Adenine DNA glycosylase MUTYH has a well-established role in preventing mutations associated with 8-oxo-gunaine damage within the cell. Remarkably, mRNA expression levels of MUTYH is highly associated with decreased survival of cells treated with the SN1 methylating agent N-methyl-N’-nitro-N-nitrosoguanidine (MNNG), contradicting a pro-survival mechanism for the enzyme (Fry et al. Genes Dev. 22(19):2621-6, 2008). Mutyh-/- mouse embryonic fibroblasts (MEFs) ectopically expressing catalytically-inactive (D222N) MUTYH have survival levels compared to knockout cells when treated with MNNG, supporting a MUTYH-mediated catalytic mechanism for potentiating MNNG cytotoxicity. Furthermore, small molecules that form adducts with apurinic/apyrimidinic (AP) sites induced by MNNG abolish the enhanced survival of Mutyh-/- MEFs while having no additive cytotoxicity to wild-type MEFs. Here, the ability of catalytically-inactive and wild-type mouse Mutyh to form covalent interactions with MNNG-treated DNA was tested in vitro. Protein-DNA trapped intermediates were then subjected to proteomics analysis using MALDI-MS to identify potential amino acid residues involved in AP-site interactions. To test the hypothesis that Mutyh has novel methylpurine glycosylase activity, an LC-tandem mass spectrometry method was used to detect and quantify methylpurine adducts excised from genomic DNA treated with MNNG.

Elucidation of the role of adenine DNA glycosylase mutyh in MNNG-mediated cell cytotoxicity

253rd National Meeting (2017)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.