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Mutagenic potential of DNA glycation: miscoding by (R)- and (S)-N2-(1-carboxyethyl)-2'-deoxyguanosine.

ABSTRACT: Elevated circulating glucose resulting from complications of obesity and metabolic disease can result in the accumulation of advanced glycation end products (AGEs) of proteins, lipids, and DNA. The formation of DNA-AGEs assumes particular importance as these adducts may contribute to genetic instability and elevated cancer risk associated with metabolic disease. The principal DNA-AGE, N(2)-(1-carboxyethyl)-2'-deoxyguanosine (CEdG), is formed as a mixture of R and S isomers at both the polymer and monomer levels. In order to examine the miscoding potential of this adduct, oligonucleotides substituted with (R)- and (S)-CEdG and the corresponding triphosphates (R)- and (S)-CEdGTP were synthesized, and base-pairing preferences for each stereoisomer were examined using steady-state kinetic approaches. Purine dNTPs were preferentially incorporated opposite template CEdG when either the Klenow (Kf(-)) or Thermus aquaticus (Taq) polymerases were used. The Kf(-) polymerase preferentially incorporated dGTP, whereas Taq demonstrated a bias for dATP. Kf(-) incorporated purines opposite the R isomer with greater efficiency, but Taq favored the S isomer. Incorporation of (R)- and (S)-CEdGTP only occurred opposite dC and was catalyzed by Kf(-) with equal efficiencies. Primer extension from a 3'-terminal CEdG was observed only for the R isomer. These data suggest CEdG is the likely adduct responsible for the observed pattern of G transversions induced by exposure to elevated glucose or its alpha-oxoaldehyde decomposition product methylglyoxal. The results imply that CEdG within template DNA and the corresponding triphosphate possess different syn/anti conformations during replication which influence base-pairing preferences. The implications for CEdG-induced mutagenesis in vivo are discussed.

SUBMITTER: Wuenschell GE

PROVIDER: S-EPMC3446853 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

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Mutagenic potential of DNA glycation: miscoding by (R)- and (S)-N2-(1-carboxyethyl)-2'-deoxyguanosine.

Wuenschell Gerald E GE Tamae Daniel D Cercillieux Angelique A Yamanaka Rio R Yu Calvin C Termini John J

Biochemistry 20100301 9

Elevated circulating glucose resulting from complications of obesity and metabolic disease can result in the accumulation of advanced glycation end products (AGEs) of proteins, lipids, and DNA. The formation of DNA-AGEs assumes particular importance as these adducts may contribute to genetic instability and elevated cancer risk associated with metabolic disease. The principal DNA-AGE, N(2)-(1-carboxyethyl)-2'-deoxyguanosine (CEdG), is formed as a mixture of R and S isomers at both the polymer an ...[more]

PMID: 20143879

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Project description:DNA interstrand cross-links (ICLs) are cytotoxic DNA lesions derived from reactions of DNA with a number of anti-cancer reagents as well as endogenous bifunctional electrophiles. Deciphering the DNA repair mechanisms of ICLs is important for understanding the toxicity of DNA cross-linking agents and for developing effective chemotherapies. Previous research has focused on ICLs cross-linked with the N7 and N2 atoms of guanine as well as those formed at the N6 atom of adenine; however, little is known about the mutagenicity of O6-dG-derived ICLs. Although less abundant, O6-alkylated guanine DNA lesions are chemically stable and highly mutagenic. Here, O6-2'-deoxyguanosine-butylene-O6-2'-deoxyguanosine (O6-dG-C4-O6-dG) is designed as a chemically stable ICL, which can be induced by the action of bifunctional alkylating agents. We investigate the DNA replication-blocking and mutagenic properties of O6-dG-C4-O6-dG ICLs during an important step in ICL repair, translesion DNA synthesis (TLS). The model replicative DNA polymerase (pol) Sulfolobus solfataricus P2 DNA polymerase B1 (Dpo1) is able to incorporate a correct nucleotide opposite the cross-linked template guanine of ICLs with low efficiency and fidelity but cannot extend beyond the ICLs. Translesion synthesis by human pol ? is completely inhibited by O6-dG-C4-O6-dG ICLs. Moderate bypass activities are observed for human pol ? and S. solfataricus P2 DNA polymerase IV (Dpo4). Among the pols tested, pol ? exhibits the highest bypass activity; however, 70% of the bypass products are mutagenic containing substitutions or deletions. The increase in the size of unhooked repair intermediates elevates the frequency of deletion mutation. Lastly, the importance of pol ? in O6-dG-derived ICL bypass is demonstrated using whole cell extracts of Xeroderma pigmentosum variant patient cells and those complemented with pol ?. Together, this study provides the first set of biochemical evidence for the mutagenicity of O6-dG-derived ICLs.

Dataset Information

Mutagenic potential of DNA glycation: miscoding by (R)- and (S)-N2-(1-carboxyethyl)-2'-deoxyguanosine.

Publications

Mutagenic potential of DNA glycation: miscoding by (R)- and (S)-N2-(1-carboxyethyl)-2'-deoxyguanosine.

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