Interactions of growth inhibitory factor with hydroxyl and superoxide radicals

To show the effects of growth inhibitory factor (Cu4Zn3MT-III) involved in the scavenging of reactive oxygen species (ROS), a pulse radiolytic study was employed using N2O-saturated Cu4Zn3MT-III aqueous solutions. It was demonstrated that the oxidizing OH⋅ radical efficiently reacted with Cu4Zn3MT-III by forming a thiyl radical RS⋅ with a second-order constant of 1.46×1011 mol l−1s−1, which was determined by competition kinetics against KSCN. The thiyl radical RS⋅ reacted rapidly and reversibly with a thiolate in Cu4Zn3MT-III to form radical anion RSSR⋅ − with a constant of 1.65×109 mol lL−1s−1 per thiolate, while the constant of the decay of this radical anion was 2.72×105 s−1, and the equilibrium constant of the formation for RSSR⋅ − was 6.08×103 mol−1 l. These values were close to those of Cd5Zn2MT-II. The SOD activity of Cu4Zn3MT-III to quench O2⋅ −was assayed by the riboflavine-methionine-nitrobluetetrazolium (NBT) method which catalyzed the dismutation of superoxide (O2⋅ −) at pH 7.8 with an IC50 value of 1.50×10−6 M for Cu4Zn3MT-III and 1.62×10−6 M for Cd5Zn2MT-II. Additionally, the down-regulation of GIF may be a main factor in the decrease of the scavenging ability for the free OH⋅ and O2⋅ −radicals, which is possibly associated with the pathogenesis of neurodegenerative disease.