Mechanism of benzo[a]pyrene diol epoxide induced deoxyribonucleic acid strand scission

Approximately 1% of (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BaP-diol epoxide) DNA alkylation sites rearrange with strand scission at neutral pH. Phosphotriester hydrolysis and depurination/depyrimidination strand scission were critically examined as possible mechanisms for this phenomenon. The catalysis of nicking by alkali and the inhibition of nicking by counterions were consistent with either mechanism. The kinetics of nicking, however, were characteristic of a multistep reaction such as depurination/depyrimidination strand scission and the detection of apurinic sites in BaP-diol epoxide alkylated DNA strongly supported this mechanism. The number of such sites, especially at lower reaction levels, was probably sufficient to account for strand scission. No direct evidence was obtained for nicking occurring through phosphotriester hydrolysis. Studies with model substrates, including dibutyl phosphate, DNA homopolymers, and TMV RNA, indicated that if BaP-diol epoxide forms phosphotriesters in DNA or RNA, they do not hydrolyze with strand scission. Besides apurinic/apyrimidinic sites, a second alkali-sensitive rearrangement product was present in BaP-diol epoxide modified DNA. These latter sites accumulated with time and after 24 h accounted for as much as 4% of the initial alkylation events. Although relatively stable at neutrality, they spontaneously nicked the DNA backbone at high pH. It is possible that these sites represent a rearrangement of the major N2 guanine adduct.