Brain metal dyshomeostasis and altered structural dynamics of the presynaptic protein α-synuclein (αS) are both implicated in the pathology of Parkinson's disease (PD), yet a mechanistic understanding of disease progression in the context of αS structure and metal interactions remains elusive. In this Communication, we detail the influence of iron, a prevalent redox-active brain biometal, on the aggregation propensity and secondary structure of N-terminally acetylated αS (αS), the physiologically relevant form in humans. We demonstrate that under aerobic conditions, Fe(II) commits αS to a PD-relevant oligomeric assembly, verified by the oligomer-selective A11 antibody, that does not have any parallel β-sheet character but contains a substantial right-twisted antiparallel β-sheet component based on CD analyses and descriptive deconvolution of the secondary structure. ThisαS-Fe oligomer does not develop into the β-sheet fibrils that have become hallmarks of PD, even after extended incubation, as verified by TEM imaging and the fibril-specific OC antibody. Thioflavin T (ThT), a fluorescent probe for β-sheet fibril formation, also lacks coordination to this antiparallel conformer. We further show that this oligomeric state is not observed when O is excluded, indicating a role for iron(II)-mediated O chemistry in locking this dynamic protein into a conformation that may have physiological or pathological implications.

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.

The etiology of Parkinson's disease (PD) is linked with cellular inclusions in the substantia nigra pars compacta region of the brain that are enriched in the misfolded presynaptic protein α-synuclein (αS) and death of the dopaminergic neurons. Brain iron homeostasis governs both neurotransmission and neurodegeneration; hence, the role of iron in PD progression and neuronal health is apparent. Elevated iron deposits become prevalent in the cerebral region upon aging and even more so in the PD brain. Structural as well as oxidative modifications can result from coordination of αS with redox active iron, which could have functional and/or pathological implications. In this review, we will discuss iron-mediated αS aggregation, alterations in iron metabolism, and the role of the iron-dopamine couple. Moreover, iron interactions with N-terminally acetylated αS, the physiologically relevant form of the human protein, will be addressed to shed light on the current understanding of protein dynamics and the physiological environment in the disease state. Oxidative pathways and biochemical alterations resulting from aberrant iron-induced chemistry are the principal focus of this review in order to highlight the plethora of research that has uncovered this emerging dichotomy of iron playing both functional and disruptive roles in PD pathology.

Novel therapeutics are needed to treat pathologies associated with the Clostridioides difficile binary toxin (CDT), particularly when C. difficile infection (CDI) occurs in the elderly or in hospitalized patients having illnesses, in addition to CDI, such as cancer. While therapies are available to block toxicities associated with the large clostridial toxins (TcdA and TcdB) in this nosocomial disease, nothing is available yet to treat toxicities arising from strains of CDI having the binary toxin. Like other binary toxins, the active CDTa catalytic subunit of CDT is delivered into host cells together with an oligomeric assembly of CDTb subunits via host cell receptor-mediated endocytosis. Once CDT arrives in the host cell’s cytoplasm, CDTa catalyzes the ADP-ribosylation of G-actin leading to degradation of the cytoskeleton and rapid cell death. Although a detailed molecular mechanism for CDT entry and host cell toxicity is not yet fully established, structural and functional resemblances to other binary toxins are described. Additionally, unique conformational assemblies of individual CDT components are highlighted herein to refine our mechanistic understanding of this deadly toxin as is needed to develop effective new therapeutic strategies for treating some of the most hypervirulent and lethal strains of CDT-containing strains of CDI.