Galantamine suppresses α-synuclein aggregation by inducing autophagy via the activation of α7 nicotinic acetylcholine receptors.

Synucleinopathies, including Parkinson's disease and dementia with Lewy bodies, are neurodegenerative disorders characterized by the aberrant accumulation of α-synuclein (α-syn). Although no treatment is effective for synucleinopathies, the suppression of α-syn aggregation may contribute to the development of numerous novel therapeutic targets. Recent research revealed that nicotinic acetylcholine (nACh) receptor activation has neuroprotective effects and promotes the degradation of amyloid protein by activating autophagy. In an in vitro human-derived cell line model, we demonstrated that galantamine, the nAChR allosteric potentiating ligand, significantly reduced the cell number of SH-SY5Y cells with intracellular Lewy body-like aggregates by enhancing the sensitivity of α7-nAChR. In addition, galantamine promoted autophagic flux, and prevented the formation of Lewy body-resembled aggregates. In an in vivo synucleinopathy mouse model, the propagation of α-syn aggregation in the cerebral cortex was inhibited by galantamine administration for 90 days. These results suggest that α7-nAChR is expected to be a novel therapeutic target, and galantamine is a potential agent for synucleinopathies.

Optimizing start-up strategies for the two-inflow nitritation/anammox process: Influence on biofilm microbial community composition.

Low-energy nitrogen removal from ammonium-rich wastewater is crucial in preserving the water environment. A one-stage nitritation/anammox process with two inflows treating ammonium-containing wastewater, supplied from inside and outside the wound filter, is expected to stably remove nitrogen. Laboratory-scale reactors were operated using different start-up strategies; the first involved adding nitritation inoculum after anammox biomass formation in the filter, which presented a relatively low nitrogen removal rate (0.171 kg N/m3 · d), at a nitrogen loading rate of 1.0 kg N/m3 · d. Conversely, the second involved the gradual cultivation of anammox and nitritation microorganisms, which increased the nitrogen removal rate (0.276 kg N/m3 · d). Furthermore, anammox (Candidatus Brocadia) and nitritation bacteria (Nitrosomonadaceae) coexisted in the biofilm formed on the filter surface. The abundance of nitritation bacteria (10.5%) in the reactor biofilm using the second start-up strategy was higher than that using the first (3.7%). Thus, the two-inflow nitritation/anammox process effectively induced habitat segregation using a suitable start-up strategy.

Antimicrobial resistome and mobilome in the urban river affected by combined sewer overflows and wastewater treatment effluent.

The dissemination of antimicrobial resistance in the environment is an emerging global health problem. Wastewater treatment effluent and combined sewer overflows (CSOs) are major sources of antimicrobial resistance in urban rivers. This study aimed to clarify the effect of municipal wastewater treatment effluent and CSO on antimicrobial resistance genes (ARGs), mobile gene elements, and the microbial community in an urban river. The ARG abundance per 16S-based microbial population in the target river was 0.37-0.54 and 0.030-0.097 during the CSO event and dry weather, respectively. During the CSO event, the antimicrobial resistome in the river shifted toward a higher abundance of ARGs to clinically important drug classes, including macrolide, fluoroquinolone, and ß-lactam, whereas ARGs to sulfonamide and multidrug by efflux pump were relatively abundant in dry weather. The abundance of intI1 and tnpA genes were highly associated with the total ARG abundance, suggesting their potential application as an indicator for estimating resistome contamination. Increase of prophage during the CSO event suggested that impact of CSO has a greater potential for horizontal gene transfer (HGT) via transduction. Consequently, CSO not only increases the abundance of ARGs to clinically important antimicrobials but also possibly enhances potential of HGT in urban rivers.

Effects of organic carbon and sulfide on the anammox reaction in the anoxic column in the SRDAPN process for treating high-strength wastewater.

Low energy consumption treatment of high-strength wastewater is crucial in controlling groundwater pollution and eutrophication in closed waterbodies. In this study, the sulfate reduction, denitrification/anammox, and partial nitrification (SRDAPN) process, which is an effective organic carbon and nitrogen removal process with low energy consumption for low strength wastewater, was applied to treat livestock wastewater with high COD and sulfate concentration, and microbial reaction and community were examined using an anaerobic-anoxic biological filter reactor that simulates circulation from an aerobic reactor. At a total organic carbon loading rate of 2.7-5.8 kgC/m3·day, sulfate reduction and methane production occurred simultaneously in the anaerobic column of the reactor. Specifically, sulfate reduction resulted in organic matter removal rates of 38 and 26% at ambient temperature and 25 °C, respectively. Furthermore, both heterotrophic and autotrophic denitrification occurred in the anoxic column, and when the organic loading rate in the anoxic reactor was below 0.2 kgC/m3·day, 33%-37% of ammonium and 33%-34% of nitrite were removed by the anammox reaction. Heterotrophic denitrification bacteria (Thauera, Comamonas, and Denitratisoma) and sulfur denitrification bacteria (Sulfurimonas denitrificans) grew in the lower and middle parts of the anoxic column, whereas anammox bacteria (2.5% of Candidatus Brocadia at ambient temperature and 9.4% of Candidatus Kuenenia at 25 °C) grew in the upper part of the anoxic column. These results indicate that the SRDAPN process based on sulfur cycle and anammox is useful for treatment of high strength wastewater with low energy consumption.

Effects of high-strength landfill leachate effluent on stress-induced microalgae lipid production and post-treatment micropollutant degradation.

This research investigates the effects of landfill leachate effluent concentrations from moving bed biofilm reactor (MBBR) on stress-induced Chlorella vulgaris and Scenedesmus armatus lipid production and post-treatment micropollutant degradation. The effluent concentrations were varied between 25%, 50%, 75%, and 100% (v/v). The landfill leachate influent was treated using two-stage moving bed biofilm reactor under 24 h and 18 h hydraulic retention time (HRT). The results indicated that the effluent concentration was positively correlated with the stress-induced microalgae lipid production in the post-treatment of residual micropollutants. C. vulgaris and S. armatus completely remove residual micropollutants in the effluent. The superoxide dismutase and peroxidase activity were positively correlated with the cellular lipid content. The lipid content of C. vulgaris and S. armatus cultivated in the 18 h HRT effluent were 31-51% and 51-64%, while those in the 24 h HRT effluent were 15-16% and 5-19%. The optimal condition of microalgae cultivation for the post-treatment of residual micropollutants was 50-75% (v/v) effluent concentrations under 18 h HRT, achieving the highest lipid production of 113-116 mg/L for C. vulgaris and 74-75 mg/L for S. armatus. Essentially, the MBBR landfill leachate effluent holds promising potential as a substrate for microalgae lipid production.

Molecular Mechanisms of Aggregation of Canine SOD1 E40K Amyloidogenic Mutant Protein.

Canine degenerative myelopathy (DM) is a human amyotrophic lateral sclerosis (ALS)-like neurodegenerative disease. It is a unique, naturally occurring animal model of human ALS. Canine DM is associated with the aggregation of canine superoxide dismutase 1 (cSOD1), which is similar to human ALS. Almost 100% of cases in dogs are familial, and the E40K mutation in cSOD1 is a major causative mutation of DM. Therefore, it is important to understand the molecular mechanisms underlying cSOD1(E40K) aggregation. To address this, we first analyzed the structural model of wild type cSOD1. Interactions were evident between amino acid E40 and K91. Therefore, the mutation at residue E40 causes loss of the interaction and may destabilize the native structure of cSOD1. Differential scanning fluorimetry revealed that the E40K mutant was less stable than the wild type. Moreover, stability could be recovered by the E40K and K91E double mutation. Acceleration of amyloid fibril formation in vitro and aggregate formation in cells of cSOD1(E40K) was also suppressed by the introduction of this double mutation in thioflavin T fluorescence assay results and in transfectant cells, respectively. These results clearly show the importance of the interaction between amino acid residues E40 and K91 in cSOD1 for the stability of the native structure and aggregation.

Metagenomic insights into the effect of sulfate on enhanced biological phosphorus removal.

Excess phosphorus in water supplies causes eutrophication, which degrades water quality. Hence, the efficient removal of phosphorus from wastewater represents a highly desirable process. Here, we evaluated the effect of sulfate concentration on enhanced biological phosphorus removal (EBPR), in which phosphorus is typically removed under anaerobic-oxic cycles, with sulfate reduction the predominant process in the anaerobic phase. Two sequencing batch EBPR reactors operated under high- (SBR-H) vs. low-sulfate (SBR-L) concentrations for 189 days and under three periods, i.e., start-up, sufficient acetate, and limited acetate. Under acetate-rich conditions, phosphorus removal efficiency was > 90% for both reactors; however, under acetate-limited conditions, only 34% and 91.3% of the phosphorus were removed for the SBR-L and the SBR-H, respectively. Metagenomic sequencing of the reactors showed that the relative abundance of the polyphosphate-accumulating and sulfur-reducing bacteria (SRB) was higher in the SBR-H, consistent with its higher phosphorus removal activity. Ten high-quality metagenome-assembled genomes, including one closely related to the genus Thiothrix disciformis (99.81% average amino acid identity), were recovered and predicted to simultaneously metabolize phosphorus and sulfur by the presence of phosphorus (ppk, ppx, pst, and pit) and sulfur (sul, sox, dsr, sqr, apr, cys, and sat) metabolism marker genes. The omics-based analysis provided a holistic view of the microbial ecosystem in the EBPR process and revealed that SRB and Thiothrix play key roles in the presence of high sulfate.Key points• We observed high phosphorus-removal efficiency in high-sulfate EBPR.• Metagenome-based analysis revealed sulfate-related metabolic mechanisms in EBPR.• SRB and PAOs showed interrelationships in the EBPR-sulfur systems.

Kaempferol Has Potent Protective and Antifibrillogenic Effects for α-Synuclein Neurotoxicity In Vitro.

Aggregation of α-synuclein (α-Syn) is implicated in the pathogenesis of Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA). Therefore, the removal of α-Syn aggregation could lead to the development of many new therapeutic agents for neurodegenerative diseases. In the present study, we succeeded in generating a new α-Syn stably expressing cell line using a piggyBac transposon system to investigate the neuroprotective effect of the flavonoid kaempferol on α-Syn toxicity. We found that kaempferol provided significant protection against α-Syn-related neurotoxicity. Furthermore, kaempferol induced autophagy through an increase in the biogenesis of lysosomes by inducing the expression of transcription factor EB and reducing the accumulation of α-Syn; thus, kaempferol prevented neuronal cell death. Moreover, kaempferol directly blocked the amyloid fibril formation of α-Syn. These results support the therapeutic potential of kaempferol in diseases such as synucleinopathies that are characterized by α-Syn aggregates.

Initial behaviors and removal of extracellular plasmid gene in membrane bioreactor.

Extracellular antibiotic resistance genes (eARG) are considered to play an important role in spread of antimicrobial resistance (AMR) in wastewater treatment and water environment. Membrane bioreactor (MBR) reportedly has better removal of ARGs in wastewater than conventional activated sludge process. However, removal of eARG is possibly limited because eARG is small to pass through microfiltration (MF) membranes. To evaluate potential removal of eARG in MBR, this study aimed to understand the initial behaviors of eARG received in MBR. The recombinant plasmid with artificial marker gene was spiked in lab-scale MBR to trace fate of eARG in MBR. Among 10 10 copies/L of the spiked gene, 2.6 × 109 copies/L was adsorbed on sludge particles at 6 h after spiking, while only 2.2 × 108-3.6 × 108 copies/L of the spiked gene was remained but constant in sludge liquid phase from 6 until 48 h. This result suggests that adsorption on sludge particles served as the main mechanism to govern the initial fate of eARG in MBR. Meanwhile, the spiked gene concentrations in membrane permeate was lower than sludge liquid phase and decreased overtime, suggesting retention of eARG in membrane filtration. Total LRV of the spiked extracellular gene were 3.4 ± 0.8 log at 48 h after spiking. LRV by adsorption corresponded to 1.7 ± 0.7 log constantly since 3 h after spiking, while LRV by membrane filtration increased from 0 to 1.7 ± 0.6 log. Linear correlation of LRV by membrane filtration with transmembrane pressure (TMP) suggested that foulant deposition on membrane governs removal of eARG by membrane filtration in MBR.

α-Synuclein chaperone suppresses nucleation and amyloidogenesis of prion protein.

Protein misfolding diseases are a group of devastating disorders characterized by structural conversion of a soluble protein into an amyloid-like aggregate. Typically, the structural conversion occurs by misfolding of a single disease-associated protein, such as α-synuclein (αS) in Parkinson's disease, amyloid-ß in Alzheimer's disease, and prion protein (PrP) in transmissible spongiform encephalopathies (TSEs). However, accumulating evidence has implicated that cross-interactions between heterologous amyloidogenic proteins dramatically impact on amyloidogenesis and disease pathology. Here we show αS in a monomeric state can suppress amyloidogenesis of PrP in vitro. Thioflavin-T assays and transmission electron miscopy revealed that monomeric αS inhibits the nucleation step of amyloidogenesis without inhibiting the growing step. Surface plasmon resonance and co-sedimentation assays neither detected interaction between αS and monomeric PrP nor fibrillar PrP. These results suggested that αS suppress amyloidogenesis of PrP by binding to a transiently accumulated intermediate, such as a partially unfolded state. Moreover, we found that oligomeric αS, which was recently suggested to interact with PrP, also did not interact with PrP. Taken together, our study revealed a chaperon-like activity of αS against PrP amyloidogenesis, suggesting a possible involvement of αS in the pathology of TSEs.

Treatment enhances the prevalence of antibiotic-resistant bacteria and antibiotic resistance genes in the wastewater of Sri Lanka, and India.

Wastewater treatment plants (WWTPs) are being debated for being the hot spots for the development of antibiotic resistance in pathogenic microbial communities. We observed the prevalence of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARG), and multidrug resistance (MDR) in two municipal WWTPs and one hospital WWTP in Western and Southern Sri Lanka, and compared the results with particular reference to Indian and the World scenario to trace the imprints of treatment on ARB and ARG. Result suggests that although wastewater treatment resulted in higher than 1.06 log Escherichia coli (E. coli) reduction at all WWTPs, yet the percent of E. coli resistant to most of the antibiotics increased from influent to effluent. Higher prevalence of ARB, ARG, and MDR were noted in hospital WWTP owing to the higher antibiotic concentrations used and excreted by the patients. With reference to India, the WWTPs in Sri Lanka showed more ARB and a consistent increase in its percentages after the treatment but were less resistant to Fluoroquinolone (FQ). E. coli strains isolated from each location of both countries showed multidrug resistance, which has increased after the treatment and was strongly correlated with FQ in every WWTP. Resistant genes for Fluoroquinolone (FQ) (aac-(6')-1b-cr, qnrB, qnrS), ß-lactams (ampC), and sulphonamides (sul1) were common in all the wastewaters except additional parC gene in the hospital effluent of Sri Lanka, implying much higher resistance for quinolones, especially for Ciprofloxacin. Multivariate statistical treatments suggest that effluent showed higher loadings and association for MDR/ARB, where pH change and more extensive interaction with metals during the treatment processes seem to have profound effects.

Prevalence of antibiotic resistance in the tropical rivers of Sri Lanka and India.

We evaluate the imprints of urbanization, landuse and lifestyle on the prevalence and provenance of antibiotic resistance in the tropical rivers of Sri Lanka (Kelani and Gin) and India (Sabarmati, and Brahmaputra River). The prevalence of E. coli in the Kelani, Sabarmati, and Brahmaputra Rivers was in the range of 10-27, 267-76,600, and <50 CFU ml-1 respectively. Isolated E. coli colonies were subjected to six antibiotics to assess their resistance. We found higher resistance to old generation antibiotics like tetracycline (TC), and sulfamethoxazole (ST) transcends the resistance for fluoroquinolones like norfloxacin (NFX), ciprofloxacin (CIP), and levofloxacin (LVX). Interestingly, both Indian rivers had exhibited relatively higher resistance to TC and ST than the Kelani river or Gin River, implying that the Sri Lankan situation is relatively less critical. At genetic level the resistance for ß-lactams, fluoroquinolones and sulphonamides, were detected in many samples, as reported globally. While the resistance genes for aac-(6')-1b-cr, qnrS and sul1 were detected in both Sri Lankan and Indian Rivers, blaTEM and ampC were specific to the Indian Rivers only. Decoupling of the prevalence of metal contamination and antibiotic resistance has been noticed in India and Sri Lanka. Study implies that urbanization, landuse, and lifestyle (ULL) are the three most critical factors governing multidrug resistance (MDR) and fecal contamination.

Fate and seasonal change of Escherichia coli resistant to different antibiotic classes at each stage of conventional activated sludge process.

This study investigated the impact of each treatment stage of the activated sludge process on the fate of antibiotic resistant bacteria (ARB) in wastewater treatment plants (WWTPs). Wastewater and sludge samples were collected monthly at each stage of a commercial-scale WWTP. After 20-25 strains of indicator Escherichia coli were isolated from each sample on Chromocult Coliform Agar, antibiotic resistance of the isolates to amoxicillin (AMX), ciprofloxacin (CIP), norfloxacin (NFX), kanamycin (KM), sulfamethoxazole/trimethoprim (ST) and tetracycline (TC) were tested with the Kirby-Bauer disk diffusion method. As a result, activated sludge in the aeration tank and return sludge had higher abundance of antibiotic resistant E. coli than influent wastewater and secondary treatment effluent. AMX resistant E. coli was enriched in return sludge at the secondary clarifier. Higher temperature was also likely to cause an increase of AMX resistant E. coli in sludge. The antibiotic resistance profile of E. coli in secondary treatment effluent was more dependent on activated sludge than influent wastewater. These results suggested that activated sludge in WWTP possibly serves as a reservoir of ARB, and that behavior of ARB in WWTP differs by antibiotic classes.

Enhancement of methane production and phosphorus recovery with a novel pre-treatment of excess sludge using waste plaster board.

A new pre-treatment process for excess sludge is proposed to increase methane production and recover phosphorus by adding waste plaster board as calcium sulfate. The content of calcium sulfate in the plaster granules (PG) used in this study is 99%. When PG and calcium sulfate are added to the excess sludge generated from a municipal wastewater treatment plant, acetate production is enhanced as per sulfate reduction and phosphorus release is reduced via the formation of calcium phosphate. In the continuous pre-treatment experiment performed at 25 °C and for 10 days of sludge retention time (SRT) using calcium sulfate, 1935 ±â€¯395 mg/L of acetate is produced with 1070 ±â€¯255 mg/L of sulfate, which is reduced. Desulfobulbus spp., which can oxidize organic matter to acetate incompletely, have been observed in the pre-treated sludge. The pre-treated sludge has subsequently been used for methophiric anaerobic digestion. The methane yield from the pre-treated sludge is found to be 1.2 times that of the non-pretreated sludge at an SRT of 30 days, indicating that the pre-treatment using PG can improve methane production. Phosphorus is released from the non-pretreated sludge in the digester. Nevertheless, a decrease in phosphorus content has been observed, resulting in the digested sludge containing calcium phosphate that is useful for agriculture.

MicroRNA-143/Musashi-2/KRAS cascade contributes positively to carcinogenesis in human bladder cancer.

It has been well established that microRNA (miR)-143 is downregulated in human bladder cancer (BC). Recent precision medicine has shown that mutations in BC are frequently observed in FGFR3, RAS and PIK3CA genes, all of which correlate with RAS signaling networks. We have previously shown that miR-143 suppresses cell growth by inhibiting RAS signaling networks in several cancers including BC. In the present study, we showed that synthetic miR-143 negatively regulated the RNA-binding protein Musashi-2 (MSI2) in BC cell lines. MSI2 is an RNA-binding protein that regulates the stability of certain mRNAs and their translation by binding to the target sequences of the mRNAs. Of note, the present study clarified that MSI2 positively regulated KRAS expression through directly binding to the target sequence of KRAS mRNA and promoting its translation, thus contributing to the maintenance of KRAS expression. Thus, miR-143 silenced KRAS and MSI2, which further downregulated KRAS expression through perturbation of the MSI2/KRAS cascade.

Structural and functional characterization of fast-cycling RhoF GTPase.

Ras superfamily GTPases are molecular switches that cycle between GDP-bound inactive state and GTP-bound active state to control many signaling pathways. Emerging evidence suggests that several Ras superfamily GTPases, including RhoF, do not follow the classical GDP/GTP exchange cycle; they act as constitutively active GTP-bound proteins due to their fast activities of GDP/GTP exchange (termed as 'fast-cycling' GTPases). To understand the molecular basis of the fast-cycling GTPases, we generated a GTPase active recombinant RhoF and examined its function and structure. Two point mutations in the switch I/II regions (Q77L and P45S, corresponding to Q61L and P29S of Rac1) significantly reduced the GTPase activity of RhoF, suggesting a conserved mechanism of GTP hydrolysis between RhoF and other RAS superfamily GTPases. However, in contrary to the previous evidence, RhoF represented a slow GDP/GTP exchange activity that dissociates GDP very slowly on a day-to-week time scale, in our experiment using fluorescently labeled GDP. The slow GDP dissociation was accelerated by Mg2+ chelation and canonical fast-cycling mutations, F44L (corresponding to F28L of Rac1) and P45S. NMR and dynamic light scattering data revealed a multimeric structure of RhoF that can switch between different conformations depending on the GTP/GDP-bound state. Overall, our study suggests that (1) RhoF shares a conserved mechanism of GTP hydrolysis with other RAS superfamily GTPases, but (2) RhoF adopts a unique multimeric structure. Our study also argues that (3) the emerging concept of the fast-cycling GTPases for RhoF should be validated using an alternative assay that does not rely on fluorescently labeled GDP (251 words).

Evidence for a central role of PrP helix 2 in the nucleation of amyloid fibrils.

Amyloid fibrils are filamentous protein aggregates associated with the pathogenesis of a wide variety of human diseases. The formation of such aggregates typically follows nucleation-dependent kinetics, wherein the assembly and structural conversion of amyloidogenic proteins into oligomeric aggregates (nuclei) is the rate-limiting step of the overall reaction. In this study, we sought to gain structural insights into the oligomeric nuclei of the human prion protein (PrP) by preparing a series of deletion mutants lacking 14-44 of the C-terminal 107 residues of PrP and examined the kinetics and thermodynamics of these mutants in amyloid formation. An analysis of the experimental data using the concepts of the Φ-value analysis indicated that the helix 2 region (residues 168-196) acquires an amyloid-like ß-sheet during nucleation, whereas the other regions preserves a relatively disordered structure in the nuclei. This finding suggests that the helix 2 region serves as the nucleation site for the assembly of amyloid fibrils.-Honda, R., Kuwata, K. Evidence for a central role of PrP helix 2 in the nucleation of amyloid fibrils.

Role of the Disulfide Bond in Prion Protein Amyloid Formation: A Thermodynamic and Kinetic Analysis.

Prion diseases are associated with the structural conversion of prion protein (PrP) to a ß-sheet-rich aggregate, PrPSc. Previous studies have indicated that a reduction of the disulfide bond linking C179 and C214 of PrP yields an amyloidlike ß-rich aggregate in vitro. To gain mechanistic insights into the reduction-induced aggregation, here I characterized how disulfide bond reduction modulates the protein folding/misfolding landscape of PrP, by examining 1) the equilibrium stabilities of the native (N) and aggregated states relative to the unfolded (U) state, 2) the transition barrier separating the U and aggregated states, and 3) the final structure of amyloidlike misfolded aggregates. Kinetic and thermodynamic experiments revealed that disulfide bond reduction decreases the equilibrium stabilities of both the N and aggregated states by ∼3 kcal/mol, without changing either the amyloidlike aggregate structure, at least at the secondary structural level, or the transition barrier of aggregation. Therefore, disulfide bond reduction modulates the protein folding/misfolding landscape by entropically stabilizing disordered states, including the U and transition state of aggregation. This also indicates that the equilibrium stability of the N state, but not the transition barrier of aggregation, is the dominant factor determining the reduction-induced aggregation of PrP.

A valine-to-lysine substitution at position 210 induces structural conversion of prion protein into a ß-sheet rich oligomer.

Prion diseases are fatal neurodegenerative diseases associated with structural conversion of α-helical prion protein (PrP) into its ß-sheet rich isoform (PrPSc). Previous genetic analyses have indicated that several amino acid residues involved in the hydrophobic core of PrP (such as V180, F198, and V210) play a critical role in the development of prion diseases. To understand how these hydrophobic residues would contribute to the α-to-ß conversion process of PrP, we substituted the V210 residue with bulkier (V210F, V210I, and V210L), smaller (V210A), and charged amino acids (V210K) and characterized its effects. Interestingly, although most of the mutations had little or no effect on the biochemical properties of PrP, the V210K mutation induced structural conversion of PrP into a ß-structure. The ß-inducing effect was prominent and observed even under a physiological condition (i.e., in the absence of denaturant, acidic pH, reducing agent, and high temperature) in contrast to the disease-associated mutations in the PrP gene. We also examined structural features of V210K PrP using guanidine-hydrochloride unfolding, dynamic light scattering, 8-anilino-1-naphthalene sulfonate fluorescence, and electron microscopy, and revealed that V210K PrP assembles into a non-fibrillar ß-rich oligomer. Thus, the α-to-ß conversion can be induced by introduction of a charged residue into the hydrophobic core, which provide novel insight into the structural dynamics of PrP.

Amyloid-ß Peptide Induces Prion Protein Amyloid Formation: Evidence for Its Widespread Amyloidogenic Effect.

Transmissible spongiform encephalopathy is associated with misfolding of prion protein (PrP) into an amyloid ß-rich aggregate. Previous studies have indicated that PrP interacts with Alzheimer's disease amyloid-ß peptide (Aß), but it remains elusive how this interaction impacts on the misfolding of PrP. This study presents the first in vitro evidence that Aß induces PrP-amyloid formation at submicromolar concentrations. Interestingly, systematic mutagenesis of PrP revealed that Aß requires no specific amino acid sequences in PrP, and induces the misfolding of other unrelated proteins (insulin and lysozyme) into amyloid fibrils in a manner analogous to PrP. This unanticipated nonspecific amyloidogenic effect of Aß indicates that this peptide might be involved in widespread protein aggregation, regardless of the amino acid sequences of target proteins, and exacerbate the pathology of many neurodegenerative diseases.