Publications (69) View all
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Article: Glutathione Conjugates of Ochratoxin a as Biomarkers of Exposure / Glutationski Konjugati Okratoksina A Kao Biomarkeri Izloženosti.
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ABSTRACT: Abstract In the present study the photoreactivity of the fungal carcinogen ochratoxin A (OTA) has been utilised to generate authentic samples of reduced glutathione (GSH) and N-acetylcysteine (NAC) conjugates of the parent toxin. These conjugates, along with the nontoxic OTα, which is generated through hydrolysis of the amide bond of OTA by carboxypeptidase A, were utilised as biomarkers to study the metabolism of OTA in the liver and kidney of male and female Dark Agouti rats. Male rats are more susceptible than female rats to OTA carcinogenesis with the kidney being the target organ. Our studies show that the distribution of OTA in male and female rat kidney is not significantly different. However, the extent of OTA metabolism was greater in male than female rats. Much higher levels of OTα were detected in the liver compared to the kidney, and formation of OTα is a detoxification pathway for OTA. These findings suggest that differences in metabolism between male and female rats could provide an explanation for the higher sensitivity of male rats to OTA toxicity.Archives of Industrial Hygiene and Toxicology 12/2012; 63(4):417-427. · 1.05 Impact Factor -
Article: Fluorescent C-linked C8-aryl-guanine probe for distinguishing syn from anti structures in duplex DNA.
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ABSTRACT: The synthesis and optical properties of the carbon (C)-linked C(8)-(2"-benzo[b]thienyl)-2'-deoxyguanosine ((Bth)dG), which acts as a fluorescent reporter of syn versus anti glycosidic conformations in duplex DNA, are described. In the syn-conformation, the probe stabilizes a G:G mismatch, emits at ∼385 nm (excitation ∼285 nm), and shows an induced circular dichroism (ICD) signal at ∼320 nm. Molecular dynamics (MD) simulations predict a wedge (W)-conformation for the mismatched duplex with the C(8)-benzo[b]thienyl moiety residing in the minor groove. In contrast, the probe destabilizes the duplex when base paired with its normal pyrimidine partner C. With flanking purine bases, a major groove B-type duplex is favored with (Bth)dG present in the anti-conformation emitting at ∼413 nm (excitation ∼326 nm) and no ICD signal. However, with flanking pyrimidine bases, (Bth)dG adopts the syn-conformation when base paired with C, and MD simulations predict a base-displaced stacked (S)-conformation, with the opposing C flipped out of the helix. The different duplex (B-, S-, and W-) conformers formed upon incorporation of (Bth)dG are known to play a critical role in the biological activity of N-linked C8-dG adducts formed by arylamine carcinogens. Bulky environment-sensitive fluorescent C(8)-dG adducts that mimic the duplex structures formed by carcinogens may be useful in luminescence-based DNA polymerase assays.Chemical Research in Toxicology 06/2012; 25(6):1271-82. · 3.78 Impact Factor -
Article: Application of a fluorescent C-linked phenolic purine adduct for selective N7-metalation of DNA.
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ABSTRACT: The C-linked phenolic adduct, C8-(2″-hydroxyphenyl)-2'-deoxyguanosine (o-PhOHdG), has been employed to study the impact of N7-metalation of 2'-deoxyguanosine (dG) within duplex DNA. The phenolic group of o-PhOHdG assists selective metal ion coordination by the N7-site of the attached dG moiety, which is the most important metal binding site in duplex DNA. The biaryl nucleobase probe o-PhOHdG is highly fluorescent in water (Φ(fl) = 0.44), and changes in its absorption and emission were used to determine apparent association constants (K(a)) for binding to Cu(II), Ni(II), and Zn(II). The nucleoside was found to bind Cu(II) (log K(a) = 4.59) and Ni(II) (log K(a) = 3.65) effectively, but it showed relatively poor affinity for Zn(II) (log K(a) = 2.55). The fluorescent nucleobase o-PhOHdG was incorporated into a pyrimidine-rich oligonucleotide substrate (ODN1) and a purine-rich (ODN2) substrate to monitor selective binding of Cu(II) through fluorescence quenching of the enol emission of o-PhOHdG within the DNA substrates. The pyrimidine-rich substrate ODN1 was found to possess greater affinity for Cu(II) than the free nucleobase, while the purine-rich substrate ODN2 exhibited diminished Cu(II) binding affinity. The impact of Cu(II) on duplex stability and structure was determined using UV melting temperature analysis and circular dichroism (CD) measurements. These studies highlight the syn preference for Cu(II)-bound o-PhOHdG within ODN1 duplexes and demonstrate competitive Cu(II) binding by other natural dG nucleobases within ODN2. The metal binding properties of o-PhOHdG are compared to the structurally similar 2-(2'-hydroxyphenyl)benzoxazole (HBO) derivatives and the nucleoside C8-(2-pyridyl)-dG (2PydG) that has also been used to control N7-metal coordination in DNA. Our results show certain advantages to the use of o-PhOHdG that stem from its highly fluorescent nature in aqueous media and provide additional tools for studying the effects of N7-metalation on the structure and stability of duplex DNA.The Journal of Physical Chemistry B 05/2012; 116(21):6158-65. · 3.70 Impact Factor -
Article: An indole-linked C8-deoxyguanosine nucleoside acts as a fluorescent reporter of Watson-Crick versus Hoogsteen base pairing.
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ABSTRACT: Pyrrole- and indole-linked C(8)-deoxyguanosine nucleosides act as fluorescent reporters of H-bonding specificity. Their fluorescence is quenched upon Watson-Crick H-bonding to dC, while Hoogsteen H-bonding to G enhances emission intensity. The indole-linked probe is ∼ 10-fold brighter and shows promise as a fluorescent reporter of Hoogsteen base pairing.Organic & Biomolecular Chemistry 03/2011; 9(5):1565-71. · 3.70 Impact Factor -
Article: Tautomerization in gas-phase ion chemistry of isomeric C-8 deoxyguanosine adducts from phenol-induced DNA damage.
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ABSTRACT: Collision-induced dissociation (CID) of 8-(4''-hydroxyphenyl)-2'-deoxyguanosine and 8-(2''-hydroxyphenyl)-2'-deoxyguanosine was investigated using sequential tandem mass spectrometry. These adducts represent biomarkers of DNA damage linked to phenolic radicals and were investigated to gain insight into the effects of chemical structure of a C-8 modification on fragmentation pathways of modified 2'-deoxyguanosine (dG). CID in MS(2) of the deprotonated molecules of both the isomers generated the same product ion having the same m/z values. CID in MS(3) of the product ion at m/z 242 and CID in MS(4) experiments carried out on the selected product ions at m/z 225 and m/z 218 afford distinct fragmentation patterns. The conformational properties of isomeric product ions from CID showed that the ortho-isomers possess the unique ability to tautomerize through an intramolecular proton transfer between the phenolic OH group and the imine nitrogen (N7). Tautomerization of ortho-isomers to their keto-tautomers led to differences in their system of conjugated double bonds compared with either their enol-tautomer or the para-isomer. The charge redistribution through the N-7 site on the imidazole ring is a critical step in guanosine adduct fragmentation which is disrupted by the formation of the keto-tautomer. For this reason, different reaction pathways are observed for 8-(4''-hydroxyphenyl)-2'-deoxyguanosine and 8-(2''-hydroxyphenyl)-2'-deoxyguanosine. We present herein the dissociation and the gas-phase ion-molecule reactions for highly conjugated ions involved in the CID ion chemistry of the investigated adducts. These will be useful for those using tandem mass spectrometry for structural elucidation of C-8 modified dG adducts. This study demonstrates that the modification at the C-8 site of dG has the potential to significantly alter the reactivity of adducts. We also show the ability of tandem mass spectrometry to completely differentiate between the isomeric dG adducts investigated.Biological Mass Spectrometry 01/2011; 46(1):41-9. · 3.41 Impact Factor
