Synthesis of stable-isotope enriched 5-methylpyrimidines and their use as probes of base reactivity in DNA

Artur Burdzy, Katherine T. Noyes, Victoria Valinluck, Lawrence Sowers

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

A specific and efficient method is presented for the conversion of 2′-deoxyuridine to thymidine via formation and reduction of the intermediate 5-hydroxy-methyl derivative. The method has been used to generate both thymidine and 5-methyl-2′-deoxycytidine containing the stable isotopes 2H, 13C and 15N. Oligodeoxyribonucleotides have been constructed with these mass-tagged bases to investigate sequence-selectivity in hydroxyl radical reactions of pyrimidine methyl groups monitored by mass spectrometry. Studying the reactivity of 5-methyl-cytosine (5mC) is difficult as the reaction products can deaminate to the corresponding thymine derivatives, making the origin of the reaction products ambiguous. The method reported here can distinguish products derived from 5mC and thymine as well as investigate differences in reactivity for either base in different sequence contexts. The efficiency of formation of 5-hydroxymethyluracil from thymine is observed to be similar in magnitude in two different sequence contexts and when present in a mispair with guanine. The oxidation of 5mC proceeds slightly more efficiently than that of thymine and generates both 5-hydroxymethylcytosine and 5-formylcytosine but not the deaminated products. Thymine glycol is generated by both thymine and 5mC, although with reduced efficiency for 5mC. The method presented here should be widely applicable, enabling the examination of the reactivity of selected bases in DNA.

Original languageEnglish (US)
Pages (from-to)4068-4074
Number of pages7
JournalNucleic Acids Research
Volume30
Issue number18
StatePublished - Sep 15 2002
Externally publishedYes

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Thymine
Cytosine
Isotopes
DNA
5-methyldeoxycytidine
Thymidine
Deoxyuridine
Oligodeoxyribonucleotides
Guanine
Hydroxyl Radical
Mass Spectrometry

ASJC Scopus subject areas

  • Genetics

Cite this

Synthesis of stable-isotope enriched 5-methylpyrimidines and their use as probes of base reactivity in DNA. / Burdzy, Artur; Noyes, Katherine T.; Valinluck, Victoria; Sowers, Lawrence.

In: Nucleic Acids Research, Vol. 30, No. 18, 15.09.2002, p. 4068-4074.

Research output: Contribution to journalArticle

Burdzy, Artur ; Noyes, Katherine T. ; Valinluck, Victoria ; Sowers, Lawrence. / Synthesis of stable-isotope enriched 5-methylpyrimidines and their use as probes of base reactivity in DNA. In: Nucleic Acids Research. 2002 ; Vol. 30, No. 18. pp. 4068-4074.
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