Monomers, Dimers, and Oligomers of Pyroglutamate-Modified α-Synuclein Fragments Exhibit Distinct Biophysical Characteristics.

α-Synuclein (aSyn) aggregation represents a key event in the neurodegenerative cascade of synucleinopathies. Initially, aSyn appears as an intrinsically disordered protein. However, its structural flexibility allows aSyn to either adopt α-helical conformations, relevant for physiological functions at presynaptic vesicles, or form β-strand-rich aggregates, leading to toxic oligomers. This relation between structure, function, and toxicity can be influenced by post-translational modifications such as the recently identified glutaminyl cyclase-catalyzed pyroglutamate (pE) modification. Here, we investigated (i) structural characteristics of monomeric, dimeric, and oligomeric states of N-terminal truncated, pE-modified aSyn variants, pE24-, pE62-, and pE79-aSyn by a complementary biophysical approach including DLS, SEC-MALS, SRCD, SEC-SAXS, and AUC and (ii) the toxicity of oligomeric pE-aSyn variants compared to full-length aSyn. Overall, pE62-aSyn showed an immediate fibril formation, reflecting the aggregation-prone properties of this particular variant. Furthermore, in a membrane-like environment, the secondary aSyn structure shifted toward α-helical folding depending on the degree of N-terminal truncation. pE79-aSyn showed a significantly reduced level of structural adaptation, reflecting compromised functions at presynaptic vesicles. In addition, the comparative analysis indicates the presence of a dimeric aSyn intermediate, the initial and potentially crucial step in aSyn aggregation, and supports the hypothesis of a toxic porous oligomeric state. For the first time, based on SAXS data, EOM models of the dimeric aSyn state are proposed.