1996 Fiscal Year Final Research Report Summary
Mutagenesis by Nucleoside Analogs and its Mechanism
| Project/Area Number |
08044291
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Institution | Okayama University |
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Principal Investigator |
HAYATSU Hikoya Okayama University, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (10012593)
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| Co-Investigator(Kenkyū-buntansha) |
LOAKES David Medecal Research Council, Laboratory of Molecular Biology, Scientist, 分子生物学研究部, 研究員
BROWN Daniel.M Medical Research Council, Laboratory of Molecular Biology, Attached Scientific W, 分子生物学研究部, 特別研究職
SHIMAMOTO Tadashi Okayama University, Gene Research Center, Assistant Professor, 遺伝子実験施設, 助手 (90187443)
NEGISHI Kazuo Okayama University, Gene Research Center, Associate Professor, 遺伝子実験施設, 助教授 (70116490)
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| Project Period (FY) |
1996
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| Keywords | P-nucleoside / Escherichia coli / Thymidine Kinase / Nucleoside analog / DNA polymerase / N^4-aminocytidine / Nickelion / Salmonella typhimurium |
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| Research Abstract |
We have shown that deoxyribosyl dihydropyrimido [4,5-c] [1,2] oxazin-7-one (dP) is a potent mutagen in Escherichia coli and Salmonella typhimurium. In E.coli, this deoxycytidine analog induces both GC-to-AT and AT-to GC transitions. No induced transversions are observed. It is highly mutagenic in a wild-type E.coli, but this is much reduced in a strain lacking thymidine kinase. The mutagenesis induced by dP is efficiently inhibited by the addition of thymidine. Partially purified thymidine kinase from E.coli catalyzes the phosphorylation of dP to its 5'-monophosphate. When E.coli was grown in the presence of dP,the nucleoside analogue was incorporated into its DNA.The content of dP in DNA was dependent on the concentration of dP added to the medium. The incorporation characteristics of the 5'-triphosphate of dP (dPTP) were also studied using E.coli DNA polymerase I large fragment. The results confirm that this triphosphate can be incorporated opposite both A and G in the template with similar efficiencies. This indicates that dP is metabolized as a thymidine analogue and the resulting triphosphate induces a high rate of mutagensis through replicational errors with the error level quite high.
N^4-Aminocytidine is a ptently mutagenic nucleoside analog, causing AT-to-GC and GC-to-AT transitions. This mutagenicity was enhanced by the presence of NiCl_2 in the assay on S.typhimurium and E.coli. We also found that N^4-aminocytidine and N^4-aminodeoxycytidine triphosphate can form a complex with Ni^<++>ion. Spectral changes of N^4-aminocytidene induced by Ni^<++> indicated that twe N^4-aminocytidene molecules can be complexed with one Ni^<++>ion. This complex formation may result in the inhibition of the enzymatic degradation of N^4-aminocytidine and/or in the enhancement of the error in DNA replication.
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