Analysis of mitochondrial protein sulfenation during aging in the rat brain

Purpose: The purpose of the present study was to identify mitochondrial proteins that undergo changes in cysteine sulfenation during aging. Methods: Studies were conducted in rats when they were 5 or 30 months of age. Following blocking of free protein thiols with N-ethylmaleimide, protein sulfenic acids were reduced by arsenite to free thiol groups that were subsequently labeled with biotin-maleimide. Samples were then comparatively analyzed by 2-dimensional Western blots, and proteins showing changes in sulfenation were selectively identified by mass spectrometry peptide sequencing. Result: Five proteins were identified. Proteins showing an age-related decrease in sulfenation include pyruvate carboxylase and pyruvate dehydrogenase; while those showing an age-related increase in sulfenation include aconitase, mitofilin, and tubulin (α-1). Conclusion: Results of the present study provide a general picture of mitochondrial protein sulfenation in brain oxidative stress and implicate the involvement of protein sulfenation in overall decline of mitochondrial function during brain aging.