The binding of benzo(a)pyrene to DNA components of differing sequence complexity.

Abstract

An examination has been made of the binding, both in vitro and in vivo, of the benzo(a)pyrene (BP) adduct to DNA components of differing sequence complexity. Annealing was performed at low renaturation temperatures in the presence of high concentrations of formamide to minimize hydrocarbon-induced depurination. BHK-DNA was modified in vitro using a tritiated derivative of the ultimate carcinogenic metabolite of BP, 7alpha,8beta-di-hydroxy-9beta,10beta-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (BPDE). Co-renaturation of this modfied DNA with [14C]-thymidine-labelled BHK-DNA demonstrated that the hydrocarbon adduct did not interfere with strand annealing and showed that the BP adduct was distributed randomly throughout all DNA sequence classes. However, when the DNA of cells in culture was modified by [3H]BP, following metabolic activation, and mixed with [14C]-thymidine-labelled DNA, a small but reproducible difference in the renaturation of the two labels was found. This difference in renaturation profiles was not due to base-compositional effects since a similar result was found when the alternate 14C-label was present in guanine bases, the principal site of BP modification. The small difference in the renaturation of the two radioactive labels indicated an enrichment of the hydrocarbon on the most rapidly-renaturing sequence components (the palindromic and highly repetitive sequences) where it amounted to between 19 and 64% increased modification.