1. Rates for repair of pBR 322 DNA radicals by thiols as measured by the gas explosion technique: evidence that counter-ion condensation and co-ion depletion are significant at physiological ionic strength.
- Author
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Fahey RC, Prise KM, Stratford MR, Watfa RR, and Michael BD
- Subjects
- Chromatography, High Pressure Liquid, DNA analysis, DNA drug effects, DNA radiation effects, Electrons, Electrophoresis, Agar Gel, Hydrogen-Ion Concentration, In Vitro Techniques, Ions, Plasmids radiation effects, Sulfhydryl Compounds analysis, DNA Damage, DNA Repair, Plasmids drug effects, Radiation-Protective Agents, Sulfhydryl Compounds pharmacology
- Abstract
Rates of repair of pBR 322 plasmid DNA radicals by thiols of varying net charge (Z) at pH 7 and physiological ionic strength were measured using the oxygen explosion technique. The extent of conversion of supercoiled to relaxed circular plasmid was measured by HPLC as a function of the time of oxygen exposure before or after irradiation, the time-courses being fitted by a pseudo-first-order kinetic expression with k1 = k2[RSH]. Values of k2 (M-1 S-1) were: 2.1 x 10(5) (GSH, Z = -1), 1.4 x 10(6) (2-mercaptoethanol, Z = 0), 1.2 x 10(7) (cysteamine, Z = +1), 6.6 x 10(7) (WR-1065 or N-(2-mercaptoethyl)-1,3-diaminopropane, Z = +2). The approximately 6-fold increase in rate with each unit increase in Z is attributed to concentration of cationic thiols near DNA as a consequence of counter-ion condensation and reduced levels of anionic thiols near DNA owing to co-ion depletion. The results are quantitatively consistent with chemical repair as a significant mechanism for radioprotection of cells by neutral and cationic thiols under aerobic conditions, but indicate that repair by GSH will compete effectively with oxygen only at low oxygen tension.
- Published
- 1991
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