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.
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.
Fahey
R C
RC
Cancer Research Campaign Gray Laboratory, Mount Vernon Hospital, Northwood, Middlesex, UK.
Prise
K M
KM
Stratford
M R
MR
Watfa
R R
RR
Michael
B D
BD
D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis / Q000187:drug effects / Q000528:radiation effects; D004249:DNA Damage; D004260:DNA Repair; D004583:Electrons; D004587:Electrophoresis, Agar Gel; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007477:Ions; D010957:Plasmids / Q000187:drug effects* / Q000528:radiation effects; D011837:Radiation-Protective Agents; D013438:Sulfhydryl Compounds / Q000032:analysis / Q000494:pharmacology*
D007477:Ions; D011837:Radiation-Protective Agents; D013438:Sulfhydryl Compounds; D004247:DNA