Dataset Information

Ring-opening of the ?-OH-PdG adduct promotes error-free bypass by the Sulfolobus solfataricus DNA polymerase Dpo4.

ABSTRACT: Acrolein, a mutagenic aldehyde, reacts with deoxyguanosine (dG) to form 3-(2'-deoxy-?-d-erythro-pentofuranosyl)-5,6,7,8-tetrahydro-8-hydroxypyrimido[1,2-a] purin-10(3H)-one (?-OH-PdG). When placed opposite deoxycytosine (dC) in DNA, ?-OH-PdG undergoes ring-opening to the N(2)-(3-oxopropyl)-dG. Ring-opening of the adduct has been hypothesized to facilitate nonmutagenic bypass, particularly by DNA polymerases of the Y family. This study examined the bypass of ?-OH-PdG by Sulfolobus solfataricus Dpo4, the prototypic Y-family DNA polymerase, using templates that contained the adduct in either the 5'-CXG-3' or the 5'-TXG-3' sequence context. Although ?-OH-PdG partially blocked Dpo4-catalyzed DNA synthesis, full primer extension was observed, and the majority of bypass products were error-free. Conversion of the adduct into an irreversibly ring-opened derivative prior to reaction facilitated bypass and further improved the fidelity. Structures of ternary Dpo4·DNA·dNTP complexes were determined with primers that either were positioned immediately upstream of the lesion (preinsertion complexes) or had a 3'-terminal dC opposite the lesion (postinsertion complexes); the incoming nucleotides, either dGTP or dATP, were complementary to the template 5'-neighbor nucleotide. In both postinsertion complexes, the adduct existed as ring-opened species, and the resulting base-pair featured Watson-Crick hydrogen bonding. The incoming nucleotide paired with the 5'-neighbor template, while the primer 3'-hydroxyl was positioned to facilitate extension. In contrast, ?-OH-PdG was in the ring-closed form in both preinsertion complexes, and the overall structure did not favor catalysis. These data provide insights into ?-OH-PdG chemistry during replication bypass by the Dpo4 DNA polymerase and may explain why ?-OH-PdG-induced mutations due to primer-template misalignment are uncommon.

SUBMITTER: Shanmugam G

PROVIDER: S-EPMC3775444 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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Ring-opening of the γ-OH-PdG adduct promotes error-free bypass by the Sulfolobus solfataricus DNA polymerase Dpo4.

Shanmugam Ganesh G Minko Irina G IG Banerjee Surajit S Christov Plamen P PP Kozekov Ivan D ID Rizzo Carmelo J CJ Lloyd R Stephen RS Egli Martin M Stone Michael P MP

Chemical research in toxicology 20130816 9

Acrolein, a mutagenic aldehyde, reacts with deoxyguanosine (dG) to form 3-(2'-deoxy-β-d-erythro-pentofuranosyl)-5,6,7,8-tetrahydro-8-hydroxypyrimido[1,2-a] purin-10(3H)-one (γ-OH-PdG). When placed opposite deoxycytosine (dC) in DNA, γ-OH-PdG undergoes ring-opening to the N(2)-(3-oxopropyl)-dG. Ring-opening of the adduct has been hypothesized to facilitate nonmutagenic bypass, particularly by DNA polymerases of the Y family. This study examined the bypass of γ-OH-PdG by Sulfolobus solfataricus Dp ...[more]

PMID: 23947567

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Project description:Aflatoxin B(1) (AFB(1)) is oxidized to an epoxide in vivo, which forms an N7-dG DNA adduct (AFB(1)-N7-dG). The AFB(1)-N7-dG can rearrange to a formamidopyrimidine (AFB(1)-FAPY) derivative. Both AFB(1)-N7-dG and the β-anomer of the AFB(1)-FAPY adduct yield G→T transversions in Escherichia coli, but the latter is more mutagenic. We show that the Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) bypasses AFB(1)-N7-dG in an error-free manner but conducts error-prone replication past the AFB(1)-FAPY adduct, including misinsertion of dATP, consistent with the G→T mutations observed in E. coli. Three ternary (Dpo4-DNA-dNTP) structures with AFB(1)-N7-dG adducted template:primers have been solved. These demonstrate insertion of dCTP opposite the AFB(1)-N7-dG adduct, and correct vs incorrect insertion of dATP vs dTTP opposite the 5'-template neighbor dT from a primed AFB(1)-N7-dG:dC pair. The insertion of dTTP reveals hydrogen bonding between the template N3 imino proton and the O(2) oxygen of dTTP, and between the template T O(4) oxygen and the N3 imino proton of dTTP, perhaps explaining why this polymerase does not efficiently catalyze phosphodiester bond formation from this mispair. The AFB(1)-N7-dG maintains the 5'-intercalation of the AFB(1) moiety observed in DNA. The bond between N7-dG and C8 of the AFB(1) moiety remains in plane with the alkylated guanine, creating a 16° inclination of the AFB(1) moiety with respect to the guanine. A binary (Dpo4-DNA) structure with an AFB(1)-FAPY adducted template:primer also maintains 5'-intercalation of the AFB(1) moiety. The β-deoxyribose anomer is observed. Rotation about the FAPY C5-N(5) bond orients the bond between N(5) and C8 of the AFB(1) moiety out of plane in the 5'-direction, with respect to the FAPY base. The formamide group extends in the 3'-direction. This improves stacking of the AFB(1) moiety above the 5'-face of the FAPY base, as compared to the AFB(1)-N7-dG adduct. Ternary structures with AFB(1)-β-FAPY adducted template:primers show correct vs incorrect insertion of dATP vs dTTP opposite the 5'-template neighbor dT from a primed AFB(1)-β-FAPY:dC pair. For dATP, the oxygen atom of the FAPY formamide group participates in a water-mediated hydrogen bond with Arg332. The insertion of dTTP yields a structure similar to that observed for the AFB(1)-N7-dG adduct. The differential accommodation of these AFB(1) adducts within the active site may, in part, modulate lesion bypass.

Dataset Information

Ring-opening of the ?-OH-PdG adduct promotes error-free bypass by the Sulfolobus solfataricus DNA polymerase Dpo4.

Publications

Ring-opening of the γ-OH-PdG adduct promotes error-free bypass by the Sulfolobus solfataricus DNA polymerase Dpo4.

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