involved in DNA replication and thus link DNA recombination with replication.
We studied the function of Mgs (Maintenance of Genome Stability) proteins which are homologous to RuvB motor protein and highly conserved from bacteria to humans. It has DNA-dependent ATPase and DNA annealing activities. We demonstrated that it is required to maintain genome stability by regulating DNA superhelicity during DNA replication together with RecQ helicases and topoisomerases. Less
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[Publications] Iwasaki,H.,YW.Han,T.Okamoto,T.Ohnishi,M.Yoshikawa,K.Yamada,H.Toh,H.Daiyasu,T.Ogura & H.Shinagawa: "Mutational analysis of the functional motifs of RuvB, an AAA^+ class helicase and motor protein for Holliday junction branch migration."Mol.Microbiol.. 36. 528-538 (2000)
Description
「研究成果報告書概要(和文)」より
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[Publications] Iwasaki, H., YW.Han, T.Okamoto, T.Ohnishi, M.Yoshikawa, K.Yamada, H.Toh, H.Daiyasu, T.Ogura & H.Shinagawa.: "Mutational analysis of the functional motifs of RuvB, an AAA^+ class helicase and motor protein for Holliday junction branch migration."Mol.Microbiol.. 36. 528-538 (2000)
Description
「研究成果報告書概要(欧文)」より
