Generation and reactions of the disulphide radical anion derived from metallothionein: a pulse radiolytic study.

OH-radicals were generated by pulse radiolysis of aqueous solutions of rabbit (Zn,Cd)-metallothionein (MT). They react with MT mainly by forming a thiyl radical with a rate constant of 1.7 x 10(12) dm3 mol-1 s-1. The thiyl radical reacts rapidly but reversibly with a thiolate function to form RSSR.-: RS + RS- reversible RSSR.-. The kinetics of the formation and decay of this radical anion have been studied pulse radiolytically by monitoring the evolution of the optical absorption of RSSR.- at 450 nm. This process is mostly intermolecular, i.e. bimolecular in MT. In the absence of O2, RSSR.- decays bimolecularly: RSSR.(-)+RS.-->RSSR + RS-. In the presence of O2, RS. may be scavenged by O2 and thus the yield of RSSR.- decreases: RS.+O2 reversible RSOO.. Under these conditions RSSR.- decays by first-order kinetics: RSSR.(-)+O2-->RSSR + O2.-. The rate constants of these reactions have been determined at room temperature: k4 = 1.8 x 10(9) dm3 mol-1 s-1, k5 = 7 x 10(4) s-1, k10 = 9.2 x 10(8) dm3 mol-1 s-1, and k18 about 3 x 10(7) dm3 mol-1 s-1. From the dependence of the maximal absorbance at 450 nm on the thiolate concentration in the absence of oxygen, epsilon (RSSR.-) = 9 x 10(3) dm3 mol-1 cm-1 and the stability constant (K4/5) of 2.3 x 10(4) dm3 ml-1 was determined. K4/5 is in good agreement with that determined kinetically, k4/k5 = 2.6 x 10(4) dm3 mol-1. The stability constant K15/16 of the thiylperoxyl radical, RSOO., was determined to be 5.5 x 10(3) dm3 mol-1.