Interaction of nitric oxide with glutathione or cysteine generates reactive oxygen species causing DNA single strand breaks

It was found that reactive oxygen species (ROS) were generated in the interactions of nitric oxide (NO) with glutathione (GSH) or cysteine (CySH) under aerobic conditions. When supercoiled DNA was incubated with a mixture of NO/GSH, NO/CySH, NOC-7 (a NO donor)/GSH or NOC-7/CySH under aerobic conditions, DNA single-strand breaks were observed on agarose gel electrophoresis. The strand breaks were inhibited by common ROS scavengers: superoxide dismutase + catalase, the spin trapping agent 5,5-dimethyl-1-pyrroline-N oxide (DMPO), ethanol, and EDTA. The strand breaks were also caused by incubation with a mixture of S-nitrosoglutathione (GSNO) with GSH or CySH, which was inhibited by ROS scavengers. In the reaction of NO/GSH, GSNO rapidly formed and then gradually decreased, and in the reaction of GSNO/GSH, GSNO was gradually decreased. The decrease inf GSNO was accelerated in the presence of superoxide + catalase. Hydroxyl radical was detected in the mixtures of NO with GSH or CySH under aerobic conditions, and thiyl radicals were detected in the mixtures of GSNO with GSH or CySH under anaerobic conditions as examined in electron spin resonance studies using DMPO as a spin trap. The results indicate that the interaction of NO with thiols in the presence of 0, generates ROS that caused DNA single-strand breaks.