Dopaminergic neurons in the substantia nigra, a small region of the mid brain producing dopamine, undergo extensive and selective loss with the progression of Parkinson's disease (PD), with less substantial death in other brain regions. The surviving dopaminergic cells of PD patients contain cytosolic filamentous inclusions known as Lewy bodies, within which ŕ-synuclein (ŕSyn) is a major component. Neurofibrillary tangles, containing aggregated tau, are also present. Accumulation of toxic forms of ŕSyn is considered a critical step in the development of PD, and it is currently accepted that these toxic forms are protofibrillar aggregates of ŕSyn and post-transactionally modified ŕSyn. The pattern of post-translational ŕSyn modifications identified in cytosolic aggregates in experimental models and PD brains include oxidation, nitration, and phosphorylation, but the form of ŕSyn that is thought to be mostly responsible for neuron toxicity is the oligomeric protein resulting from the interaction with oxidized dopamine. The role of dopamine is intriguing, as this neurotransmitter is also involved in the biosynthesis of neuromelanin (NM), the pigment contained in substantia nigra neurons, which is formed upon extensive oxidative oligomerization of dopamine and conjugation with protein/peptides. Oxidative stress and redox metal ions are strong determinants of post-translational modifications of ŕSyn and other neuronal proteins like tau. In this project, we plan to perform a systematic, quantitative and comparative study of the redox reactivity, capacity of generating ROS and RNS, and catalytic oxidative activity of thoroughly characterized iron and copper complexes with ŕSyn and tau. We believe this is the necessary step to enable a reliable assessment of the toxicity of metal ions bound to neuronal proteins and peptides in a given environment. The toxicity associated with ROS production by metal-ŕSyn and metal-tau peptides will be evaluated in cellular models. The project develops along several, inter-related research lines: 1) Binding characteristics, ROS production, dopamine oxidation, and protein modification by Cu and Fe complexes with N-acetylated ŕSyn (Ac-ŕSyn) in solution and membrane environment. 2) Reactions of Cu- and Fe-Ac-ŕSyn complexes with NO donors, production of RNS, and protein nitration. 3) Binding and reactivity behavior of Cu and Fe complexes with oxidized, nitrated, and phosphorylated Ac-ŕSyn. 4) Interaction of Fe and Cu with tau fragments: binding, redox behavior, ROS and RNS production, reactivity towards biological substrates. 5) Heme toxicity: binding and reactivity behavior with tau and ?-amyloid 6) ROS production by Cu and Fe complexes with ŕSyn and tau in cellular models 7) Protein target modification by ROS/RNS and dopamine in cells 8) Dopamination of proteins and biosynthetic pathway to NM 9) Melanin-protein conjugates as NM analogues: microglia activation in neuronal cell models.