We have obtained evidence the anti-BaP diol epoxide, the activated form of the pre-carcinogen benzo[a]pyrene (BaP), intercalates into DNA. This conclusion is based on the following evidence, 1) the hydrocarbon in the complex exhibits a red shift in its uv spectrum 2) fluorescence of the hydrocarbon is quenched 3) magnesium and spermidine result in dissociation of the complex and 4) the hydrocarbon unwinds SV40 DNA by 18 degrees. Preliminary temparture-jump relaxation kinetics exhibit three exponentials with kinetic constants 60, 650 and 4200 Mus, results which suggest that intercalation is a three step process. Alternative explanations for these three exponentials are different binding sites or stereoselective binding of the two enantiomers of the anti-BaP diol epoxide. This proposal is designed to investigate the mechanism of intercalation of BaP diol epoxides. We propose to synthesize and resolve the metabolically produced (+) and (-) anti- and (+) and (-) syn-BaP diol epoxides. We will investigate the mechanism of intercalation in terms of hydrocarbon structural modifications (enantiomers and diastereomers) and polydeoxynucleotide composition (sequences and left handed helix). We propose to synthesize a stable aziridine analog of the BaP diol epoxide (replacing the epoxy oxygen with nitrogen), which will be used in spectroscopic studies on the conformation of the intercalation complex. We will investigate the mechanism of intercalation of t-jump kinetics and by a determination of thermodynamic and equioibrium parameters of the physical reactions. Sequence specificity will be investigated by measuring complex formation with the 16 possible deoxydinucleoside phosphates. It will be of interest to compare the mechanism of polycyclic aromatic hydrocarbon intercalation with that of water soluble drugs since the latter can under simple electrostatic interactions and the former cannot. The biological properties of the four diol epoxides differ considerably, e.g., the (+)anti-BaP diol epoxide is the most mutagenic and carcinogenic form of the activated carcinogen. The physical interactions that these hydrocarbons undergo with DNA will help explain these differences at the molecular level.
Showing the most recent 10 out of 11 publications
