title: Polymerase Bypass of N(6)-Deoxyadenosine Adducts Derived from Epoxide Metabolites of 1,3-Butadiene.
abstract: N(6)-(2-Hydroxy-3-buten-1-yl)-2'-deoxyadenosine (N(6)-HB-dA I) and N(6),N(6)-(2,3-dihydroxybutan-1,4-diyl)-2'-deoxyadenosine (N(6),N(6)-DHB-dA) are exocyclic DNA adducts formed upon alkylation of the N(6) position of adenine in DNA by epoxide metabolites of 1,3-butadiene (BD), a common industrial and environmental chemical classified as a human and animal carcinogen. Since the N(6)-H atom of adenine is required for Watson-Crick hydrogen bonding with thymine, N(6)-alkylation can prevent adenine from normal pairing with thymine, potentially compromising the accuracy of DNA replication. To evaluate the ability of BD-derived N(6)-alkyladenine lesions to induce mutations, synthetic oligodeoxynucleotides containing site-specific (S)-N(6)-HB-dA I and (R,R)-N(6),N(6)-DHB-dA adducts were subjected to in vitro translesion synthesis in the presence of human DNA polymerases β, η, ι, and κ. While (S)-N(6)-HB-dA I was readily bypassed by all four enzymes, only polymerases η and κ were able to carry out DNA synthesis past (R,R)-N(6),N(6)-DHB-dA. Steady-state kinetic analyses indicated that all four DNA polymerases preferentially incorporated the correct base (T) opposite (S)-N(6)-HB-dA I. In contrast, hPol β was completely blocked by (R,R)-N(6),N(6)-DHB-dA, while hPol η and κ inserted A, G, C, or T opposite the adduct with similar frequency. HPLC-ESI-MS/MS analysis of primer extension products confirmed that while translesion synthesis past (S)-N(6)-HB-dA I was mostly error-free, replication of DNA containing (R,R)-N(6),N(6)-DHB-dA induced significant numbers of A, C, and G insertions and small deletions. These results indicate that singly substituted (S)-N(6)-HB-dA I lesions are not miscoding, but that exocyclic (R,R)-N(6),N(6)-DHB-dA adducts are strongly mispairing, probably due to their inability to form stable Watson-Crick pairs with dT.
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Srikanth
authors.initials: S
authors.lastname: Kotapati
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Susith
authors.initials: S
authors.lastname: Wickramaratne
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Amanda
authors.initials: A
authors.lastname: Esades
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Emily J
authors.initials: EJ
authors.lastname: Boldry
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Danae
authors.initials: D
authors.lastname: Quirk Dorr
authors.affiliation: ‡Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, United States.
authors.forename: Matthew G
authors.initials: MG
authors.lastname: Pence
authors.affiliation: ‡Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, United States.
authors.forename: F Peter
authors.initials: FP
authors.lastname: Guengerich
authors.affiliation: †Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, United States.
authors.forename: Natalia Y
authors.initials: NY
authors.lastname: Tretyakova
mesh_terms: D002070:Butadienes / Q000737:chemistry / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis / Q000378:metabolism; D018736:DNA Adducts / Q000737:chemistry / Q000378:metabolism*; D017931:DNA Primers / Q000378:metabolism; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003839:Deoxyadenosines / Q000737:chemistry / Q000378:metabolism*; D004852:Epoxy Compounds / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D002070:Butadienes; D018736:DNA Adducts; D017931:DNA Primers; D003839:Deoxyadenosines; D004852:Epoxy Compounds; C000598755:N6-(2-hydroxy-3-buten-1-yl)-2'-deoxyadenosine; D009838:Oligodeoxyribonucleotides; D004247:DNA; D004259:DNA-Directed DNA Polymerase; C031763:1,3-butadiene