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Oxidative stress appears to be directly involved in the pathogenesis of several neurodegenerative disorders, including Alzheimer and Parkinson diseases. Nigral dopaminergic neurons are particularly exposed to oxidative stress because a pathological accumulation of cytosolic dopamine gives rise to various toxic molecules, including free radicals and reactive quinones. These latter species can react with proteins preventing them from exerting their physiological functions. Among the possible targets of quinones, alpha-synuclein is of primary interest because of its direct involvement in dopamine metabolism. Contrary to the neurotoxic processes, neuromelanin synthesis seems to play a protective role by its ability to sequester a variety of potentially damaging substances. In this study, we carried out a kinetic and structural analysis of the early oxidation products of dopamine. Specifically, considering the potential high toxicity of aminochrome for both cells and mitochondria, we focused our attention on its rearrangement to 5,6-dihydroxyindole. After the spectroscopic characterization of the products derived from the oxidation of dopamine, the structural information obtained was used to analyze the reactivity of quinones toward alpha-synuclein. Our results suggest that indole-5,6-quinone, rather than dopamine-o-quinone or aminochrome, is the reactive species. We propose that the observed reactivity could represent a general reaction pathway whenever cysteinyl residues are absent in proteins or if they are sterically protected.

The preferential degeneration of dopaminergic neurons in the substantia nigra pars compacta is responsible for the motor impairment associated with Parkinson's disease. Dopamine is a highly reactive molecule, which is usually stored inside synaptic vesicles where it is stabilized by the ambient low pH. However, free cytosolic dopamine can auto-oxidize, generating reactive oxygen species, and lead to the formation of toxic quinones. In the present work, we have analyzed the mechanisms through which the dysfunction of dopamine homeostasis could induce cell toxicity, by focusing in particular on the damage induced by dopamine oxidation products at the mitochondrial level. Our results indicate that dopamine derivatives affect mitochondrial morphology and induce mitochondrial membrane depolarization, leading to a reduction of ATP synthesis. Moreover, our results suggest that opening of the mitochondrial transition pore induced by dopamine-derived quinones may contribute to the specific Parkinson's disease-associated vulnerability of dopamine containing neurons.

Tyrosinase (Ty) is a copper-containing enzyme ubiquitously distributed in nature. In recent years, Ty has attracted interest as a potential detoxifying agent for xenobiotic compounds with phenolic structure. Among these, chlorophenols are particularly relevant pollutants, commonly found in waste waters. The activity of Streptomyces antibioticus tyrosinase toward isomeric monochlorophenols was studied. Tyrosinase oxidizes both 3- and 4-chlorophenol to the same product, 4-chloro-1,2-ortho-quinone, which subsequently undergoes a nucleophilic substitution reaction at the chlorine atom by excess phenol to give the corresponding phenol-quinone adduct. By contrast, 2-chlorophenol is not reactive and acts as a competitive inhibitor. Docking calculations suggest that the substrates point to one of the copper atoms of the dinuclear center (copper B) and appear to interact preferentially with one of the two coordinated oxygen atoms. The approach of the substrate toward the active site is favored by a π-stacking interaction with one of the copper-coordinated histidines (His194) and by a hydrogen bonding interaction with the O1 oxygen. With this study, we provide the first characterization of the early intermediates in the biotechnologically relevant reaction of Ty with chlorophenols. Additionally, combining experimental evidences with molecular modeling simulations, we propose a detailed reaction scheme for Ty-mediated oxidation of monochlorophenols. © 2010 Elsevier Inc. All rights reserved.