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AlkB-mediated oxidative demethylation reverses DNA damage in Escherichia coli

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Abstract

The bacterial AlkB protein is known to be involved in cellular recovery from alkylation damage; however, the function of this protein remains unknown. AlkB homologues have been identified in several organisms, including humans, and a recent sequence alignment study has suggested that these proteins may belong to a superfamily of 2-oxoglutarate-dependent and iron-dependent oxygenases (2OG-Fe(ii)-oxygenases)1. Here we show that AlkB from Escherichia coli is indeed a 2-oxoglutarate-dependent and iron-dependent DNA repair enzyme that releases replication blocks in alkylated DNA by a mechanism involving oxidative demethylation of 1-methyladenine residues. This mechanism represents a new pathway for DNA repair and the third type of DNA damage reversal mechanism so far discovered.

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Figure 1: Repair of MMS-treated M13mp18 ssDNA by treatment with AlkB.
Figure 2: AlkB-mediated release of radioactivity from [3H]methyl DNA.
Figure 3: Reverse phase HPLC elution profiles of radioactivity released from [3H]methyl ssDNA by AlkB and/or acid treatment.
Figure 4: Oxidative demethylation of 1-methyladenine lesions in DNA by the AlkB protein.

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References

  1. Aravind, L. & Koonin, E. V. The DNA-repair protein AlkB, EGL-9, and leprecan define new families of 2-oxoglutarate- and iron-dependent dioxygenases. Genome Biol. 2, 0007.1–0007.8 (2001)

    Article  Google Scholar 

  2. Pegg, A. E. Repair of O(6)-alkylguanine by alkyltransferases. Mutat. Res. 462, 83–100 (2000)

    Article  CAS  Google Scholar 

  3. Evensen, G. & Seeberg, E. Adaptation to alkylation resistance involves the induction of a DNA glycosylases. Nature 296, 773–775 (1982)

    Article  ADS  CAS  Google Scholar 

  4. Karran, P., Hjelmgren, T. & Lindahl, T. Induction of a DNA glycosylase for N-methylated purines is part of the adaptive response to alkylating agents. Nature 296, 770–773 (1982)

    Article  ADS  CAS  Google Scholar 

  5. Karran, P., Lindahl, T. & Griffin, B. Adaptive response to alkylating agents involves alternation in situ of O6-methylguanine residues in DNA. Nature 280, 76–77 (1979)

    Article  ADS  CAS  Google Scholar 

  6. Kataoka, H., Yamamoto, Y. & Sekiguchi, M. A new gene (alkB) of Escherichia coli that controls sensitivity to methyl methane sulfonate. J. Bacteriol. 153, 1301–1307 (1983)

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Wei, Y. F., Carter, K. C., Wang, R. P. & Shell, B. K. Molecular cloning and functional analysis of a human cDNA encoding an Escherichia coli AlkB homolog, a protein involved in DNA alkylation damage repair. Nucleic Acids Res. 24, 931–937 (1996)

    Article  CAS  Google Scholar 

  8. Dinglay, S., Trewick, S. C., Lindahl, T. & Sedgwick, B. Defective processing of methylated single-stranded DNA by E. coli AlkB mutants. Genes Dev. 14, 2097–2105 (2000)

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Parkinson, A. Casarett and Doull's Toxicology (ed. Claassen, C. D.) 113–186 (McGraw-Hill, New York, 1996)

    Google Scholar 

  10. Nash, T. The colorimetric estimation of formaldehyde by means of the Hantzsch reaction. Biochem. J. 55, 416–421 (1953)

    Article  CAS  Google Scholar 

  11. Rapoport, R., Hanukoglu, I. & Sklan, D. A fluorimetric assay for hydrogen peroxide, suitable for NAD(P)H-dependent superoxide generating redox systems. Anal. Biochem. 218, 309–313 (1994)

    Article  CAS  Google Scholar 

  12. Chen, B. J., Carroll, P. & Samson, L. The Escherichia coli AlkB protein protects human cells against alkylation-induced toxicity. J. Bacteriol. 176, 6255–6261 (1994)

    Article  CAS  Google Scholar 

  13. Singer, B. & Grunberger, D. Molecular Biology of Mutagens and Carcinogens (Plenum, New York, 1983)

    Book  Google Scholar 

  14. Amersham Pharmacia Biotech GST Gene Fusion System: User Manual; (1997) 〈www.amershambiosciences.com〉

    Google Scholar 

  15. Bjelland, S., Bjørås, M. & Seeberg, E. Excision of 3-methylguanine from alkylated DNA by 3-methyladenine DNA glycosylase I of Escherichia coli. Nucleic Acids Res. 21, 2045–2049 (1993)

    Article  CAS  Google Scholar 

  16. Wang, G., Rahman, M. S. & Humayun, M. Z. Replication of M13 single-stranded viral DNA bearing single site-specific adducts by Escherichia coli cell extracts: differential efficiency of translesion DNA synthesis for SOS-dependent and SOS-independent lesions. Biochemistry 36, 9486–9492 (1997)

    Article  CAS  Google Scholar 

  17. Palejwala, V. A., Simha, D. & Humayun, M. Z. Mechanisms of mutagenesis by exocyclic DNA adducts. Transfection of M13 viral DNA bearing a site-specific adduct shows that ethenocytosine is a highly efficient RecA-independent mutagenic noninstructional lesion. Biochemistry 30, 8736–8743 (1991)

    Article  CAS  Google Scholar 

  18. Chung, C. T., Niemela, S. L. & Miller, R. H. One-step preparation of competent Escherichia coli: transformation and storage of bacterial cells in the same solution. Proc. Natl Acad. Sci. USA 86, 2172–2175 (1989)

    Article  ADS  CAS  Google Scholar 

  19. Hofer, T. & Moller, L. Reduction of oxidation during the preparation of DNA and analysis of 8-hydroxy-2′-deoxyguanosine. Chem. Res. Toxicol. 11, 882–887 (1998)

    Article  CAS  Google Scholar 

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Acknowledgements

We thank M. Bjørås, L. Eide, K. Baynton, K. I. Kristiansen, J. Myllyharju, K. Skarstad and J. Klaveness for help and discussions, and L. Eide, K. Baynton and A. Klungland for critical reading of the manuscript. We are grateful to M. Bjørås for preparation of [3H]MNU-labelled DNA substrates, to L. Eide for the construction of the plasmid for expression of GST–AlkB, and to D. Daoudi for technical assistance. This work was supported by the Research Council of Norway and the Norwegian Cancer Society. E.S. also acknowledges support from the European Commission.

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  1. Centre for Molecular Biology and Neuroscience, and Institute of Medical Microbiology, University of Oslo, National Hospital, 0027, Oslo, Norway

    Pål Ø. Falnes, Rune F. Johansen & Erling Seeberg

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  1. Pål Ø. Falnes
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  2. Rune F. Johansen
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  3. Erling Seeberg
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Correspondence to Pål Ø. Falnes.

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Falnes, P., Johansen, R. & Seeberg, E. AlkB-mediated oxidative demethylation reverses DNA damage in Escherichia coli. Nature 419, 178–182 (2002). https://doi.org/10.1038/nature01048

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