Flavonoid inhibitors of alpha-synuclein fibrillation: A therapeutic strategy for Parkinson's disease, and, The effect of membranes on SMA fibrillation.

摘要

I. alpha-Synuclein fibrillation is closely associated with the formation of Lewy bodies in neurons, and is implicated in the causative pathogenesis of Parkinson's disease (PD). Currently, there is no approved therapeutic agent directed toward preventing the formation of fibrils. which has been recently shown to have a key role in the cytotoxic nature of amyloidogenic proteins. Flavonoids, known as plant pigments, belong to a vast group of polyphenolic compounds. Over 4000 flavonoids have been identified from various plants and foodstuffs derived from plants, and have been demonstrated as potential neuroprotective agents.; In this study 48 flavonoids belonging to several classes with structures differing in the position of double bonds and ring substituents were tested for their ability to inhibit the fibrillation of alpha-synuclein in vitro. A variety of flavonoids inhibited alpha-synuclein fibrillation, and most of the strong inhibitory flavonoids were also found to disaggregate existing fibrils.; The molecular mechanism underlying the inhibition of alpha-synuclein fibrillation induced by the flavonoid was thoroughly examined via various biochemical and biophysical probes. The binding of inhibitory flavonoids to alpha-synuclein via both the covalent modification by the flavonoid quinone (through gel and MS studies) and noncovalent interactions, most likely through H-bonding between the Tyr residues of alpha-synuclein and the flavonoid (through the mutant studies), lead to the restriction of the conformational changes in the protein during incubation, as revealed by unchanged far-UV CD, FTIR spectra and the small peak shift in ANS, as well as the stabilization of soluble species of alpha-synuclein (monomers and oligomers). All of these factors rather than a single one contribute to the inhibition of WT alpha-synuclein fibrillation induced by the flavonoid.; The molecular structural requirements that appear necessary to provide a flavonoid the ability to inhibit a-synuclein fibrillation were determined to be a vicinal dihydroxyphenyl moiety, no matter in which ring position they are located. Flavonoids with three vicinal hydroxyl groups exhibit enhanced inhibitory effects on alpha-synuclein fibrillation.; Our results in this study show that antioxidant activities of flavonoids are roughly correlated with the in vitro inhibitory effects on alpha-synuclein fibrillation despite a few exceptions, which could be due to variation in the reported antioxidant activities of flavonoids from different groups by using different assay systems.; Take together, the inhibitory flavonoids on alpha-synuclein fibrillation may have potential as therapeutic leads in combating Parkinson's disease, and that diets rich in flavonoids may be effective in preventing the disorder.; II. Light chain (or AL) amyloidosis is the most common form of systemic amyloidosis, characterized by the pathological deposition of insoluble fibrils of immunoglobulin light chain fragments in various organs and tissues, especially the kidney and heart. The etiology of the disease is believed to involve the proliferation of monoclonal plasma/B cells, followed by proteolysis of the light chains and deposition of the variable domains, leading to localized cell and tissue death. However, both the triggering factors and the mechanisms involved in the abnormal formation of the insoluble fibrillar aggregates from the soluble proteins are poorly understood. For example, although the fibrillar deposits are typically found associated with the ECM and basement membranes, it is not clear whether fibrils are initially formed intra- or extra-cellularly, nor is it understood what determines where the deposits will occur, i. e. site tropism.; In the present investigation, we studied the interaction of a recombinant amyloidogenic light chain variable domain, SMA, with lipid vesicles. The nature of the interaction was dependent on the lipid composition and the ratio of SMA to lipid. The most p