Mechanisms of photodynamic inactivation of acridine orange-sensitized transfer RNA : Participation of singlet oxygen and base damage leading to inactivation.

Abstract

The photodynamic inactivation of acridine orange (AO)-sensitized tRNA was investigated with respect to the type of photochemical pathways and the base damage leading to the inactivation. The diagnostic tests for singlet oxygen (¹O₂), namely the enhancement effect of deuterium oxide (D₂O) and the inhibition effect by sodium azide, indicated that ¹O₂ was involved as the reactive intermediate in the photoinactivation (the Type II mechanism). Furthermore, the D₂O effect was more pronounced at high ionic strength and under low dye to RNA ratio conditions, indicating that monomerically bound AO acts exclusively through the Type II mechanism. With respect to base damage, the present studies show that guanine is destroyed and tRNA-AO occurs as a photoadduct. The formation of photoadduct was enhanced under N₂, and little affected by D₂O, excluding the possibility that the photoadduct is responsible for the photoinactivation. Conversely, effects of D₂O and azide on the photoinactivation, in conjunction with the predominance of the Type II mechanism in the known cases of guanine destruction by AO-photosensitization, indicate that the destruction of guanine by ¹O₂ is the most probable critical damage in the photodynamic inactivation of tRNA.