2001年
Oxidation of Thymine to 5-Formyluracil in DNA Promotes Misincorporation of dGMP and Subsequent Elongation of a Mismatched Primer Terminus by DNA Polymerase
Journal of Biological Chemistry
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- 巻
- 276
- 号
- 19
- 開始ページ
- 16501
- 終了ページ
- 16510
- 記述言語
- 英語
- 掲載種別
- 研究論文(学術雑誌)
- DOI
- 10.1074/jbc.M008598200
- 出版者・発行元
- AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
5-Formyluracil (fU) is a major oxidative thymine lesion generated by ionizing radiation and reactive oxygen species. In the present study, we have assessed the influence of fU on DNA replication to elucidate its genotoxic potential. Oligonucleotide templates containing fU at defined sites were replicated in vitro by Escherichia coli DNA polymerase I Klenow fragment deficient in 3'-5'-exonuclease, Gel electrophoretic analysis of the reaction products showed that fU constituted very weak replication blocks to DNA synthesis, suggesting a weak to negligible cytotoxic effect of this lesion. However, primer extension assays with a single dNTP revealed that fU directed incorporation of not only correct dAMP but also incorrect dGMP, although much less efficiently. No incorporation of dCMP and dTMP was observed. When fU was substituted for T in templates, the incorporation efficiency of dAMP (f(A) = V-max/K-m) decreased to 1/4 to 1/2, depending on the nearest neighbor base pair, and that of dGMP (f(G)) increased 1.1-5.6-fold. Thus, the increase in the replication error frequency (f(G)/f(A) for fU versus T) was 3.1-14.3-fold. The misincorporation rate of dGMP opposite fU (pK(a) = 8.6) but not T (pK(a) = 10.0) increased with pH (7.2-8.6) of the reaction mixture, indicating the participation of the ionized (or enolate) form of fU in the mispairing with G, The resulting mismatched fU:G primer terminus was more efficiently extended than the T:G terminus (8.2-11.3-fold). These results show that when T is oxidized to fU in DNA, fU promotes both misincorporation of dGMP at this site and subsequent elongation of the mismatched primer, hence potentially mutagenic.
- リンク情報
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- DOI
- https://doi.org/10.1074/jbc.M008598200
- CiNii Articles
- http://ci.nii.ac.jp/naid/30016334868
- PubMed
- https://www.ncbi.nlm.nih.gov/pubmed/11278425
- Web of Science
- https://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcAuth=JSTA_CEL&SrcApp=J_Gate_JST&DestLinkType=FullRecord&KeyUT=WOS:000168623100124&DestApp=WOS_CPL
- URL
- http://www.scopus.com/inward/record.url?eid=2-s2.0-0035844134&partnerID=MN8TOARS
- ID情報
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- DOI : 10.1074/jbc.M008598200
- ISSN : 0021-9258
- eISSN : 1083-351X
- CiNii Articles ID : 30016334868
- ORCIDのPut Code : 80281160
- PubMed ID : 11278425
- SCOPUS ID : 0035844134
- Web of Science ID : WOS:000168623100124