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Maddukuri L.,A B Hancock Jr Memorial Laboratory For Cancer Research | Eoff R.L.,A B Hancock Jr Memorial Laboratory For Cancer Research | Choi J.-Y.,A B Hancock Jr Memorial Laboratory For Cancer Research | Choi J.-Y.,Ewha Womans University | And 5 more authors.
Biochemistry | Year: 2010

3-(2′-Deoxy-β-d-erythro-pentofuranosyl)pyrimido-[1,2-a] purin-10(3H)-one (M1dG) is the major adduct derived from the reaction of DNA with the lipid peroxidation product malondialdehyde and the DNA peroxidation product base propenal. M1dG is mutagenic in Escherichia coli and mammalian cells, inducing base-pair substitutions (M1dG → A and M1dG → T) and frameshift mutations. Y-family polymerases may contribute to the mutations induced by M1dG in vivo. Previous reports described the bypass of M1dG by DNA polymerases η and Dpo4. The present experiments were conducted to evaluate bypass of M1dG by the human Y-family DNA polymerases κ, τ, and Rev1. M1dG was incorporated into template-primers containing either dC or dT residues 5′ to the adduct, and the template-primers were subjected to in vitro replication by the individual DNA polymerases. Steady-state kinetic analysis of single nucleotide incorporation indicates that dCMP is most frequently inserted by hPol κ opposite the adduct in both sequence contexts, followed by dTMP and dGMP. dCMP and dTMP were most frequently inserted by hPol τ, and only dCMP was inserted by Rev1. hPol κ extended template-primers in the order M1dG:dC > M1dG:dG > M1dG:dT ∼ M1dG:dA, but neither hPol τ nor Rev1 extended M1dG-containing template-primers. Liquid chromatography-mass spectrometry analysis of the products of hPol κ-catalyzed extension verified this preference in the 3′-GXC-5′ template sequence but revealed the generation of a series of complex products in which dAMP is incorporated opposite M1dG in the 3′-GXT-5′ template sequence. The results indicate that DNA hPol κ or the combined action of hPol τ or Rev1 and hPol κ bypass M1dG residues in DNA and generate products that are consistent with some of the mutations induced by M1dG in mammalian cells. © 2010 American Chemical Society.

Galligan J.J.,Vanderbilt University | Rose K.L.,Vanderbilt University | Beavers W.N.,A B Hancock Jr Memorial Laboratory For Cancer Research | Hill S.,Vanderbilt University | And 4 more authors.
Journal of the American Chemical Society | Year: 2014

Lipid electrophiles modify cellular targets, altering their function. Here, we describe histones as major targets for modification by 4-oxo-2-nonenal, resulting in a stable Lys modification structurally analogous to other histone Lys acylations. Seven adducts were identified in chromatin isolated from intact cells: four 4-ketoamides to Lys and three Michael adducts to His. A 4-ketoamide adduct residing at H3K27 was identified in stimulated macrophages. Modification of histones H3 and H4 prevented nucleosome assembly. © 2014 American Chemical Society.

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