Skip to Content
Merck
  • Structural and biochemical impact of C8-aryl-guanine adducts within the NarI recognition DNA sequence: influence of aryl ring size on targeted and semi-targeted mutagenicity.

Structural and biochemical impact of C8-aryl-guanine adducts within the NarI recognition DNA sequence: influence of aryl ring size on targeted and semi-targeted mutagenicity.

Nucleic acids research (2014-11-02)
Michael Sproviero, Anne M R Verwey, Katherine M Rankin, Aaron A Witham, Dmitriy V Soldatov, Richard A Manderville, Mostafa I Fekry, Shana J Sturla, Purshotam Sharma, Stacey D Wetmore
ABSTRACT

Chemical mutagens with an aromatic ring system may be enzymatically transformed to afford aryl radical species that preferentially react at the C8-site of 2'-deoxyguanosine (dG). The resulting carbon-linked C8-aryl-dG adduct possesses altered biophysical and genetic coding properties compared to the precursor nucleoside. Described herein are structural and in vitro mutagenicity studies of a series of fluorescent C8-aryl-dG analogues that differ in aryl ring size and are representative of authentic DNA adducts. These structural mimics have been inserted into a hotspot sequence for frameshift mutations, namely, the reiterated G3-position of the NarI sequence within 12mer (NarI(12)) and 22mer (NarI(22)) oligonucleotides. In the NarI(12) duplexes, the C8-aryl-dG adducts display a preference for adopting an anti-conformation opposite C, despite the strong syn preference of the free nucleoside. Using the NarI(22) sequence as a template for DNA synthesis in vitro, mutagenicity of the C8-aryl-dG adducts was assayed with representative high-fidelity replicative versus lesion bypass Y-family DNA polymerases, namely, Escherichia coli pol I Klenow fragment exo(-) (Kf(-)) and Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4). Our experiments provide a basis for a model involving a two-base slippage and subsequent realignment process to relate the miscoding properties of C-linked C8-aryl-dG adducts with their chemical structures.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Urea solution, BioUltra, ~8 M in H2O
Sigma-Aldrich
Urea-12C, 99.9 atom % 12C
Sigma-Aldrich
Formamide, ACS reagent, ≥99.5%
Sigma-Aldrich
Formamide, BioReagent, ≥99.5% (GC), for molecular biology
Sigma-Aldrich
L-Thyroxine sodium salt pentahydrate, ≥98% (HPLC), powder
Sigma-Aldrich
L-Thyroxine sodium salt pentahydrate, γ-irradiated, powder, BioXtra, suitable for cell culture
Sigma-Aldrich
Urea solution, 40 % (w/v) in H2O
Millipore
Urea solution, suitable for microbiology, 40% in H2O
Sigma-Aldrich
Formamide, JIS special grade, ≥98.5%
Sigma-Aldrich
Formamide, SAJ first grade, ≥98.5%
Sigma-Aldrich
Formamide, BioUltra, for molecular biology, ≥99.5% (T)
Sigma-Aldrich
Formamide, ReagentPlus®, ≥99.0% (GC)
Sigma-Aldrich
Formamide, spectrophotometric grade, ≥99%
Sigma-Aldrich
Urea, SAJ first grade, ≥98.0%
Sigma-Aldrich
Urea, JIS special grade, ≥99.0%
Sigma-Aldrich
Ethylenediaminetetraacetic acid, SAJ special grade, ≥99.0%
Sigma-Aldrich
o-Xylene, SAJ special grade, ≥98.5%
Supelco
Urea, 8 M (after reconstitution with 16 mL high purity water)
Supelco
o-Xylene, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Urea, ≥99.0%
Supelco
Formamide solution, NMR reference standard, 90% in DMSO-d6 (99.9 atom % D), NMR tube size 5 mm × 8 in.
Sigma-Aldrich
Ethylenediaminetetraacetic acid, 99.995% trace metals basis
Sigma-Aldrich
o-Xylene, anhydrous, 97%
Sigma-Aldrich
Ethylenediaminetetraacetic acid, BioUltra, ≥99.0% (KT)
Sigma-Aldrich
Urea, BioUltra, for molecular biology, 99% (T)
Sigma-Aldrich
Ethylenediaminetetraacetic acid, ≥98.0% (KT)
Sigma-Aldrich
Bromophenol Blue, ACS reagent
Sigma-Aldrich
Urea, puriss., meets analytical specification of Ph. Eur., BP, USP, 99.0-100.5%, 99.0-101.0% (calc. on dry substance)
Sigma-Aldrich
Ethylenediaminetetraacetic acid solution, 0.02% in DPBS (0.5 mM), sterile-filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Ethylenediaminetetraacetic acid, ACS reagent, 99.4-100.6%, powder