Effect of oxidative stress on stability and structure of neurofilament proteins

Neurofilament proteins are highly phosphorylated molecules in the axonal co mpartment of the adult nervous system. We report the structural analysis of neurofilament proteins after oxidative damage. SDS-PAGE, immunoblotting, c ircular dichroism, and Fourier transform infrared spectroscopy were used to investigate the relative sensitivity of neurofilaments to oxidative stress and to identify changes in their molecular organization. An ascorbate-Fe+3 -O-2 buffer system as well as catechols were used to generate free radicals on a substrate of phosphorylated and dephosphorylated neurofilaments. By F ourier Transform Infrared spectroscopy and circular dichroism, we establish ed that the neurofilament secondary structure is mainly composed of alpha - helices and that after free radical damage of the peptide backbone of neuro filaments, those helices are partly modified into beta -sheet and random co il structures. These characteristic reorganizations of the neurofilament st ructure after oxidative exposure suggest that free radical activity might p lay an important role in the biogenesis of the cytoplasmic inclusions found in several neurodegenerative diseases.