8-Methyl-2'-deoxyguanosine (8-medGuo) has been shown to be a major stable alkylation product of 2'-deoxyguanosine induced by methyl radical attack on DNA. Moreover, by using primer extension assays, the latter DNA modification has recently been reported to be a miscoding lesion by generating G to C and G to T transversions and deletions in vitro. However, no data have been reported up to now, concerning the processing of this C8-alkylated nucleoside by the DNA repair machinery. Therefore, we have investigated the capability of excision of 8-methylguanine (8-meGua) site specifically incorporated into oligonucleotide substrates by several bacterial, yeast and mammalian DNA N-glycosylases. The results show that the 3-methyladenine (3-meAde) DNA glycosylase II (AlkA protein) from Escherichia coli is the only DNA N-glycosylase tested able to remove 8-meGua from double-stranded DNA fragments. Moreover, the activity of AlkA for 8-meGua varied markedly depending on the opposite base in DNA, being the highest with Adenine and Thymine and the lowest with Cytosine and Guanine. The removal of 8-meGua by AlkA protein was compared to that of 7-methylguanine (7-meGua) and hypoxanthine (Hx). The rank of damage as a substrate for AlkA being 7-meGua>8-meGua>Hx. In contrast, the human 3-meAde DNA N-glycosylase (Mpg) is not able to release 8-meGua paired with any of the four DNA bases. We also show that, DNA N-glycosylases involved in the removal of oxidative damage, such as Fpg or Nth proteins from E. coli, Ntg1, Ntg2 or Ogg1 proteins of Saccharomyces cerevisiae, or human Ogg1 do not release 8-meGua placed opposite any of the four DNA bases. Furthermore, HeLa and Chinese hamster ovary (CHO) cell free protein extracts do not show any cleavage activity at 8-meGua paired with adenine or cytosine, which suggests the absence of base excision repair (BER) of this lesion in mammalian cells.

• PMID: 12509232
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Gasparutto D, Dhérin C, Boiteux S, Cadet J

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