α-Synuclein (α-syn) is a major protein component of the neuropathological hallmarks of Parkinson's disease and related neurodegenerative disorders termed synucleinopathies. Neither the mechanism of α-syn fibrillization nor the degradative process for α-syn has been elucidated. Previously, we showed that wild-type, mutated, and fibrillar α-syn proteins are substrates of calpain I in vitro. In this study, we demonstrate that calpain-mediated cleavage near and within the middle region of soluble α-syn with/without tyrosine nitration and oxidation generates fragments that are unable to self-fibrillize. More importantly, these fragments prevent full-length α-syn from fibrillizing. Calpain-mediated cleavage of α-syn fibrils composed of wild-type or nitrated α-syn generate C-terminally truncated fragments that retain their fibrillar structure and induce soluble full-length α-syn to co-assemble. Therefore, calpain-cleaved soluble α-syn inhibits fibrillization, whereas calpain-cleaved fibrillar α-syn promotes furt...