Studies on the binding and transcriptional properties of aflatoxin B1-8,9-epoxide.

[3H]Aflatoxin B1-8,9-epoxide ([3H]AFB1-8,9-epoxide), the putative ultimate carcinogen of AFB1, was synthesized and tested for its binding specificity to and transcriptional effect on several single- and double-stranded DNAs containing cytosine. The test was carried out over a 200-fold concentration range (i.e. 0.1-20 microgram [3H]AFB1-8,9-epoxide per 0.025 A260 units of DNA). The results show: (i) [3H]AFB1-8,9-epoxide bound preferentially to the double-stranded alternating co-polymer poly[d(G-C)] over the double-stranded poly(dG).poly(dC) and single-stranded poly(dG) or poly(dC) homopolymers. (ii) The binding affinity of [3H]AFB1-8,9-epoxide to poly(dC) was essentially the same as the poly(dG). (iii) Under identical conditions, [3H]AFB1-8,9-epoxide bound to poly(dG).poly(dC) 2.5-3 times more than to poly[d(I-C)]; however, poly[d(I-C)]-directed RNA synthesis was clearly more sensitive to [3H]AFB1-8,9-epoxide inhibition than poly(dG).poly(dC). Conversely, the binding affinity of [3H]AFB1-8,9-epoxide to poly(dC) and to poly[d(I-C)] was quite similar, yet poly(dC)-directed RNA synthesis was much more resistant to [3H]AFB1-8,9-epoxide inhibition than poly[d(I-C)]. (iv) After [3H]AFB1-8,9-epoxide was hydrolyzed to [3H]AFB1-8,9-dihydrodiol (0.01 N NaOH, 10 min 23 degrees C), it was no longer able to bind poly[d(G-C)] or to inhibit poly[d(G-C)]-directed RNA synthesis. These results confirm our earlier studies using microsome-activated AFB1 and AFB1-Cl2 that AFB1 after activation is able to bind cytosine in DNA, and the binding is not via AFB1-8,9-dihydrodiol. Furthermore, the results also suggest that AFB1 adducts may not have the same biological effect depending on the base, sequence as well as the conformation of the DNA where the adducts are formed.