Methylation of 2′-deoxyguanosine by a free radical mechanism

Conor Crean, Nicholas Geacintov, Vladimir Shafirovich

Research output: Contribution to journalArticle

Abstract

The mechanistic aspects of the methylation of guanine in DNA initiated by methyl radicals that are derived from the metabolic oxidation of some chemical carcinogens remain poorly understood. In this work, we investigated the kinetics and the formation of methylated guanine products by two methods: (i) the combination of CH3 radicals and guanine neutral radicals, G(-H), and (ii) the direct addition of CH3 radicals to guanine bases. The simultaneous generation of CH3 and dG(-H) radicals was triggered by the competitive oneelectron oxidation of dimethyl sulfoxide (DMSO) and 2'-deoxyguanosine (dG) by photochemically generated sulfate radicals in deoxygenated aqueous buffer solutions (pH 7.5). The photolysis of methylcob(III)alamin to form CH3 radicals was used to investigate the direct addition of these radicals to guanine bases. The major end products of the radical combination reactions are the 8-methyl-dG and N2-methyl-dG products formed in a ratio of 1:0.7. In contrast, the methylation of dG by CH3 radicals generates mostly the 8-methyl-dG adduct and only minor quantities of N2-methyl-dG (1:0.13 ratio). The methylation of the self-complementary 5'd(AACGCGAATTCGCGTT) duplexes was achieved by the selective oxidation of the guanines with carbonate radical anions in the presence of DMSO as the precursor of CH3 radicals. The methyl-G lesions formed were excised by the enzymatic digestion and identified by LC-MS/MS methods using uniformly 15N-labeled 8-methyldG and N2-methyl-dG adducts as internal standards. The ratios of S-methyl-G/N2-methyl-G lesions derived from the combination of methyl radicals with G(-H) radicals positioned in double-stranded DNA or that with the free nucleoside dG(-H) radicals were found to be similar. Utilizing the photochemical method and dipropyl or dibutyl sulfoxides as sources of alkyl radicals, the corresponding 8-alkyl-dG and N2-alkyl-dG adducts were also generated in ratios similar to those obtained with DMSO.

Original languageEnglish (US)
Pages (from-to)12773-12781
Number of pages9
JournalJournal of Physical Chemistry B
Volume113
Issue number38
DOIs
StatePublished - Sep 24 2009

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Deoxyguanosine
methylation
Methylation
Dimethyl sulfoxide
Free radicals
free radicals
Free Radicals
Oxidation
Guanine
DNA
guanines
Carcinogens
Photolysis
Carbonates
Dimethyl Sulfoxide
Negative ions
Kinetics
adducts
Sulfoxides
lesions

ASJC Scopus subject areas

  • Physical and Theoretical Chemistry
  • Materials Chemistry
  • Surfaces, Coatings and Films

Cite this

Methylation of 2′-deoxyguanosine by a free radical mechanism. / Crean, Conor; Geacintov, Nicholas; Shafirovich, Vladimir.

In: Journal of Physical Chemistry B, Vol. 113, No. 38, 24.09.2009, p. 12773-12781.

Research output: Contribution to journalArticle

Crean, Conor ; Geacintov, Nicholas ; Shafirovich, Vladimir. / Methylation of 2′-deoxyguanosine by a free radical mechanism. In: Journal of Physical Chemistry B. 2009 ; Vol. 113, No. 38. pp. 12773-12781.
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