Edited by Wolfgang Peti ␣-Synuclein (␣S) is the primary protein associated with Parkinson's disease, and it undergoes aggregation from its intrinsically disordered monomeric form to a cross-␤ fibrillar form. The closely related homolog ␤-synuclein (␤S) is essentially fibril-resistant under cytoplasmic physiological conditions. Toxic gain-of-function by ␤S has been linked to dysfunction, but the aggregation behavior of ␤S under altered pH is not wellunderstood. In this work, we compare fibril formation of ␣S and ␤S at pH 7.3 and mildly acidic pH 5.8, and we demonstrate that pH serves as an on/off switch for ␤S fibrillation. Using ␣S/␤S domain-swapped chimera constructs and single residue substitutions in ␤S, we localized the switch to acidic residues in the N-terminal and non-amyloid component domains of ␤S. Computational models of ␤S fibril structures indicate that key glutamate residues (Glu-31 and Glu-61) in these domains may be sites of pH-sensitive interactions, and variants E31A and E61A show dramatically altered pH sensitivity for fibril formation supporting the importance of these charged side chains in fibril formation of ␤S. Our results demonstrate that relatively small changes in pH, which occur frequently in the cytoplasm and in secretory pathways, may induce the formation of ␤S fibrils and suggest a complex role for ␤S in synuclein cellular homeostasis and Parkinson's disease. ␣-Synuclein (␣S),5 the primary protein component of intracytoplasmic inclusions known as Lewy bodies (LB) in Parkinson's disease (PD), is a 140-residue, predominantly monomeric, intrinsically disordered protein (IDP) (1-6). It is abundant in the cytosol but present in many organelles (7,8). The monomers of ␣S can aggregate to soluble oligomers as well as insoluble oligomers and fibrils, which are considered to be associated with the pathogenesis of PD (9). A closely related family member, ␤-synuclein (␤S) that co-localizes with ␣S, has ϳ60% sequence identity with ␣S but has not been detected in LBs of PD patients (10, 11). Instead, it has been proposed that ␤S may delay ␣S fibril formation and ameliorate ␣S toxicity in vivo by inhibiting ␣S aggregation (12-14). Furthermore, despite the high sequence similarity, ␤S itself does not form fibrils in vitro at cytoplasmic pH without facilitating agents (15). However, recent reports highlight a role for a possible toxic gain-of-function of ␤S in two different model systems (16,17), and ␤S is a component of vesicle-like lesions in the hippocampus, whose formation accompanies dementia in PD (18). In addition, two ␤S mutations, V70M and P123H, were found in sporadic and familial dementia with LBs and have been suggested to be involved in lysosomal pathology (19 -21). There is an emerging role for ␤S in the pathophysiology of PD, but the molecular determinants of this role remain poorly studied.Understanding the complex relationship of synucleins with neurodegeneration requires consideration of their structures and functions in diverse subcellular environments, beyond the cytoplasm. Altho...

The intrinsically disordered protein β-synuclein is known to inhibit the aggregation of its intrinsically disordered homolog, α-synuclein, which is implicated in Parkinson's disease. While β-synuclein itself does not form fibrils at the cytoplasmic pH 7.4, alteration of pH and other environmental perturbations are known to induce its fibrilization. However, the sequence and structural determinants of β-synuclein inhibition and self-aggregation are not well understood. We have utilized a series of domain-swapped chimeras of α-synuclein and β-synuclein to probe the relative contributions of the N-terminal, C-terminal, and the central non-amyloid-β component domains to the inhibition of α-synuclein aggregation. Changes in the rates of α-synuclein fibril formation in the presence of the chimeras indicate that the non-amyloid-β component domain is the primary determinant of self-association leading to fibril formation, while the N- and C-terminal domains play critical roles in the fibril inhibition process. Our data provide evidence that all three domains of β-synuclein together contribute to providing effective inhibition, and support a model of transient, multi-pronged interactions between IDP chains in both processes. Inclusion of such multi-site inhibitory interactions spread over the length of synuclein chains may be critical for the development of therapeutics that are designed to mimic the inhibitory effects of β-synuclein.

Glycation of α-synuclein (αSyn), as occurs with aging, has been linked to the progression of Parkinson’s disease (PD) through the promotion of advanced glycation end-products and the formation of toxic oligomers that cannot be properly cleared from neurons. DJ-1, an antioxidative protein that plays a critical role in PD pathology, has been proposed to repair glycation in proteins, yet a mechanism has not been elucidated. In this study, we integrate solution nuclear magnetic resonance (NMR) spectroscopy and liquid atomic force microscopy (AFM) techniques to characterize glycated N-terminally acetylated-αSyn (glyc-ac-αSyn) and its interaction with DJ-1. Glycation of ac-αSyn by methylglyoxal increases oligomer formation, as visualized by AFM in solution, resulting in decreased dynamics of the monomer amide backbone around the Lys residues, as measured using NMR. Upon addition of DJ-1, this NMR signature of glyc-ac-αSyn monomers reverts to a native ac-αSyn-like character. This phenomenon is reversible upon removal of DJ-1 from the solution. Using relaxation-based NMR, we have identified the binding site on DJ-1 for glycated and native ac-αSyn as the catalytic pocket and established that the oxidation state of the catalytic cysteine is imperative for binding. Based on our results, we propose a novel mechanism by which DJ-1 scavenges glyc-ac-αSyn oligomers without chemical deglycation, suppresses glyc-ac-αSyn monomer–oligomer interactions, and releases free glyc-ac-αSyn monomers in solution. The interference of DJ-1 with ac-αSyn oligomers may promote free ac-αSyn monomer in solution and suppress the propagation of toxic oligomer and fibril species. These results expand the understanding of the role of DJ-1 in PD pathology by acting as a scavenger for aggregated αSyn.