Oxidative DNA damage contributes to aging, cancer, chronic inflammatory diseases, and degenerative diseases.  Guanine is particularly vulnerable to oxidation, forming guanine radicals which can form irreversible crosslinks to proteins.  DNA-protein crosslinks are an understudied yet significant form of oxidative damage.  Here, we sought to use fluorescence polarization to detect DNA-protein crosslinking caused by guanine oxidation. The unnatural base, 2-aminopurine (Ap), is a fluorescent adenine derivative and was incorporated into a 29-mer oligonucleotide. After annealing with its complement, the aminopurine-containing DNA was subjected to flash-quench treatment in the presence of histone of poly-Lysine. Without irradiation, the Ap duplex gives a fluorescence polarization value of 0.049. In the presence of histone of poly-Lysine, the numbers increase to 0.072 and 0.058, consistent with crosslinking to DNA.

Use of aminopurine-containing DNA to detect DNA-protein crosslinking by fluorescence polarization

243rd National Meeting (2012)

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