The use of an artificial nucleotide for polymerase-based recognition of carcinogenic O6-alkylguanine DNA adducts
| Autor: | Wyss, Laura A., Nilforoushan, Arman, Williams, David M., Marx, Andreas, Sturla, Shana J. |
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| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: | |
| Zdroj: | Nucleic Acids Research, 44 (14) Nucleic Acids Research |
| Popis: | Enzymatic approaches for locating alkylation adducts at single-base resolution in DNA could enable new technologies for understanding carcinogenesis and supporting personalized chemotherapy. Artificial nucleotides that specifically pair with alkylated bases offer a possible strategy for recognition and amplification of adducted DNA, and adduct-templated incorporation of an artificial nucleotide has been demonstrated for a model DNA adduct O6-benzylguanine by a DNA polymerase. In this study, DNA adducts of biological relevance, O6-methylguanine (O6-MeG) and O6-carboxymethylguanine (O6-CMG), were characterized to be effective templates for the incorporation of benzimidazole-derived 2'-deoxynucleoside-5'-O-triphosphates ( BENZI: TP and BIM: TP) by an engineered KlenTaq DNA polymerase. The enzyme catalyzed specific incorporation of the artificial nucleotide BENZI: opposite adducts, with up to 150-fold higher catalytic efficiency for O6-MeG over guanine in the template. Furthermore, addition of artificial nucleotide BENZI: was required for full-length DNA synthesis during bypass of O6-CMG. Selective incorporation of the artificial nucleotide opposite an O6-alkylguanine DNA adduct was verified using a novel 2',3'-dideoxy derivative of BENZI: TP. The strategy was used to recognize adducts in the presence of excess unmodified DNA. The specific processing of BENZI: TP opposite biologically relevant O6-alkylguanine adducts is characterized herein as a basis for potential future DNA adduct sequencing technologies. published |
| Databáze: | OpenAIRE |
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