The Substrate Specificity of MutY for Hyperoxidized Guanine Lesions in Vivo

Abstract

The DNA damage product 7,8-dihydro-8-oxo-2'-deoxyguanine (8-oxoG) is a commonly used biomarker of oxidative stress. The mutagenic potential of this DNA lesion is mitigated in Escherichia coli by multiple enzymes. One of these enzymes, MutY, excises an A mispaired with 8-oxoG as part of the process to restore the original G:C base pair. However, numerous studies have shown that 8-oxoG is chemically labile toward further oxidation. Here, we examine the activity of MutY on the 8-oxoG oxidation products guanidinohydantoin (Gh), two diastereomers of spiroiminodihydantoin (Sp1 and Sp2), oxaluric acid (Oa), and urea (Ur). Single-stranded viral genomes containing a site-specific lesion were constructed and replicated in E. coli that are either proficient in DNA repair or that lack MutY. These lesions were found previously to be potently mutagenic in repair competent bacteria, and we report here that these 8-oxoG-derived lesions are equally miscoding when replicated in E. coli lacking MutY; no significant change in mutation identity or frequency is observed. Interestingly, however, in the presence of MutY, Sp1 and Sp2 are more toxic than in cells lacking this repair enzyme.

Intrinsic mutagenic properties of 5-chlorocytosine: A mechanistic connection between chronic inflammation and cancer

“…This result therefore suggests that 5ClC base-paired with guanine most of the time (∼95%), but it also base-paired with adenine (∼5%), an unexpected result. To put this value in perspective, 5ClC is approximately as mutagenic as 8oxoG, evaluated with a similar assay in a wild-type E. coli strain (i.e., a strain with normal levels of the 8oxoG repair enzymes MutM and MutY) (30,31). The positive control m3C behaved as previously reported (27,29).…”

Postgenomic Analysis of Streptococcus thermophilus Cocultivated in Milk with Lactobacillus delbrueckii subsp. bulgaricus : Involvement of Nitrogen, Purine, and Iron Metabolism

“…bulgaricus. Beyond the upregulation of dpr, the upregulation of htrA (serine protease) and htpX (heat shock protein) could be related to stress, as is the upregulation of the mutS, mutY, and ung genes, coding for DNA glycosylases involved in DNA repair relative to oxidative stress damages (18,66). However, we observed a downregulation of Gor, the glutathione reductase, and radA, which belong to the oxidative stress response.…”

Superior Removal of Hydantoin Lesions Relative to Other Oxidized Bases by the Human DNA Glycosylase hNEIL1

“…Single nucleotide insertion and primer extension experiments using Escherichia coli (E. coli) Klenow fragment exonuclease I (KF exo − ) DNA polymerase indicate that dAMP and dGMP are inserted opposite these oxidized lesions (25,26). Moreover, in E. coli based mutagenesis assays with singlestranded lesion-containing viral DNA, Gh and Sp are potently mutagenic causing both G→ T and G→ C transversion mutations (27,28).…”

“…Single nucleotide insertion and primer extension experiments using Escherichia coli (E. coli) Klenow fragment exonuclease I (KF exo − ) DNA polymerase indicate that dAMP and dGMP are inserted opposite these oxidized lesions (25,26). Moreover, in E. coli based mutagenesis assays with singlestranded lesion-containing viral DNA, Gh and Sp are potently mutagenic causing both G→ T and G→ C transversion mutations (27,28).Oxidative damage to individual DNA bases may be repaired via the base excision repair (BER) pathway that is initiated by damage-specific glycosylases that excise damaged DNA bases (7). Other enzymes in the BER pathway remove the remaining sugar fragment and incorporate an undamaged nucleotide at the site of the original damaged base.…”