Halogenated hydrocarbons are major industrial chemicals and are of environmental and lexicological concern because of potential hazards to human health. Several of these compounds are bifunctional electrophiles and are bioactivated to crosslinking agents, linking glutathione (GSH) or the DMArepair protein C^-alklyguanine DNA-alkyltransferase (AGT) to DNA. Some compounds of interest in this group are ethylene and methylene dihalides and the related bis-electrophile butadiene diepoxide, a metabolite of the monomer butadiene. Wepropose to: (i) Characterize mechanisms of mutagenesis related to the AGT-DNA crosslinks, including the chemical characterization of DNA adducts and crosslinks, the possible involvement of translesion polymerases and nucleotide excision repair inmutagenic processing, and the relative contribution of the GSH and AGT pathways in producing mutationsin vivo, (ii) Characterize interactions of AGT with DNA and alkylated DNA, includingrates of AGT: DNA association and dissociation and of baseflipping,(iii) Search for other nuclear/DNA-binding proteins that also enhance Ws-electrophile-generated mutations via an AGT-like mechanism, both in bacteria and mammaliancells. Candidates are proliferating cell nuclear antigen (PCNA), uracil DNAglyosylase, and possibly OxyR. (iv) Quantify DNA adducts formed with GSH and CH2C12, CHzBr2, and ethylene dichloride in vitro and in vivo using rapid digestion, chromatography, and accelerator mass spectrometry for unstable DNA adducts.This information will be related to the halogen order, rates of GSH conjugation, and mutagenicity, and should be useful in assessments of the carcinogenic potentials of these and related compounds, (v) Characterize the mechanism by which GSH enhances the mutations produced by the Ms-functional electrophile butadiene diepoxide. Progress in the five Specific Aims described in this work will provide a better understanding of the interactions of these and other bis- functional electrophiles with DNA, and several aspects of the work have relevance :o the risk assessment of halogenated hydrocarbons.

National Institute of Health (NIH)
National Institute of Environmental Health Sciences (NIEHS)
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Cancer Etiology Study Section (CE)
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Carlin, Danielle J
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Vanderbilt University Medical Center
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Su, Yan; Egli, Martin; Guengerich, F Peter (2017) Human DNA polymerase ? accommodates RNA for strand extension. J Biol Chem 292:18044-18051
Sedgeman, Carl A; Su, Yan; Guengerich, F Peter (2017) Formation of S-[2-(N6-Deoxyadenosinyl)ethyl]glutathione in DNA and Replication Past the Adduct by Translesion DNA Polymerases. Chem Res Toxicol 30:1188-1196
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Patra, Amitraj; Zhang, Qianqian; Guengerich, F Peter et al. (2016) Mechanisms of Insertion of dCTP and dTTP Opposite the DNA Lesion O6-Methyl-2'-deoxyguanosine by Human DNA Polymerase ?. J Biol Chem 291:24304-24313
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