Inactivation of NADP(+)-dependent isocitrate dehydrogenase by singlet oxygen derived from photoactivated rose bengal

Abstract

Recently, we demonstrated that the control of cytosolic and mitochondrial redox balance and the cellular defense against oxidative damage is one of the primary functions of NADP(+)-dependent isocitrate dehydrogenase (ICDH) by supplying NADPH for antioxidant systems. When exposed to a singlet oxygen-producing system composed of rose bengal (RB) and visible light, ICDH was susceptible to oxidative modification and damage as indicated by the loss of activity and by the formation of carbonyl groups. The structural alterations of modified enzyme were indicated by the increase in susceptibility to proteases and the change in intrinsic fluorescence spectra. Upon exposure to photoactivated RB, a significant decrease in both cytosolic and mitochondrial ICDH activities was observed in HL-60 cells. The singlet oxygen-mediated damage to ICDH may result in the perturbation of cellular antioxidant defense mechanisms and subsequently lead to a pro-oxidant condition. When we examined the antioxidant role of cytosolic ICDH against singlet oxygen-induced damage with HL-60 cells transfected with the cDNA for mouse cytosolic ICDH in sense and antisense orientations, a clear inverse relationship was observed between the amount of cytosolic ICDH expressed in target cells and their susceptibility to singlet oxygen-mediated oxidative damage.