Novel post-synthetic generation, isomeric resolution, and characterization of Fapy-dG within oligodeoxynucleotides: Differential anomeric impacts on DNA duplex properties

Mark Lukin, Conceião A.S.A. Minetti, David P. Remeta, Sivaprasad Attaluri, Francis Johnson, Kenneth J. Breslauer, Carlos De Los Santos

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

Accumulation of damaged guanine nucleobases within genomic DNA, including the imidazole ring opened N 6-(2-Deoxy-α,β-D-erythro- pentafuranosyl)-2,6-diamino-4-hydroxy-5-formylamidopyrimidine (Fapy-dG), is associated with progression of age-related diseases and cancer. To evaluate the impact of this mutagenic lesion on DNA structure and energetics, we have developed a novel synthetic strategy to incorporate cognate Fapy-dG site-specifically within any oligodeoxynucleotide sequence. The scheme involves the synthesis of an oligonucleotide precursor containing a 5-nitropyrimidine moiety at the desired lesion site via standard solid-phase procedures. Following deprotection and isolation, the Fapy-dG lesion is generated by catalytic hydrogenation and subsequent formylation. NMR assignment of the Fapy-dG lesion (X) embedded within a TXT trimer reveals the presence of rotameric and anomeric species. The latter have been characterized by synthesizing the tridecamer oligodeoxynucleotide d(GCGTACXCATGCG) harboring Fapy-dG as the central residue and developing a protocol to resolve the isomeric components. Hybridization of the chromatographically isolated fractions with their complementary d(CGCATGCGTACGC) counterpart yields two Fapy-dGC duplexes that are differentially destabilized relative to the canonical GC parent. The resultant duplexes exhibit distinct thermal and thermodynamic profiles that are characteristic of α-and β-anomers, the former more destabilizing than the latter. These anomer-specific impacts are discussed in terms of differential repair enzyme recognition, processing and translesion synthesis.

Original languageEnglish (US)
Pages (from-to)5776-5789
Number of pages14
JournalNucleic acids research
Volume39
Issue number13
DOIs
StatePublished - Jul 2011

All Science Journal Classification (ASJC) codes

  • Genetics

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