Electrochemical Study on the Effects of Epigenetic Cytosine Methylation on <i>Anti</i>-Benzo[<i>a</i>]pyrene Diol Epoxide Damage at TP53 Oligomers

<i>Anti</i>-benzo­[<i>a</i>]­pyrene-<i>r</i>-7,<i>t</i>-8-dihydrodiol-<i>t</i>-9,10-epoxide (<i>anti</i>-BPDE) is a known carcinogen that damages DNA, and this damage is influenced by the DNA sequence and epigenetic factors. The influence of epigenetic cytosine methylation on the reaction with <i>anti</i>-BPDE at a known hotspot DNA damage site was studied electrochemically. Gold electrodes were modified with thiolated DNA oligomers spanning codons 270–276 of the TP53 gene. The oligomers exhibited 5-carbon cytosine methylation at the codon 273 location on the bound probe, the acquired complementary target, or both. Redox active diviologen compounds of the form C₁₂H₂₅V²⁺C₆H₁₂V²⁺C₁₂H₂₅ (V²⁺ = 4,4′-bipyridyl or viologen, C12-Viologen) were employed to detect <i>anti</i>-BPDE damage to DNA. DNA was exposed to racemic (±)- or enantiomerically pure (+)-<i>anti</i>-BPDE solutions followed by electrochemical interrogation in the presence of C12-Viologen. Background subtracted square wave voltammograms (SWV) showed the appearance of two peaks at approximately −0.38 V and −0.55 V vs Ag/AgCl upon <i>anti</i>-BPDE exposure. The acquired voltammetry is consistent with singly reduced C12-Viologen dimers bound at two different DNA environments, which arise from BPDE damage and are influenced by cytosine methylation and BPDE stereochemical considerations. UV spectroscopic and mass spectrometric methods employed to validate the electrochemical responses showed that (+)-<i>anti</i>-BPDE primarily adopts a minor groove bound orientation within the oligomers while selectively targeting the nontranscribed ssDNA sequence within the duplexes

Prospects for applying synthetic biology to toxicology: future opportunities and current limitations for the repurposing of cytochrome P450 systems

The 30 years since the inception of Chemical Research in Toxicology, game-changing advances in chemical and molecular biology, the fundamental disciplines underpinning molecular toxicology, have been made. While these have led to important advances in the study of mechanisms by which chemicals damage cells and systems, there has been less focus on applying these advances to prediction, detection, and mitigation of toxicity. Over the last ∼15 years, synthetic biology, the repurposing of biological "parts" in systems engineered for useful ends, has been explored in other areas of the biomedical and life sciences, for such applications as detecting metabolites, drug discovery and delivery, investigating disease mechanisms, improving medical treatment, and producing useful chemicals. These examples provide models for the application of synthetic biology to toxicology, which, for the most part, has not yet benefited from such approaches. In this perspective, we review the synthetic biology approaches that have been applied to date and speculate on possible short to medium term and "blue sky" aspirations for synthetic biology, particularly in clinical and environmental toxicology. Finally, we point out key hurdles that must be overcome for the full potential of synthetic biology to be realized