Dnmt3a-CD Is Less Susceptible to Bulky Benzo[a]pyrene Diol Epoxide-Derived DNA Lesions Than Prokaryotic DNA Methyltransferases

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• 作者：Olga V. Lukashevich ; Vladimir B. Baskunov ; Maria V. Darii ; Alexander Kolbanovskiy ; Alexander A. Baykov ; Elizaveta S. Gromova
• 刊名：Biochemistry
• 出版年：2011
• 出版时间：February 8, 2011
• 年：2011
• 卷：50
• 期：5
• 页码：875-881
• 全文大小：800K
• 年卷期：v.50,no.5(February 8, 2011)
• ISSN：1520-4995

文摘

Benzo[a]pyrene (B[a]P) is a well-characterized environmental polycyclic aromatic hydrocarbon pollutant. In living organisms, B[a]P is metabolized to the genotoxic anti-benzo[a]pyrene diol epoxide that reacts with cellular DNA to form stereoisomeric anti-B[a]PDE-N²-dG adducts. In this study, we explored the effects of adduct stereochemistry and position in double-stranded DNA substrates on the functional characteristics of the catalytic domain of murine de novo DNA methyltransferase Dnmt3a (Dnmt3a-CD). A number of 18-mer duplexes containing site-specifically incorporated (+)- and (-)-trans-anti-B[a]PDE-N²-dG lesions located 3鈥? and 5鈥?adjacent to and opposite the target cytosine residue were prepared. Dnmt3a-CD binds cooperatively to the DNA duplexes with an up to 5-fold greater affinity compared to that for the undamaged DNA duplexes. Methylation assays showed a 1.7鈭?.3-fold decrease in the methylation reaction rates for the damaged duplexes. B[a]PDE modifications stimulated a nonproductive binding and markedly favored substrate inhibition of Dnmt3a-CD in a manner independent of DNA methylation status. The latter effect was sensitive to the position and stereochemistry of the B[a]PDE-N²-dG adducts. The overall effect of trans-anti-B[a]PDE-N²-dG adducts on Dnmt3a-CD was less detrimental than in the case of the prokaryotic methyltransferases we previously investigated.