2005 Fiscal Year Final Research Report Summary
Identification and characterization of base excision repair enzymes involved in the repair of oxidative DNA damage
| Project/Area Number |
15310038
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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 | 一般 |
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| Research Field |
Risk sciences of radiation/Chemicals
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
IDE Hiroshi Hiroshima University, Graduate School of Science, Professor, 大学院・理学研究科, 教授 (30223126)
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| Co-Investigator(Kenkyū-buntansha) |
TERATO Hiroaki Hiroshima University, Graduate School of Science, Research Associate, 大学院・理学研究科, 助手 (00243543)
KUBO Kihei Osaka Prefecture University, Graduate School of Agriculture and Biological Science, Professor, 大学院・生命環境科学研究科, 教授 (40117619)
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| Project Period (FY) |
2003 – 2005
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| Keywords | DNA damage / base excision repair enzymes / DNA glycosylase / crosslink |
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| Research Abstract |
In this study, we have performed identification and characterization of mammalian DNA glycosylases to elucidate the repair mechanism of oxidative DNA damage in mammalian cells. The findings of this research are summarized as follows.
(1) DNA damage recognition proteins were isolated from HeLa cell extracts by mechanism-based trapping assays using oxanine-containing oligonucleotides as probes. About 14 proteins (28-69 kDa) were identified as trapped products in SDS-PAGE analysis. Mass fingerprinting analysis of trapped products indicates, together with histone, several proteins that might be involved in DNA repair. Detailed analysis of their function is ongoing.
(2) DNA glycosylase activity for 5-formyluracil (fU) was isolated from rat liver and identified as SMUG1. Human SMUG1 recognized uracil and its derivatives bearing an oxidized group at the ring C5 position, i.e., fU, 5-hydroxymethyluracil, and 5-hydroxyuracil. SMUG1 accounted for dominant activities for these lesions in HeLa cells. Thus, SMUG1 is a new member of DNA glycosylases involved in the repair of oxidative damage.
(3) The damage specificities of human glycosylases (hNTH1, hNEIL1, and hNEIL2) have been characterized and compared to those of E.coli counterparts. Despite being homologues, human and E.coli homologues (Endo III vs.hNTH1, Endo VIII vs.hNEIL1) exhibit significantly different damage preferences, particularly, for thymine glycol stereoisomers and formamidopymidine, hNEIL2 exhibits only very weak N-glycosylase activity.
(4) Analysis of repair activity of E.coli Endo IV and yeast APN1 suggests that they initiate nucleotide incision repair (NIR) for free radical-induced DNA lesions. NIR may constitute an alternative or backup repair pathway for the base excision repair (BER) pathway in cells.
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