Coordination Features and Affinity of the Cu2+ Site in the α-Synuclein Protein of Parkinson’s Disease

摘要

Parkinson’s disease (PD) is the second most prevalent age-related, neurodegenerative disorder,naffecting >1% of the population over the age of 60. PD pathology is marked by intracellular inclusionsncomposed primarily of the protein R-synuclein (R-syn). These inclusions also contain copper, and theninteraction of Cu2þ with R-syn may play an important role in PD fibrillogenesis. Here we report thenstoichiometry, affinity, and coordination structure of the Cu2þ-R-syn complex. Electron paramagneticnresonance (EPR) titrations show that monomeric R-syn binds 1.0 equiv of Cu2þ at the protein N-terminus.nNext, an EPR competition technique demonstrates that R-syn binds Cu2þ with a Kd of ≈0.10 nM. Finally,nEPRand electron spin echomodulation (ESEEM) applied to a suite ofmutant and truncated R-syn constructsnreveal a coordination sphere arising from the N-terminal amine, the Asp2 amide backbone and side chainncarboxyl group, and the His50 imidazole. The high binding affinity identified here, in accord with previousnmeasurements, suggests that copper uptake and sequestration may be a part of R-syn’s natural function,nperhaps modulating copper’s redox properties. The findings further suggest that the long-range interactionnbetween the N-terminus and His50 may have a weakening effect on the interaction of R-syn with lipidnmembranes, thereby mobilizing monomeric R-syn and hastening fibrillogenesis