Alkyladenine DNA glycosylase (AAG) is the only known human glycosylase capable of excising alkylated purines from DNA, including the highly mutagenic 1,N 6-ethenoadenine (εA) lesion. Here, we examine the ability of AAG to excise εA from a nucleosome core particle (NCP), which is the primary repeating unit of DNA packaging in eukaryotes. Using chemical synthesis techniques, we assembled a global population of NCPs in which A is replaced with εA. While each NCP contains no more than one εA lesion, the total population contains εA in 49 distinct geometric positions. Using this global εA-containing NCP system, we obtained kinetic parameters of AAG throughout the NCP architecture. We observed monophasic reaction kinetics across the NCP, but varying amounts of AAG excision. AAG activity is correlated with solution accessibility and local histone architecture. Notably, we identified some highly solution-accessible lesions that are not repaired well, and an increase in repair within the region of asymmetric unwrapping of the nucleosomal DNA end. These observations support in vivo work and provide molecular-level insight into the relationship between repair and NCP architecture.

If unrepaired, damage to genomic DNA can cause mutations and/or be cytotoxic. Single base lesions are repaired via the base excision repair (BER) pathway. The first step in BER is the recognition and removal of the nucleobase lesion by a glycosylase enzyme. For example, human oxoguanine glycosylase 1 (hOGG1) is responsible for removal of the prototypic oxidatively damaged nucleobase, 8-oxo-7,8-dihydroguanine (8-oxoG). To date, most studies of glycosylases have used free duplex DNA substrates. However, cellular DNA is packaged as repeating nucleosome units, with 145 base pair segments of DNA wrapped around histone protein octamers. Previous studies revealed inhibition of hOGG1 at the nucleosome dyad axis and in the absence of chromatin remodelers. In this study, we reveal that even in the absence of chromatin remodelers or external cofactors, hOGG1 can initiate BER at positions off the dyad axis and that this activity is facilitated by spontaneous and transient unwrapping of DNA from the histones. Additionally, we find that solution accessibility as determined by hydroxyl radical footprinting is not fully predictive of glycosylase activity and that histone tails can suppress hOGG1 activity. We therefore suggest that local nuances in the nucleosome environment and histone-DNA interactions can impact glycosylase activity.

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