Copper Induced Radical Dimerization of α-Synuclein Requires Histidine

Aggregation of the neuronal protein α-synuclein (αS) is a critical factor in the pathogenesis of Parkinson’s disease. Analytical methods to detect post-translational modifications of αS are under development, yet the mechanistic underpinnings of biomarkers like dityrosine formation within αS have yet to be established. In our work, we demonstrate that Cu I -bound N-terminally acetylated αS ( NAc αS) activates O 2 resulting in both intermolecular dityrosine cross-linking within the fibrillar core as well as intramolecular cross-linking within the C-terminal region. Substitution of the H50 residue with a disease relevant Q mutation abolishes intermolecular dityrosine cross-linking and limits the Cu I /O 2 promoted cross-linking to the C-terminal region. Such a dramatic change in reaction behavior establishes a previously unidentified role for H50 in facilitating intermolecular cross-linking. Involvement of H50 in the reaction profile implies that long-range histidine coordination with the upstream Cu I coordination site is necessary to stabilize the transition of Cu I to Cu II as is a required mechanistic outcome of Cu I /O 2 reactivity. The aggregation propensity of NAc H50Q–Cu I is also enhanced in comparison to NAc αS–Cu I, suggesting a potential functional role for both copper and intermolecular cross-linking in attenuating NAc αS fibrillization.