Reactions of deoxyribonucleotide bases with sulfooxymethyl or halomethyl polycyclic aromatic hydrocarbons induce unwinding of DNA supercoils

Abstract

Torsional stress in double-stranded DNA enables and regulates facets of chromosomal metabolism, replication, and transcription and requires regulatory enzymatic systems including topoisomerases and histone methyltransferases. As such, this machinery may be subject to deleterious effects from reactive mutagens, including ones from carcinogenic polycyclic aromatic hydrocarbon (PAH) adduct formation with DNA. Supercoiled plasmid DNA was investigated for its torsional responses to adducts formed in vitro from PAH benzylic carbocation reactive intermediates created spontaneously by release of leaving groups. PAH sulfate esters were found to (1) unwind DNA in a concentration dependent manner, and (2) provide maximum unwinding in a pattern consistent with known carcinogenicities of the parent PAHs, that is, 6-methylbenzo[a]pyrene > 7,12-methylbenz[a]anthracene > 3-methylcholanthrene > 9-methylanthracene > 7-methylbenz[a]anthracene > 1-methylpyrene. Supercoil unwinding was demonstrated to be dependent on the presence of sulfate or chloride leaving groups such that reactive carbocations were generated in situ by hydrolysis. In silico modeling of intercalative complex topology showed PAH benzylic carbocation reactive functional groups in alignment with target nucleophiles on guanine bases in a 5’-dCdG-3’ pocket in agreement with known formation of nucleotide adducts. Inhibitory or modulatory effects on PAH-induced supercoil unwinding were seen with ascorbic acid and an experimental antineoplastic agent Antineoplaston A10 in agreement with their known anticarcinogenic properties. In summary, the reactive PAH intermediates studied here undoubtedly participate in well-known mutational mechanisms such as frameshifts and apurinic site generation. However, they are also capable of random disruption of chromosomal supercoiling in a manner consistent with the known carcinogenicities of the parent compounds, and this mechanism may represent an additional detrimental motif worthy of further study for a more complete understanding of chemical carcinogenicity.

Keywords:

• DNA supercoiling
• DNA unwinding
• polycyclic aromatic hydrocarbons
• carcinogenicity
• sulfate ester reactivity

Acknowledgements

This paper is dedicated to the memory of Dr. James W. Flesher, without whose friendship, mentoring, inspiration, and provision of lab space and synthetic compounds this research would not have been possible. Thanks to Dr. Jamie Horn, Christine Kuhn and Mindy Leisure for their valuable assistance in this research. Thanks also to Dr. David M. Kaetzel for access to gel electrophoretic, UV transilluminator and photographic equipment. Finally, thanks to Dr. Jessica Fortin and Ms. Doreen Lehner for their helpful suggestions on review of the manuscript.

Disclosure statement

The author declares that he has no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Data availability

The author declares that reasonable requests for copies of original data will be reviewed and given appropriate responses.