Characterization of α-Synuclein Interactions with Selected Aggregation-Inhibiting Small Molecules

Abstract

The 140-residue protein alpha-synuclein (aS) has been implicated in the molecular chain of events leading to Parkinson's disease, which relates to the hierarchical aggregation of aS into soluble oligomers and insoluble fibrils. A number of small organic molecules have been reported to inhibit aS aggregation. Here, the interactions of chlorazole black E, Congo red, lacmoid, PcTS-Cu (2+), and rosmarinic acid with aS are examined by NMR spectroscopy to identify aS sequence elements that are masked by these compounds. Surprisingly, similar aS interaction sites, encompassing residues 3-18 and 38-51, were obtained for all molecules at equimolar small molecule:aS ratios. At higher ratios, virtually the entire amphiphilic region of aS (residues 2-92) is affected, revealing the presence of additional, lower affinity interaction sites. Upon rearranging the high-affinity interaction sites over the aS amphiphilic region in an aS mutant form, perturbations of the entire amphiphilic region were found to have already been obtained at equimolar ratios, indicating a high specificity for the original binding sites. CD spectroscopy reveals that, in the presence of the small molecules, the aS structure is still dominated by random-coil characteristics. The strongest effects are exerted by molecules that contain sulfonate groups adjacent to aromatic systems, often present in multiple copies in a symmetrical arrangement, suggesting that these elements are useful for developing an aS-specific chemical chaperone.

Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein α-Synuclein

“…Moreover, our data highlight the key role of the N-terminal part of the protein on early oligomeric interactions. In this regard, we observe that a number of small-molecule anti-amyloid compounds preferentially target the regions enclosing residues ϳVal 3 -Lys 23 and/or Leu 38 -Gly 51 (39,43). As discussed previously, our excimer profile suggests that ␤5 region is not proximal in oAS (Fig.…”

A Novel “Molecular Tweezer” Inhibitor of α-Synuclein Neurotoxicity in Vitro and in Vivo

“…Several groups have reported inhibitors of α-syn assembly and toxicity, including N-methylated peptides [28] dopamine and L-dopa [29], rifampicin [30], curcumin [31], and various other compounds [32][33][34]. In most cases, the compounds have been shown to inhibit α-syn aggregation in vitro, whereas inhibition of toxicity was not studied.…”

Structural Determinants of Tau Aggregation Inhibitor Potency

“…For example, NMR spectroscopy can detect direct interactions between pthalocyanine and Tau protein (24). More generally, direct interactions between diazo dyes, phenothiazines, polyalcohols, and pthalocyanine with monomeric ␤-amyloid (67) and ␣-synuclein (25,68,69) have been reported as well. However, this approach requires high-micromolar to low-millimolar concentrations of ligand, and it is not clear that natively unfolded protein structures could support such interactions at the low protein concentrations used to demonstrate aggregation inhibition.…”