Base Pairing Configuration and Stability of an Oligonucleotide Duplex Containing a 5-Chlorouracil-Adenine Base Pair

Abstract

Inflammation-mediated reactive molecules can damage DNA by oxidation and chlorination. The biological consequences of this damage are as yet incompletely understood. In this paper, we have constructed oligonucleotides containing 5-chlorouracil (ClU), one of the known inflammation damage products. The thermodynamic stability, base pairing configuration and duplex conformation of oligonucleotides containing ClU paired opposite adenine have been examined. NMR spectra reveal that the ClU-A base pair adopts a geometry similar to that of the T-A base pair, and the ClU-A containing duplex adopts a normal B-form conformation. The linewidth of the imino proton of the ClU residue is substantially greater than that of the corresponding T imino proton; however, this difference is not attributed to a reduced thermal or thermodynamic stability or to increased proton exchange with solvent. While the NMR studies reveal increased chemical exchange for the ClU imino proton of the ClU-A base pair, the ClU residue is not a target for removal by the Escherichia coli mispaired uracil glycosylase, which senses damage-related helix instability. The results of this study are consistent with previous reports indicating that the DNA of replicating cells can tolerate substantial substitution with CIU. The fraudulent, pseudo-Watson-Crick ClU-A base pair is sufficiently stable to avoid glycosylase removal and, therefore, might constitute a persistent form of cellular DNA damage.

Triplex- and Duplex-Forming Abilities of Oligonucleotides Containing 2′-Deoxy-5-trifluoromethyluridine and 2′-Deoxy-5-trifluoromethylcytidine

“…Analogous to m C, 5-bromocytosine ( Br C) stabilizes duplex formation when Br C-modified oligonucleotides are used. 5,8) Introduction of substituents at the C-5 position of uracil, such as a methyl group or halogen atoms, increases the stability of triplexes and duplexes formed 5,[9][10][11] (Fig. 1).…”

Chemical Evolution of a Bacterium’s Genome

“…[14] We chose chlorouracil (c) as the most promising candidate for DNAbase transliteration (see Figure S1b) because: 1) it closely resembles thymine in structural studies of synthetic DNA duplexes, which suggests that the stability of the A:c pair is close to that of A:T; [15] 2) it is readily converted into the chlorodeoxyuridine nucleoside and the dcMP, dcDP, and dcTP nucleotides by nucleoside phosphorylases, thymidine kinase, thymidylate kinase, and nucleoside diphosphate kinase, respectively; [2] 3) unlike fluorouracil, it is less liable to elevated tautomerism, which causes ambiguous pairing with guanine as well as adenine; [15] 4) unlike the bromo and iodo analogues, it is not reduced to uracil at the redox potential of the cytoplasm. [16] Chlorine is present in numerous natural compounds, especially secondary metabolites of marine organisms, and a number of enzymatic mechanisms that enable the formation of C À Cl bonds have been elucidated.…”

“…[30] Of these possibilities, formation of the anionic enol tautomer can be considered as a major factor that influences mispairing. The imino proton in chlorouracil (pK a = 7.9) is known to be more acidic than that in thymine (pK a = 9.7) but less acidic than in fluorouracil (pK a = 7.7), [15] which suggests that tautomerism Figure 3. Evolutionary kinetics of thymine replacement with chlorouracil in E. coli.…”

Comparison of the Structural and Dynamic Effects of 5-Methylcytosine and 5-Chlorocytosine in a CpG Dinucleotide Sequence

“…Thermodynamic parameters were obtained by fitting the experimental melting curves or from a plot of 1/ T m versus lnCt obtained at different oligonucleotide concentrations (Ct) as described previously (32, 33; Supporting Information Figure S2). The experimental values obtained in this study for the oligonucleotides shown in Figure 1 are listed in Table 1.…”