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Chem Res Toxicol. 2013 Aug 19;26(8):1182-7. doi: 10.1021/tx400096m. Epub 2013 Jul 10.

Removal of N-alkyl modifications from N(2)-alkylguanine and N(4)-alkylcytosine in DNA by the adaptive response protein AlkB.

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1
Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.

Abstract

The AlkB enzyme is an Fe(II)- and α-ketoglutarate-dependent dioxygenase that repairs DNA alkyl lesions by a direct reversal of damage mechanism as part of the adaptive response in E. coli. The reported substrate scope of AlkB includes simple DNA alkyl adducts, such as 1-methyladenine, 3-methylcytosine, 3-ethylcytosine, 1-methylguanine, 3-methylthymine, and N(6)-methyladenine, as well as more complex DNA adducts, such as 1,N(6)-ethenoadenine, 3,N(4)-ethenocytosine, and 1,N(6)-ethanoadenine. Previous studies have revealed, in a piecemeal way, that AlkB has an impressive repertoire of substrates. The present study makes two additions to this list, showing that alkyl adducts on the N(2) position of guanine and N(4) position of cytosine are also substrates for AlkB. Using high resolution ESI-TOF mass spectrometry, we show that AlkB has the biochemical capability to repair in vitro N(2)-methylguanine, N(2)-ethylguanine, N(2)-furan-2-yl-methylguanine, N(2)-tetrahydrofuran-2-yl-methylguanine, and N(4)-methylcytosine in ssDNA but not in dsDNA. When viewed together with previous work, the experimental data herein demonstrate that AlkB is able to repair all simple N-alkyl adducts occurring at the Watson-Crick base pairing interface of the four DNA bases, confirming AlkB as a versatile gatekeeper of genomic integrity under alkylation stress.

PMID:
23773213
PMCID:
PMC3748507
DOI:
10.1021/tx400096m
[Indexed for MEDLINE]
Free PMC Article
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