Reaction of guanyl radicals in plasmid DNA with biological reductants: chemical repair of DNA damage produced by the direct effect of ionizing radiation.
OBJECTIVE
It has been previously argued that the use of the one-electron oxidants (SCN)2(*-) and Br2(*-) with plasmid DNA leads to the formation of DNA guanyl radicals. These guanyl radical species are intermediates in the DNA damage produced by processes such as photo-ionization and ionizing irradiation. The present paper evaluates the use of thallium(II) ions (Tl(II)OH(+)) as the one-electron oxidant, and also determines rate constants for the reduction (repair) of guanyl radicals in plasmid DNA by a variety of reducing agents including the biologically important compounds ascorbate and glutathione.


METHODS
Aqueous solutions of plasmid DNA containing 10(-3) mol dm(-3) thiocyanate or thallous ions and a reducing agent (azide, nitrite, ferrocyanide, hexachloroiridate(III), iodide, ascorbate, glutathione, glutathione disulphide, methionine, tyrosine, 5-hydroxyindole-3-acetic acid, 10(-7)-10(-4) mol dm(-3)) were irradiated with 137Cs gamma-rays (662 keV). After irradiation, the plasmid was incubated with the E. coli base excision repair endonuclease formamidopyrimidine-DNA N-glycosylase (FPG). Strand break yields after incubation were quantified by means of agarose gel electrophoresis.


RESULTS
High yields of FPG-sensitive sites produced by the oxidants (SCN)2(*-) and Tl(II)OH(+) were strongly attenuated by the presence of the reducing agents.


CONCLUSIONS
From the results, it is possible to arrive at estimates of the rate constants for the reduction of the DNA guanyl radical by the reducing agents. Values lie in the range 10(4)-10(7) dm(3) mol(-1) s(-1). Using the values for ascorbate and glutathione, it is possible to estimate an upper limit on the order of milliseconds for the lifetime of DNA guanyl radicals under cellular conditions. The implication is that there may well be a significant chemical repair of DNA base damage by the direct effect of ionizing radiation.
Milligan
J R
JR
Department of Radiology, University of California at San Diego, La Jolla, CA 92093-0610, USA. jmilligan@ucsd.edu
Aguilera
J A
JA
Mares
E J
EJ
Paglinawan
R A
RA
Ward
J F
JF
D004247:DNA / Q000737:chemistry / Q000528:radiation effects; D004249:DNA Damage / Q000528:radiation effects*; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000528:radiation effects; D043382:DNA-Formamidopyrimidine Glycosylase; D005295:Ferrocyanides; D016166:Free Radical Scavengers / Q000737:chemistry; D005609:Free Radicals / Q000737:chemistry*; D007700:Kinetics; D009699:N-Glycosyl Hydrolases / Q000737:chemistry; D010084:Oxidation-Reduction; D010472:Perchlorates / Q000737:chemistry; D010957:Plasmids / Q000737:chemistry* / Q000528:radiation effects*; D011839:Radiation, Ionizing; D017670:Sodium Compounds / Q000737:chemistry; D013793:Thallium / Q000737:chemistry; D013861:Thiocyanates / Q000737:chemistry; D014443:Tyrosine
D004277:DNA, Single-Stranded; D005295:Ferrocyanides; D016166:Free Radical Scavengers; D005609:Free Radicals; D010472:Perchlorates; D017670:Sodium Compounds; D013861:Thiocyanates; D014443:Tyrosine; D004247:DNA; C031068:sodium perchlorate; D013793:Thallium; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase; C020354:hexacyanoferrate II; C031760:thiocyanate