α-Synuclein and Dopamine Metabolites DOPAL and DOPAC: A Pathway to New Synthetic Neuromelanin Models

Melanin-protein-Fe conjugates serving as new synthetic analogues of brain neuromelanin were prepared by oxidative oligomerization of dopamine (DA) and its metabolites, 3,4-dihydroxyphenyl acetaldehyde (DOPAL) and 3,4dihydroxyphenylacetic acid (DOPAC) in the presence of α-synuclein (αSyn), Lcysteine, and iron salts. The initial reaction of cysteine with the quinone derivatives of the catechols yields the corresponding cysteinyl-catechols which under iron promoted oxidative conditions convert to cysteinyl-quinones that undergo nucleophilic addition by the side chain of the protein lysines and tyrosines. In the case of dopamine, this reaction is in competition with internal cyclization by the amino group to generate melanochrome so that α-synuclein was found partially unmodified and non-covalently bound to the melanin moiety. For DOPAL and DOPAC the functional groups on the catechol side chains can further stabilize the melanin-protein conjugates through condensation or charge interaction with the several lysines present in αSyn. The presence of cysteine in the melanin component allows to classify the resulting conjugates as pheomelanin-αSyn-Fe derivatives of DA, DOPAL and DOPAC. The iron ions acting as catalysts are initially bound to the catechol groups and remain entrapped into the melanin moieties mostly associated in clusters, with less than 10% present as mononuclear centers. The pheomelanin-αSyn-Fe conjugates are soluble and were characterized by NMR, LCMS, EPR and ICP-OES. The present results show the possible mechanism of incorporation of αSyn into brain neuromelanin