ROLE OF CHARGE IN THE RADIOPROTECTION OF ESCHERICHIA-COLI BY THIOLS

The role of net charge (Z) of thiols in their ability to radioprotect cells has been investigated in a glutathione (GSH)-deficient strain of E. coli. This strain, 7, is deficient in the enzyme gamma-glutamylcys teine synthetase and allows the effects of added low molecular weight thiols to be studied. Using the gas explosion system it is possible to measure the chemical repair of the free-radical precursors of lethal lesions by thiols in intact cells. The first-order chemical repair rat e in strain 7 is 280s(-1) in comparison with 1100s(-1) in the wild-typ e strain 1157. From the measured difference in the intracellular conce ntration of GSH between the wild-type and the mutant, this gives a sec ond-order repair rate, k(r)'s of 1.23 +/- 0.3 X 10(5) dm(3)mol(-1)s(-1 ). Measurement of intracellular thiol levels after addition of various low molecular weight thiols showed that uptake was rapid, leading to stable thiol levels within 1 min. The ratios of the intracellular to e xtracellular concentrations (C-in/C-out) were 0.74 for 3-mercaptopropi onic acid (Z=-1), 0.56 for 2-mercaptoethanol (Z=0), 1.47 for cysteamin e (Z=+1) and 1.04 for WR1065 (Z=+2). The k(r)'s for these thiols were 1.3 +/- 0.5 X 10(5) dm(3)mol(-1)s(-1) for 30-mercaptopropionic acid, 3 .3 +/- 1.6 x 10(5) dm(3)mol(-1)s(-1) for 2-mercaptoethanol, 3.9 +/- 1. 1 X 10(5) dm(3)mol(-1)s(-1) for cysteamine and 2.7 +/- 1.1 X 10(6) dm( 3)mol(-1)s(-1) for WR1065. These are lower and increase less with char ge than previously published values for chemical repair in isolated pB R322 DNA, probably because of the association of nucleoproteins and po lyamines with the cellular DNA of E. coli. However, the approximate th ree-fold increase in k(r) per unit increase in Z shows that the counte r-ion condensation and co-ion depletion are important in determining t he effectiveness of charged thiols in the radioprotection of E. coli.