Reactivity of the C2'-oxidized abasic lesion and its relevance to interactions with type I base excision repair enzymes.
ABSTRACT The C2'-oxidized abasic lesion (C2-AP) is produced in DNA that is subjected to oxidative stress. C2-AP is incised by phosphodiesterases, but is not a substrate for endonuclease III even though a Schiff base is formed (Greenberg, M. M., et al. (2004) Biochemistry 43, 15217). A chemically synthesized oligonucleotide was used to study C2-AP reactivity under alkaline conditions and with nitrogen nucleophiles chosen to mimic the lysine or N-terminal proline side chains present in the active site of Type I base excision repair enzymes. Alkaline cleavage of the C2-AP lesion produces 3'-phosphoglycoaldehyde and 3'-phosphate termini. The former is degraded further to 3'-hydroxyl groups. Cleavage at the C2-AP lesion is enhanced by small peptides, which form Schiff base intermediates with the lesion. C2-AP cleavage by Lys.Trp.Lys and Lys.Trp.Gly.Lys suggests that the inability of endonuclease III to cleave the lesion is due to the absence of appropriately positioned functional groups to take advantage of formation of the covalent intermediate. These observations leave open the possibility that the C2-AP lesion may be a substrate for other Type I repair enzymes.
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ABSTRACT: The oxidation of 2-deoxyribose in DNA has emerged as a critical determinant of the cellular toxicity of oxidative damage to DNA, with oxidation of each carbon producing a unique spectrum of electrophilic products. We have developed and validated an isotope-dilution gas chromatography-coupled mass spectrometry (GC-MS) method for the rigorous quantification of two major 2-deoxyribose oxidation products: the 2-deoxyribonolactone abasic site of 1'-oxidation and the nucleoside 5'-aldehyde of 5'-oxidation chemistry. The method entails elimination of these products as 5-methylene-2(5H)-furanone (5MF) and furfural, respectively, followed by derivatization with pentafluorophenylhydrazine (PFPH), addition of isotopically labeled PFPH derivatives as internal standards, extraction of the derivatives, and quantification by GC-MS analysis. The precision and accuracy of the method were validated with oligodeoxynucleotides containing the 2-deoxyribonolactone and nucleoside 5'-aldehyde lesions. Further, the well-defined 2-deoxyribose oxidation chemistry of the enediyne antibiotics, neocarzinostatin and calicheamicin gamma(1)(I), was exploited in control studies, with neocarzinostatin producing 10 2-deoxyribonolactone and 300 nucleoside 5'-aldehyde per 10(6) nt per microM in accord with its established minor 1'- and major 5'-oxidation chemistry. Calicheamicin unexpectedly caused 1'-oxidation at a low level of 10 2-deoxyribonolactone per 10(6) nt per microM in addition to the expected predominance of 5'-oxidation at 560 nucleoside 5'-aldehyde per 10(6) nt per microM. The two hydroxyl radical-mediated DNA oxidants, gamma-radiation and Fe(2+)-EDTA, produced nucleoside 5'-aldehyde at a frequency of 57 per 10(6) nt per Gy (G-value 74 nmol/J) and 3.5 per 10(6) nt per microM, respectively, which amounted to 40% and 35%, respectively, of total 2-deoxyribose oxidation as measured by a plasmid nicking assay. However, gamma-radiation and Fe(2+)-EDTA produced different proportions of 2-deoxyribonolactone at 7% and 24% of total 2-deoxyribose oxidation, respectively, with frequencies of 10 lesions per 10(6) nt per Gy (G-value, 13 nmol/J) and 2.4 lesions per 10(6) nt per microM. Studies in TK6 human lymphoblastoid cells, in which the analytical data were corrected for losses sustained during DNA isolation, revealed background levels of 2-deoxyribonolactone and nucleoside 5'-aldehyde of 9.7 and 73 lesions per 10(6) nt, respectively. Gamma-irradiation of the cells caused increases of 0.045 and 0.22 lesions per 10(6) nt per Gy, respectively, which represents a approximately 250-fold quenching effect of the cellular environment similar to that observed in previous studies. The proportions of the various 2-deoxyribose oxidation products generated by gamma-radiation are similar for purified DNA and cells. These results are consistent with solvent exposure as a major determinant of hydroxyl radical reactivity with 2-deoxyribose in DNA, but the large differences between gamma-radiation and Fe(2+)-EDTA suggest that factors other than hydroxyl radical reactivity govern DNA oxidation chemistry.Journal of the American Chemical Society 04/2010; 132(17):6145-53. · 11.44 Impact Factor
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ABSTRACT: Condensation of dialdehyde 1 with different aromatic amines gave a series of symmetrical diarylethenes containing Schiff-base moieties, which were reduced with NaBH4 to give the corresponding diarylethene amines. The structures of all new compounds were confirmed by NMR spectroscopy, mass spectrometry, X-ray diffraction and elemental analysis. Their photochromic properties were also investigated, revealing that diarylethene imines displayed better photochromic properties in solution than their amine analogues. Furthermore, we found that by varying the aromatic amine precursor, diarylethenes with different polychromic behavior could be obtained. A number of different aromatic amines were investigated, including different carbon skeletons and substitution with different halogens in various positions, to tune the polychromic behavior.Journal of Photochemistry and Photobiology A Chemistry 02/2011; 218(2):192-198. · 2.29 Impact Factor
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ABSTRACT: The spectral, angular, polarization, air and water temperatures, sea surface waves and wind features of reflected from the sea surface radar signal frequency Doppler shift distribution variance, sea surface radar cross section and brightness temperature are considered. The method and device of simultaneous Doppler-radar and cross-polarized radiometer sensing for sea surface roughness degree, wind speed, water and air temperatures determination are described01/1996;