The Carbonate Radical Is a Site-selective Oxidizing Agent of Guanine in Double-stranded Oligonucleotides

Vladimir Shafirovich, Alexander Dourandin, Weidong Huang, Nicholas E. Geacintov

Research output: Contribution to journalArticle

Abstract

The carbonate radical anion (CO3 .-) is believed to be an important intermediate oxidant derived from the oxidation of bicarbonate anions and nitrosoperoxocarboxylate anions (formed in the reaction of CO 2 with ONOO-) in cellular environments. Employing nanosecond laser flash photolysis methods, we show that the CO3 .- anion can selectively oxidize guanines in the self-complementary oligonucleotide duplex d(AACGCGAATTCGCGTT) dissolved in air-equilibrated aqueous buffer solution (pH 7.5). In these time-resolved transient absorbance experiments, the CO3 .- radicals are generated by one-electron oxidation of the bicarbonate anions (HCO3 -) with sulfate radical anions (SO4 .-) that, in turn, are derived from the photodissociation of persulfate anions (S2O 8 2-) initiated by 308-nm XeCl excimer laser pulse excitation. The kinetics of the CO3 .- anion and neutral guanine radicals, G(-H)., arising from the rapid deprotonation of the guanine radical cation, are monitored via their transient absorption spectra (characteristic maxima at 600 and 315 nm, respectively) on time scales of microseconds to seconds. The bimolecular rate constant of oxidation of guanine in this oligonucleotide duplex by CO3 .- is (1.9 ± 0.2) x 107 M-1 S-1. The decay of the CO3 .- anions and the formation of G(-H). radicals are correlated with one another on the millisecond time scale, whereas the neutral guanine radicals decay on time scales of seconds. Alkali-labile guanine lesions are produced and are revealed by treatment of the irradiated oligonucleotides in hot piperidine solution. The DNA fragments thus formed are identified by a standard polyacrylamide gel electrophoresis assay, showing that strand cleavage occurs at the guanine sites only. The biological implications of these oxidative processes are discussed.

Original languageEnglish (US)
Pages (from-to)24621-24626
Number of pages6
JournalJournal of Biological Chemistry
Volume276
Issue number27
DOIs
StatePublished - Jul 6 2001

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Carbonates
Guanine
Oxidants
Oligonucleotides
Anions
Bicarbonates
Oxidation
Photodissociation
Excimer Lasers
Deprotonation
Photolysis
Alkalies
Excimer lasers
Carbon Monoxide
Electrophoresis
Cations
Polyacrylamide Gel Electrophoresis
Absorption spectra
Laser pulses
Rate constants

ASJC Scopus subject areas

  • Biochemistry

Cite this

The Carbonate Radical Is a Site-selective Oxidizing Agent of Guanine in Double-stranded Oligonucleotides. / Shafirovich, Vladimir; Dourandin, Alexander; Huang, Weidong; Geacintov, Nicholas E.

In: Journal of Biological Chemistry, Vol. 276, No. 27, 06.07.2001, p. 24621-24626.

Research output: Contribution to journalArticle

Shafirovich, Vladimir ; Dourandin, Alexander ; Huang, Weidong ; Geacintov, Nicholas E. / The Carbonate Radical Is a Site-selective Oxidizing Agent of Guanine in Double-stranded Oligonucleotides. In: Journal of Biological Chemistry. 2001 ; Vol. 276, No. 27. pp. 24621-24626.
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