Most degenerative diseases are caused by free radicals. Antioxidin-RL peptide is a free radical scavenger found in the skin of plateau frog Odorrana livida, which is more stable than vitamin C as it resists light-induced degradation. However, whether and how antioxidin-RL protects cells from oxidative stress was not clear. Here we addressed this issue, and in addition, we designed a series of antioxidin cognates by adding tyrosine residues to enhance free radical-binding capability. We performed free radical-clearing assays in solution to screen the mutants, and found a mutant antioxidin-2 that was as stable as antioxidin-RL and cleared free radical faster. By using PC-12 cells as a model, we demonstrated that both antioxidin-2 and antioxidin-RL inhibited the accumulation of intracellular free radicals triggered by H2O2, reduced mitochondria membrane potential dissipation, maintained mitochondrial morphology, and decreased the expression of dynamin-related protein-1 in mitochondria, with antioxidin-RL more effective. Antioxidin-RL also attenuated the changes in SOD1 and GPx1 expression induced by H2O2. These findings provide insight into the anti-oxidative mechanisms of antioxidin-RL and its derivatives, which will provide rational basis for the development of more effective antioxidants to cure diseases.
Keywords: Antioxidin-RL; PC12 cell; metabolic activity; mitochondria; oxidative injury.