DNA methylation by N-methyl-N-nitrosourea: methylation pattern changes in single- and double-stranded DNA, and in DNA with mismatched or bulged guanines

Abstract

The detection of abnormal DNA base pairing arrangements and conformations is chemically probed in synthetic 32P-end-labeled deoxyribonucleotide oligomers using N-methyl-N-nitrosourea (MNU) and 2,12,-dimethyl-3,7,11,17-tetraazabicyclo-[11.3.1]heptadeca-1 -[17],2,11,13,15 pentaene-Ni (II) (Ni-complex) with KHSO5. The DNA targets studied are single-stranded (s-s) DNA, double-stranded (d-s) DNA, d-s DNA with G-G, G-A and G-T mismatches, d-s DNA with a single bulged G and d-s DNA with two bulged G's. The effect of the non-Watson--Crick structures on the formation of N7-methylguanine (N7-MeG) by MNU and the oxidation of G by Ni-complex is reported along with the Tm's and circular dichroism spectra of the different duplex oligomers. The results for MNU and Ni-complex show that the qualitative and quantitative character of the cleavage patterns at a G3 run change with the nature of the abnormal base pairing motif. Based on the DNA substrates studied, the results indicate that a combination of reagents which report electronic and steric perturbations can be a useful approach to monitor DNA mismatches and bulges.

Treatment of Human Cells with N-Nitroso(acetoxymethyl)methylamine: Distribution Patterns of Piperidine-Sensitive DNA Damage at the Nucleotide Level of Resolution Are Related to the Sequence Context

“…We observed that DNA damage induced by MNU or NDMAOAc is sequence-specific in a run of three or more guanines in purified DNA and in cellular DNA ( Figure 5). Our results, and those realized by other investigators, confirm that the sequence specificities of S N 1 agents are different from those of S N 2 agents ( Figure 5) (12)(13)(14)(15)33). S N 1 methylating agents (MNU and NDMAOAc) preferentially methylate the central guanines in a run of three or more guanines.…”

“…The yield of methylation by MNU is actually enhanced in double-stranded DNA versus singlestranded DNA, indicating that steric factors are not significant in this process (33). Metha et al (36) showed that MNU alone, or coupled with a specific major groove intercalator agent, might be hydrolyzed in the major groove, and increased the extent of N7-guanine methylation as shown by the ratio of N7-methylguanine (a major groove lesion) to N3-methyladenine (a minor groove lesion).…”

A Review of the Role of the Sequence-Dependent Electrostatic Landscape in DNA Alkylation Patterns

“…Consistent with the requirement for a canonical double helix, a G that is mispaired with G, T or A becomes less reactive than a G paired with C ( Figure 5) (47). Similarly, the creation of G's in a single or double bulge show a decrease in alkylation ( Figure 5).…”

“…The pattern observed in ds-DNA is lost in ss-DNA, and most importantly, the alkylation yield was significantly reduced ( Figure 5) (47). In contrast, DMS, which shows no sequence selectivity with either ss-or ds-DNA, reacts more efficiently with the former.…”

Monitoring of dimethyl sulphate-induced N3-methyladenine, N7-methylguanine and O6-methylguanine DNA adducts using reversed-phase high performance liquid chromatography and mass spectrometry

“…In comparison to another S N 2-methylating agent MMS, the ratio for DMS is a slightly higher than was found for MMS (N7-/O 6 -MeG ratio = 200) [40]. The above ratio appear to depend on particular reacting conditions (aqueous solution, pH, duration incubation), but not on the double-or single-strandedness of DNA [41,42].…”