Nucleic Acids Research Advance Access originally published online on February 17, 2009
Nucleic Acids Research 2009 37(7):2116-2125; doi:10.1093/nar/gkp057
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Nucleic Acids Research, 2009, Vol. 37, No. 7 2116-2125
© 2009 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Nucleic Acid Enzymes |
KsgA, a 16S rRNA adenine methyltransferase, has a novel DNA glycosylase/AP lyase activity to prevent mutations in Escherichia coli
1Laboratory of Radiation Biology, 2Laboratory of Biological Chemistry, Graduate School of Science, Kyoto University, Sakyo-ku, Kyoto 606-8502 and 3Department of Biomolecular Sciences, Graduate School of Life Sciences, Tohoku University, Aoba-ku, Sendai 980-8577, Japan
*To whom correspondence should be addressed. Tel: +81 75 753 4097; Fax: +81 75 753 4087; Email: qmzhang{at}kingyo.zool.kyoto-u.ac.jp
Received October 29, 2008. Revised January 20, 2009. Accepted January 21, 2009.
The 5-formyluracil (5-foU), a major mutagenic oxidative damage of thymine, is removed from DNA by Nth, Nei and MutM in Escherichia coli. However, DNA polymerases can also replicate past the 5-foU by incorporating C and G opposite the lesion, although the mechanism of correction of the incorporated bases is still unknown. In this study, using a borohydride-trapping assay, we identified a protein trapped by a 5-foU/C-containing oligonucleotide in an extract from E. coli mutM nth nei mutant. The protein was subsequently purified from the E. coli mutM nth nei mutant and was identified as KsgA, a 16S rRNA adenine methyltransferase. Recombinant KsgA also formed the trapped complex with 5-foU/C- and thymine glycol (Tg)/C-containing oligonucleotides. Furthermore, KsgA excised C opposite 5-foU, Tg and 5-hydroxymethyluracil (5-hmU) from duplex oligonucleotides via a β-elimination reaction, whereas it could not remove the damaged base. In contrast, KsgA did not remove C opposite normal bases, 7,8-dihydro-8-oxoguanine and 2-hydroxyadenine. Finally, the introduction of the ksgA mutation increased spontaneous mutations in E. coli mutM mutY and nth nei mutants. These results demonstrate that KsgA has a novel DNA glycosylase/AP lyase activity for C mispaired with oxidized T that prevents the formation of mutations, which is in addition to its known rRNA adenine methyltransferase activity essential for ribosome biogenesis.