Covalent Binding of Reactive Intermediates of Xenobiotics to Mitochondrial DNA

Vamvakas, Spyridon; Bittner, Detlef; Dekant, W.; Anders, Monika

doi:10.1016/b978-0-12-461205-1.50044-8

Cited by 2 publications

(1 citation statement)

References 21 publications

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“…The damage caused by 5 mM 1,2-dichlorovinyl 2-nitrophenyl disulfide was quantitatively comparable to the damage caused by 1.1 mM S -(2-chloroethyl)cysteine, an established DNA strand-breaking agent (). This result is in line with previous work by Vamvakas et al ( , ), who demonstrated DNA strand breaks in vitro and in vivo with chlorothioketene generated by β-lyase-catalyzed cleavage of S -1,2-(dichlorovinyl)- l -cysteine. Possible molecular mechanisms for the observed damage to DNA could be ester formation at the sugar/phosphate backbone (see accompanying manuscript), spontaneous depurination induced by DNA adduct formation, and cross-linking of the bifunctional chlorothioketene in analogy to chloroacetaldehyde, the rearrangement product of 2-chlorooxirane ().…”

Section: Discussionsupporting

confidence: 93%

Reactivity of Haloketenes and Halothioketenes with Nucleobases: Reactions in Vitro with DNA

Müller

Birner

Sander

et al. 1998

Chem. Res. Toxicol.

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Ketenes are important and highly reactive intermediates. Thioketenes are formed by cysteine conjugate beta-lyase-dependent biotransformation of 1-halovinylcysteine S-conjugates which are metabolites of several halogenated olefins. Nucleic acid constituents react with haloketenes and halothioketenes in vitro. Thioketenes induce DNA strand breaks in incubations of 1,2-dichlorovinyl 2-nitrophenyl disulfide, a thioketene precursor, with pBr 322 plasmid DNA. After treatment of single-stranded or native calf thymus DNA with chlorothioketene generated by the hydrolysis of 1,2-dichlorovinyl 2-nitrophenyl disulfide, the formation of 3,N4-thioacetylcytosine could be demonstrated. N6-(Dichloroacetyl)adenine and N4-(dichloroacetyl)cytosine, however, adducts formed by dichloroketene in vitro, are labile to hydrolysis. Therefore, the binding of this compound to DNA constituents in intact DNA is difficult to demonstrate. Substitution of one chlorine atom by fluorine allowed us to use 19F NMR as a tool to demonstrate the formation of adducts by dihaloketenes in intact DNA. N6-(Chlorofluoroacetyl)adenine and N4-(chlorofluoroacetyl)cytosine were synthesized (yields 77%, 15%, respectively) as references and characterized by LC/MS, 1H, 13C, and 19F NMR, FT-IR, and elemental analysis. To demonstrate the ability of dihaloketenes to bind to DNA, poly-dA (1 mg) and calf thymus DNA (10 mg) were suspended in DMF and treated with different concentrations of chlorofluoroketene (50-200 micromol). Analysis of the polymeric DNA by 19F NMR showed one doublet at -137.2 ppm downfield from the reference (CFCl3). A doublet at -146.9 ppm, characteristic for chlorofluoroacetic acid, an expected product of DNA adduct hydrolysis, was not detected. These results demonstrate the formation of a stable adenine adduct by dihaloketenes in intact calf thymus DNA.

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Section: Discussionsupporting

confidence: 93%

Reactivity of Haloketenes and Halothioketenes with Nucleobases: Reactions in Vitro with DNA

Müller

Birner

Sander

et al. 1998

Chem. Res. Toxicol.

Self Cite

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Interference of Co-Amplified Nuclear Mitochondrial DNA Sequences on the Determination of Human mtDNA Heteroplasmy by Using the SURVEYOR Nuclease and the WAVE HS System

Yen

Hsu

et al. 2014

PLoS ONE

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High-sensitivity and high-throughput mutation detection techniques are useful for screening the homoplasmy or heteroplasmy status of mitochondrial DNA (mtDNA), but might be susceptible to interference from nuclear mitochondrial DNA sequences (NUMTs) co-amplified during polymerase chain reaction (PCR). In this study, we first evaluated the platform of SURVEYOR Nuclease digestion of heteroduplexed DNA followed by the detection of cleaved DNA by using the WAVE HS System (SN/WAVE-HS) for detecting human mtDNA variants and found that its performance was slightly better than that of denaturing high-performance liquid chromatography (DHPLC). The potential interference from co-amplified NUMTs on screening mtDNA heteroplasmy when using these 2 highly sensitive techniques was further examined by using 2 published primer sets containing a total of 65 primer pairs, which were originally designed to be used with one of the 2 techniques. We confirmed that 24 primer pairs could amplify NUMTs by conducting bioinformatic analysis and PCR with the DNA from 143B-ρ0 cells. Using mtDNA extracted from the mitochondria of human 143B cells and a cybrid line with the nuclear background of 143B-ρ0 cells, we demonstrated that NUMTs could affect the patterns of chromatograms for cell DNA during SN-WAVE/HS analysis of mtDNA, leading to incorrect judgment of mtDNA homoplasmy or heteroplasmy status. However, we observed such interference only in 2 of 24 primer pairs selected, and did not observe such effects during DHPLC analysis. These results indicate that NUMTs can affect the screening of low-level mtDNA variants, but it might not be predicted by bioinformatic analysis or the amplification of DNA from 143B-ρ0 cells. Therefore, using purified mtDNA from cultured cells with proven purity to evaluate the effects of NUMTs from a primer pair on mtDNA detection by using PCR-based high-sensitivity methods prior to the use of a primer pair in real studies would be a more practical strategy.

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Covalent Binding of Reactive Intermediates of Xenobiotics to Mitochondrial DNA

Cited by 2 publications

References 21 publications

Reactivity of Haloketenes and Halothioketenes with Nucleobases: Reactions in Vitro with DNA

Reactivity of Haloketenes and Halothioketenes with Nucleobases: Reactions in Vitro with DNA

Interference of Co-Amplified Nuclear Mitochondrial DNA Sequences on the Determination of Human mtDNA Heteroplasmy by Using the SURVEYOR Nuclease and the WAVE HS System

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