α-Synuclein is one of the causative proteins of familial Parkinson disease, which is characterized by neuronal inclusions named Lewy bodies. Lewy bodies include not only α-synuclein but also aggregates of other proteins. This fact raises a question as to whether the formation of α-synuclein amyloid fibrils in Lewy bodies may occur via interaction with fibrils derived from different proteins. To probe this hypothesis, we investigated in vitro fibril formation of human α-synuclein in the presence of preformed fibril seeds of various different proteins. We used three proteins, Escherichia coli chaperonin GroES, hen lysozyme, and bovine insulin, all of which have been shown to form amyloid fibrils. Very surprisingly, the formation of α-synuclein amyloid fibril was accelerated markedly in the presence of preformed seeds of GroES, lysozyme, and insulin fibrils. The structural characteristics of the natively unfolded state of α-synuclein may allow binding to various protein particles, which in turn triggers the formation (extension) of α-synuclein amyloid fibrils. This finding is very important for understanding the molecular mechanism of Parkinson disease and also provides interesting implications into the mechanism of transmissible conformational diseases.

α‐Synuclein (140 amino acids), one of the causative proteins of Parkinson's disease, forms amyloid fibrils in brain neuronal cells. In order to further explore the contributions of the C‐terminal region of α‐synuclein in fibril formation and also to understand the overall mechanism of fibril formation, we reduced the number of negatively charged residues in the C‐terminal region using mutagenesis. Mutants with negative charges deleted displayed accelerated fibril formation compared with wild‐type α‐synuclein, demonstrating that negative charges located in the C‐terminal region of α‐synuclein modulate fibril formation. Additionally, when tyrosine residues located at position 125, 133, and 136 in the C‐terminal region were changed to alanine residue(s), we found that all mutants containing the Tyr136Ala mutation showed delays in fibril formation compared with wild type. Mutation of Tyr136 to various amino acids revealed that aromatic residues located at this position act favorably toward fibril formation. In mutants where charge neutralization and tyrosine substitution were combined, we found that these two factors influence fibril formation in complex fashion. These findings highlight the importance of negative charges and aromatic side chains in the C‐terminal region of α‐synuclein in fibril formation.

The pathogenesis of Alzheimer's disease (AD) is strongly correlated with the aggregation and deposition of the amyloid beta (Aβ1–42) peptide in fibrillar form, and many studies have shown that plant-derived polyphenols are capable of attenuating AD progression in various disease models. In this study, we set out to correlate the effects of anthocyanoside extracts (Vaccinium myrtillus anthocyanoside (VMA)) obtained from bilberry on the in vitro progression of Aβ fibril formation with the in vivo effects of this compound on AD pathogenesis. Thioflavin T fluorescence assays and atomic force microscopy were used to monitor Aβ amyloid formation in in vitro assays. Effects of Aβ amyloids on cellular viability were assayed using cultured Neuro2a cells. Cognitive effects were probed using mice that simultaneously expressed mutant human Aβ precursor and mutant presenilin-2. Addition of VMA inhibited the in vitro formation of Aβ peptide fibrils and also reduced the toxicity of these aggregates toward Neuro2a cells. A diet containing 1% VMA prevented the cognitive degeneration in AD mice. Curiously, this diet-derived retention of cognitive ability was not accompanied by a reduction in aggregate deposition in brains; rather, an increase in insoluble deposits was observed compared with mice raised on a control diet. The paradoxical increase in insoluble deposits caused by VMA suggests that these polyphenols divert Aβ aggregation to an alternate, non-toxic form. This finding underscores the complex effects that polyphenol compounds may exert on amyloid deposition in vivo.

A thermostable aspartase gene (aspB) from Bacillus sp. YM55‐1 was cloned and the gene sequenced. The aspB gene (1407 bp ORF) encodes a protein with a molecular mass of 51 627 Da, consisting of 468 amino‐acid residues. An amino‐acid sequence comparison revealed that Bacillus YM55‐1 aspartase shared 71% homology with Bacillus subtilis aspartase and 49% with Escherichia coli and Pseudomonas fluorescens aspartases. The E. coli TK237/pUCASPB strain, which was obtained by transforming E. coli TK237 (aspartase‐null strain) with a vector plasmid (pUCASPB) containing the cloned aspB gene, produced a large amount of the enzyme corresponding to > 10% of the total soluble protein. The over‐expressed recombinant enzyme (native molecular mass: 200 kDa) was purified effectively and rapidly using heat treatment and affinity chromatography. In order to probe the catalytic residues of this enzyme, two conserved amino‐acid residues, Lys183 and His134, were individually mutated to alanine. Although the tertiary structure of each mutant was estimated to be the same as that of wild‐type aspartase in CD and fluorescence measurements, the Lys183Ala mutant lost its activity completely, whereas His134Ala retained full activity. This finding suggests that Lys183 may be involved in the catalytic activity of this thermostable Bacillus YM55‐1 aspartase.