The C-terminal α-helices of mammalian Hsc70 play a critical role in the stabilization of α-synuclein binding and inhibition of aggregation.

Protein misfolding, followed by aggregation and amyloid formation is an underlying pathological hallmark in a number of prevalent diseases, including Parkinson's (PD), Alzheimer's (AD) and Type 2 diabetes (T2D). In the case of PD, the aggregation of α-synuclein protein (α-syn) has been shown to be highly cytotoxic and to play a key role in the death of dopaminergic cells. Thus, inhibition of the aggregation process may be considered as an attractive avenue for therapeutic intervention. In this respect, molecular chaperones, known to promote proper folding of proteins, are able to inhibit protein aggregation thus preventing amyloid formation. In this work, the effect of the constitutively expressed chaperone Hsc70 and its various domains on α-syn aggregation have been investigated using different approaches. The results show that the C-terminal domain alone (residues 386-646) is as efficient in inhibiting α-syn aggregation as the entire Hsc70 protein, by increasing the lag phase for α-syn oligomeric nucleus formation, suggesting that the chaperone interacts with and stabilizes α-syn monomers and/or small aggregates. Deletion of the C-terminal helices (residues 510-646), which are known to play the role of a lid locking target peptide ligands in the peptide-binding site of the chaperone, strongly reduced the efficiency of inhibition of α-syn aggregation indicating that these helices play an essential in stabilizing the interaction between Hsc70 and α-syn. Furthermore, the effects of Hsc70 and its structural domains on aggregation appear to correlate with those on cytotoxicity, by reducing the fraction of α-syn toxic species to various degrees. Together these results suggest a mechanism in which inhibition of synuclein aggregation is the result of monomeric synuclein binding to the chaperone as any monomeric target unfolded protein or peptide binding to the chaperone.