Mapping long-range interactions in alpha-synuclein using spin-label NMR and ensemble molecular dynamics simulations

The intrinsically disordered protein alpha-synuclein plays a key role in the pathogenesis of Parkinson's disease (PD). We show here that the native state of alpha-synuclein consists of a broad distribution of conformers with an ensemble-averaged hydrodynamic radius significantly smaller than that expected for a random coil structure. This partial condensation is driven by interactions between the highly charged C-terminus and a large hydrophobic central region of the protein sequence. We suggest that this structure could inhibit the formation of alpha-synuclein aggregates, which are thought to be the cytotoxic species responsible for neurodegeneration in PD.     

Proteomics of Caenorhabditis elegans over-expressing human alpha-synuclein analyzed by fluorogenic derivatization-liquid chromatography/tandem mass spectrometry: identification of actin and several ribosomal proteins as negative markers at early Parkinson's disease stages

It has been known that the over-expression of alpha-synuclein, the main protein of Lewy bodies in Parkinson's disease (PD), leads to neurodegeneration in PD models. In this study, the changes in protein expression between the transgenic over-expressing human alpha-synuclein wild type (alpha-synWT) and the control Caenorhabditis elegans were elucidated by fluorogenic derivatization-liquid chromatography/tandem mass spectrometry (FD-LC-MS/MS) proteome analysis, which is a highly selective, sensitive, repeatable and quantitative method for protein identification. Because the alpha-synuclein wild-type worms showed moderate levels of dopamine loss without overt behavioral abnormalities, it was suggested that the changes in proteins in the alpha-synWT are related in the sequence of the formation of Lewy bodies. Among more than 400 protein peaks detected, actin and several ribosomal proteins were identified for the first time as negative markers at early PD stages. Actin was suggested to be one of the important targets in the elucidation of the etiology of neuronal diseases such as PD or other synucleinopathies.     

Targeting alpha-synuclein in Parkinson's disease

alpha-Synuclein aggregation into fibrils is associated with the pathogenesis of Parkinson's disease (PD). Li et al. provide strong evidence that rifampicin interacts with alpha-synuclein and inhibits its fibrillization. Rifampicin could be a promising candidate for therapeutic application for PD.     

Anti-amyloid nanoclusters for the treatment of brain hazards associated with incurable neurodegenerative diseases

Neurodegenerative diseases such as Parkinson's disease (PD) and Alzheimer's disease (AD) have no cure and pose a serious threat to human health. The accumulated amyloid has been the therapeutic target of various studies for over a decade, but there is a lack of effective treatments due to various limitations, such as the difficulty to cross the blood-brain barrier (BBB) and unfavorable bioaccumulation. To overcome these challenges, ultra-small metal nanoclusters (MNCs) (<2 nm) have emerged as promising new agents. Simple modifications of MNCs efficiently cross the BBB to reach the brain and dissociate amyloid fibrils into less toxic species. In addition, the enzymatic behavior of MNCs facilitates the scavenging of intracellular reactive oxygen species (ROS) and alleviates neuroinflammation. Herein, we summarize the reported anti-amyloid MNCs. Multiple promising functions of MNCs that may alleviate the harms of neurodegenerative diseases are exhibited. The physicochemical properties that influence the inhibition and degradation of common amyloid fibrils, including alpha-synuclein (α-syn) and amyloid beta-peptide (Aβ) are discussed. The prospect of optimizing MNCs to suppress more harm in the brain is presented to facilitate the development of practical therapies for neurodegenerative diseases.     

In silico drug repositioning of FDA-approved drugs to predict new inhibitors for alpha-synuclein aggregation

One of the hallmarks of Parkinson’s disease (PD), a long-term neurodegenerative syndrome, is the accumulation of alpha-synuclein (α-syn) fibrils. Despite numerous studies and efforts, inhibition of α-syn protein aggregation is still a challenge. To overcome this issue, we propose an in silico pharmacophore-based repositioning strategy, to find a pharmaceutical drug that, in addition to their defined role, can be used to prevent aggregation of the α-syn protein. Ligand-based pharmacophore modeling was developed and the best model was selected with validation parameters including 72 % sensitivity, 98 % specificity and goodness score about 0.7. The optimal model has three groups of hydrogen bond donor (HBD), three groups of hydrogen bond acceptor (HBA), and two aromatic rings (AR). The FDA-Approved reports in the ZINC15 database were screened with the pharmacophore model taken from inhibitor compounds. The model identified 22 hits, as promising candidate drugs for Parkinson's therapy. It is noteworthy that among these, 10 drugs have been reported to inhibition of α-syn aggregation or treat/reduce Parkinson's pathogenesis. This model was used to virtual screen ZINC, NCI databases, and natural products from the pomegranate. The results of this screen were filtered for their inability to cross the blood-brain barrier, poor oral bioavailability, etc. Finally, the selected compounds of two ZINC and NCI databases were combined and structurally clustered. Remained compounds were clustered in 28 different clusters, and the 17 compounds were introduced as final candidates.     

Bioinorganic chemistry of Parkinson's disease: structural determinants for the copper-mediated amyloid formation of alpha-synuclein

The aggregation of alpha-synuclein (AS) is a critical step in the etiology of Parkinson's disease (PD). A central, unresolved question in the pathophysiology of PD relates to the role of AS-metal interactions in amyloid fibril formation and neurodegeneration. Our previous works established a hierarchy in alpha-synuclein-metal ion interactions, where Cu(II) binds specifically to the protein and triggers its aggregation under conditions that might be relevant for the development of PD. Two independent, non-interacting copper-binding sites were identified at the N-terminal region of AS, with significant difference in their affinities for the metal ion. In this work we have solved unknown details related to the structural binding specificity and aggregation enhancement mediated by Cu(II). The high-resolution structural characterization of the highest affinity N-terminus AS-Cu(II) complex is reported here. Through the measurement of AS aggregation kinetics we proved conclusively that the copper-enhanced AS amyloid formation is a direct consequence of the formation of the AS-Cu(II) complex at the highest affinity binding site. The kinetic behavior was not influenced by the His residue at position 50, arguing against an active role for this residue in the structural and biological events involved in the mechanism of copper-mediated AS aggregation. These new findings are central to elucidate the mechanism through which the metal ion participates in the fibrillization of AS and represent relevant progress in the understanding of the bioinorganic chemistry of PD.     

“双碳”背景下能源关键元素暴露的神经效应研究进展

为了实现“双碳”目标,能源关键元素(Energy-critical elements, ECEs)在全球范围内的应用将显著增加,增加了人群暴露风险,但是其健康效应尚未明确。ECEs主要为金属元素,如锂、钴及稀土元素。为了解金属ECEs的潜在健康效应,重点关注其神经危害或风险,为其在可持续发展过程中可能产生的健康问题提出干预策略。回顾了与金属ECEs相关的毒理学、职业安全和健康问题以及流行病学调查的文献报道,主要在分子和细胞水平、实验动物和人群流行病学研究等方面对上述金属ECEs神经效应的综述。目前,金属ECEs健康效应的研究面临一些挑战：现有的人群研究数量有限,亟需更多的研究为建立有关健康、环境影响和安全的监测系统提供科学依据,为绿色能源行业建立一个可持续、安全和健康的职业环境,助力实现“双碳”目标。     

Rifampicin inhibits alpha-synuclein fibrillation and disaggregates fibrils

The aggregation of alpha-synuclein in dopaminergic neurons of the substantia nigra is a critical step in the pathogenesis of Parkinson's disease. We show that the antibiotic rifampicin inhibited alpha-synuclein fibrillation and disaggregated existing fibrils in a concentration-dependent manner. Size-exclusion chromatography data indicated that rifampicin stabilized alpha-synuclein as both a monomer and soluble oligomers comprised of partially folded alpha-synuclein. Experiments using aged samples of rifampicin indicated that the most active species in inhibiting fibrillation and disaggregating fibrils is an oxidation product of rifampicin, which was confirmed in experiments under anaerobic conditions. These results indicate that rifampicin-mediated inhibition of alpha-synuclein fibrillation and disaggregation of fibrils involves preferential stabilization of monomeric and soluble oligomeric forms, and that rifampicin potentially may have therapeutic application for Parkinson's disease.     

In Silico Molecular Docking Analysis of Karanjin against Alzheimer’s and Parkinson’s Diseases as a Potential Natural Lead Molecule for New Drug Design,Development and Therapy

Parkinson’s disease (PD) and Alzheimer’s disease (AD) are neurodegenerative disorders that have emerged as among the serious health problems of the 21st century. The medications currently available to treat AD and PD have limited efficacy and are associated with side effects. Natural products are one of the most vital and conservative sources of medicines for treating neurological problems. Karanjin is a furanoflavonoid, isolated mainly from Pongamia pinnata with several medicinal plants, and has been reported for numerous health benefits. However, the effect of karanjin on AD and PD has not yet been systematically investigated. To evaluate the neuroprotective effect of karanjin, extensive in silico studies starting with molecular docking against five putative targets for AD and four targets for PD were conducted. The findings were compared with three standard drugs using Auto Dock 4.1 and Molegro Virtual Docker software. Additionally, the physiochemical properties (Lipinski rule of five), drug-likeness and parameters including absorption, distribution, metabolism, elimination and toxicity (ADMET) profiles of karanjin were also studied. The molecular dynamics (MD) simulations were performed with two selective karanjin docking complexes to analyze the dynamic behaviors and binding free energy at 100 ns time scale. In addition, frontier molecular orbitals (FMOs) and density-functional theory (DFT) were also investigated from computational quantum mechanism perspectives using the Avogadro-ORCA 1.2.0 platform. Karanjin complies with all five of Lipinski’s drug-likeness rules with suitable ADMET profiles for therapeutic use. The docking scores (kcal/mol) showed comparatively higher potency against AD and PD associated targets than currently used standard drugs. Overall, the potential binding affinity from molecular docking, static thermodynamics feature from MD-simulation and other multiparametric drug-ability profiles suggest that karanjin could be considered as a suitable therapeutic lead for AD and PD treatment. Furthermore, the present results were strongly correlated with the earlier study on karanjin in an Alzheimer’s animal model. However, necessary in vivo studies, clinical trials, bioavailability, permeability and safe dose administration, etc. must be required to use karanjin as a potential drug against AD and PD treatment, where the in silico results are more helpful to accelerate the drug development.     

Changes in interfacial properties of alpha-synuclein preceding its aggregation

Parkinson's disease (PD) is associated with the formation and deposition of amyloid fibrils of the protein alpha-synuclein (AS). It has been proposed that oligomeric intermediates on the pathway to fibrilization rather than the fibrils themselves are the pathogenic agents of PD, but efficient methods for their detection are lacking. We have studied the interfacial properties of wild-type AS and the course of its aggregation in vitro using electrochemical analysis and dynamic light scattering. The oxidation signals of tyrosine residues of AS at carbon electrodes and the ability of fibrils to adsorb and catalyze hydrogen evolution at hanging mercury drop electrodes (HMDEs) decreased during incubation. HMDEs were particularly sensitive to pre-aggregation changes in AS. Already after 1 h of a standard aggregation assay in vitro (stirring at 37 degrees C), the electrocatalytic peak H increased greatly and shifted to less negative potentials. Between 3 and 9 h of incubation, an interval during which dynamic light scattering indicated AS oligomerization, peak H diminished and shifted to more negative potentials, and AS adsorbability decreased. We tentatively attribute the very early changes in the interfacial behavior of the protein after the first few hours of incubation to protein destabilization with disruption of long-range interactions. The subsequent changes can be related to the onset of oligomerization. Our results demonstrate the utility of electrochemical methods as new and simple tools for the investigation of amyloid formation.     

Quantum dots protect against MPP+-induced neurotoxicity in a cell model of Parkinson’s disease through autophagy induction

Autophagy is a basic cellular process that decomposes damaged organelles and aberrant proteins. Dysregulation of autophagy is implicated in pathogenesis of neurodegenerative disorders, including Parkinson's disease(PD). Pharmacological compounds that stimulate autophagy can provide neuroprotection in models of PD. Nanoparticles have emerged as regulators of autophagy and have been tested in adjuvant therapy for diseases. In this present study, we explore the effects of quantum dots(QDs) that can induce autophagy in a cellular model of Parkinson's disease. Cd Te/Cd S/Zn S QDs protect differentiated rat pheochromocytoma PC12 cells from MPP+-induced cell damage, including reduced viability, apoptosis and accumulation of α-Synuclein, a characteristic protein of PD. The protective function of QDs is autophagy-dependent. In addition, we investigate the interaction between quantum dots and autophagic pathways and identify beclin1 as an essential factor for QDs-induced autophagy. Our results reveal new promise of QDs in the theranostic of neurodegenerative diseases.     

The effect of cations on nucleophilic additions to carbonyl compounds: carbonyl complexation control versus ionic association control. application to t

The fundamental influence of cations in nucleophilic additions to carbonyl compounds is discussed in terms of either carbonyl complexation or ionic association with the nucleophile. Since loose ion pairs react essentially under complexation control while tight ion pairs react under association control, an experimental criterion is proposed to determine which factor controls the reactivity (complexation or association) based on the kinetic effects of cryptand or crown-ether addition.An application of this duality is then examined for the regioselectivity of nucleophilic additions to α-enones and correlated with perturbational arguments based on abinitio calculations. Under complexation control, attack at carbon 2 is favoured, especially when a soft nucleophile is involved and the counter-ion is Li⁺ rather than Na⁺. Under association control two cases are considered according to whether the cation is directly bound or not to the nucleophilic center; in addition, the influence of hard or soft metals and the possibility of a cationic bridge in the transition state have been discussed. Several kinetic controlled reactions have been examined and interpreted in this way. An extension of this concept to other reactions will be considered.     

Photoinduced elimination of senescent microglia cells in vivo by chiral gold nanoparticles

Parkinson''s disease (PD) is an age-related neurodegenerative disease, and the removal of senescent cells has been proved to be beneficial for improving age-associated pathologies in neurodegeneration disease. In this study, chiral gold nanoparticles (NPs) with different helical directions were synthesized to selectively induce the apoptosis of senescent cells under light illumination. By modifying anti-B2MG and anti-DCR2 antibodies, senescent microglia cells could be cleared by chiral NPs without damaging the activities of normal cells under illumination. Notably, l-P⁺ NPs exhibited about a 2-fold higher elimination efficiency than d-P⁻ NPs for senescent microglia cells. Mechanistic studies revealed that the clearance of senescent cells was mediated by the activation of the Fas signaling pathway. The in vivo injection of chiral NPs successfully confirmed that the elimination of senescent microglia cells in the brain could further alleviate the symptoms of PD mice in which the alpha-synuclein (α-syn) in cerebrospinal fluid (CFS) decreased from 83.83 ± 4.76 ng mL⁻¹ to 8.66 ± 1.79 ng mL⁻¹ after two months of treatment. Our findings suggest a potential strategy to selectively eliminate senescent cells using chiral nanomaterials and offer a promising strategy for alleviating PD.

The apoptosis pathways of senescent microglia cells induced by chiral NPs under the irradiation of 808 nm laser in the brain of PD mice.     

Ultraviolet B Radiation of Human Skin Generates Platelet-activating Factor Receptor Agonists

Ultraviolet B radiation (UVB) is a potent stimulator of epidermal cytokine production. In addition to cytokines, such as tumor necrosis factor-alpha (TNF-α), UVB generates bioactive lipids including platelet-activating factor (PAF). Our previous in vitro studies in keratinocytes or epithelial cell lines have demonstrated that UVB-mediated production of PAF agonists is due primarily to the pro-oxidative effects of this stimulant, resulting in the nonenzymatic production of modified phosphocholines (oxidized glycerophosphocholines). The current studies use human skin to assess whether UVB irradiation generates PAF-receptor agonists, and the role of oxidative stress in their production. These studies demonstrate that UVB irradiation of human skin results in PAF agonists, which are blocked by the antioxidant vitamin C and the epidermal growth factor receptor inhibitor PD168393. Inasmuch as UVB-generated PAF agonists have been implicated in animal model systems as being involved in photobiologic processes including systemic immunosuppression and cytokine (TNF-α) production, these studies indicate that this novel activity could be involved in human disease.     

A hyphenated flow-through analytical system for the study of the mobility and fractionation of trace and major elements in environmental solid samples

A flow-through hyphenated analytical method has been tested that enables not only the accelerated and efficient fractionation of trace elements (TE) species in environmental solids to be achieved but allows real-time studies on the leaching process to be made. Rotating coiled columns (RCC), earlier used mainly in countercurrent chromatography, have been successfully applied to the dynamic fractionation of heavy metals in soil, sediment, and sludge samples. A ground solid sample (about 0.5 g) was retained in a PTFE rotating column as the stationary phase whereas different aqueous eluents, chosen according to recent data on the selectivity of leachants, were continuously pumped through. Elements were determined in the effluent on-line by inductively coupled plasma atomic emission spectrometry (ICP-AES). Since the flow rates used in the RCC are in good agreement with those needed for cross-flow nebulization in the ICP-AES spectrometer, both devices were coupled directly without any additional interface systems. Simultaneous investigation of the elution profiles of trace and major elements has made it possible to study the elements association in separated fractions and hence to prove the efficacy of extractants and their selectivity toward targeted mineralogical phases of samples. The close association of heavy metals with Mn oxides in the sediment and sludge samples was confirmed. The time-resolved dissolution of different organic complexes of metals was observed for the sediment sample. It was found that in sediment and sludge samples the dynamics of iron release under the action of Tamm's reagent is somewhat different from that of aluminium. In addition, the proposed method can also be applied to develop effective leaching schemes and in the analysis of environmental solids for risk assessment of their contaminants addressed to water quality and bioavailability.     

Alpha-synuclein misfolding and neurodegenerative diseases

Alpha-synuclein is an abundant presynaptic brain protein, misfolding, aggregation and fibrillation of which are implicated as critical factors in several neurodegenerative diseases. The list of the well-known synucleinopathies includes such devastating disorders as Parkinson's disease, Lewy body variant of Alzheimer's disease, diffuse Lewy body disease, dementia with Lewy bodies, multiple system atrophy, and neurodegeneration with brain iron accumulation type I. The precise functions of alpha-synuclein remain elusive, but there are evidence indicating its involvement in regulation vesicular release and/or turnover and synaptic function in the central nervous system. It might play a role in neuronal plasticity responses, bind fatty acids, regulate certain enzymes, transporters, and neurotransmitter vesicles, be involved in neuronal survival and even can act as a molecular chaperone. Structurally, alpha-synuclein is an illustrative member of the rapidly growing family of natively unfolded (or intrinsically disordered) proteins and considerable knowledge has been accumulated about its structural properties and conformational behavior. The molecular mechanisms underlying misfolding, aggregation and fibrillation of alpha-synuclein and the role of various environmental and genetic factors in stimulation and inhibition of these processes are relatively well understood. Here, the main structural features of alpha-synuclein, its functions, and involvement in various human diseases are summarized providing a foundation for better understanding of the biochemistry, biophysics and neuropathology of alpha-synuclein aggregation.     

Spermine binding to Parkinson's protein alpha-synuclein and its disease-related A30P and A53T mutants

Aggregation of alpha-synuclein (alpha-syn), a protein implicated in Parkinson's disease (PD), is believed to progress through formation of a partially folded intermediate. Using nanoelectrospray ionization (nano-ESI) mass spectrometry combined with ion mobility measurements we found evidence for a highly compact partially folded family of structures for alpha-syn and its disease-related A53T mutant with net charges of -6, -7, and -8. For the other early onset PD mutant, A30P, this highly compact population was only evident when the protein had a net charge of -6. When bound to spermine near physiologic pH, all three proteins underwent a charge reduction from the favored solution charge state of -10 to a net charge of -6. This charge reduction is accompanied by a dramatic size reduction of about a factor of 2 (cross section of 2600 A2 (-10 charge state) down to 1430 A2 (-6 charge state)). We conclude that spermine increases the aggregation rate of alpha-syn by inducing a collapsed conformation, which then proceeds to form aggregates.     

Development and application of methods using SPE,HPLC-DAD,LC-ESI-MS/MS and GFAAS for the determination of herbicides and metals in surface and drinking water

A preliminary study of the pollution in surface and drinking waters caused by herbicides and metals in the Municipal Water Supply System (CORSAN) in Rio Grande city, RS, Brazil, is reported. The occurrence of 5 herbicides and 9 metals was studied in surface and drinking water through the analysis of 2 sampling spots at CORSAN. The analytical determination was performed by solid-phase extraction (SPE), high performance liquid chromatography-photodiode array detection (HPLC-DAD) and liquid chromatography coupled to electrospray ionisation tandem mass spectrometry (LC-ESI-MS/MS) for herbicides, and graphite furnace atomic absorption spectrometry (GFAAS) for metals. The concentrations of herbicides in the surface water were very low; however, the herbicide clomazone was detected in more than 50% of the samples analysed in concentration exceeding 1.0?µg L^?1. The concentration of metals was below the Maximum Contaminant Level (MCL) set by the Brazilian regulation.     

Site-specific interactions of Cu(II) with alpha and beta-synuclein: bridging the molecular gap between metal binding and aggregation

The aggregation of alpha-synuclein (AS) is a critical step in the etiology of Parkinson's disease (PD) and other neurodegenerative synucleinopathies. Protein-metal interactions play a critical role in AS aggregation and might represent the link between the pathological processes of protein aggregation and oxidative damage. Our previous studies established a hierarchy in AS-metal ion interactions, where Cu(II) binds specifically to the protein and triggers its aggregation under conditions that might be relevant for the development of PD. In this work, we have addressed unresolved structural details related to the binding specificity of Cu(II) through the design of site-directed and domain-truncated mutants of AS and by the characterization of the metal-binding features of its natural homologue beta-synuclein (BS). The structural properties of the Cu(II) complexes were determined by the combined application of nuclear magnetic resonance, electron paramagnetic resonance, UV-vis, circular dichroism spectroscopy, and matrix-assisted laser desorption ionization mass spectrometry (MALDI MS). Two independent, noninteracting copper-binding sites with significantly different affinities for the metal ion were detected in the N-terminal regions of AS and BS. MALDI MS provided unique evidence for the direct involvement of Met1 as the primary anchoring residue for Cu(II) in both proteins. Comparative spectroscopic analysis of the two proteins allowed us to deconvolute the Cu(II) binding modes and unequivocally assign the higher-affinity site to the N-terminal amino group of Met1 and the lower-affinity site to the imidazol ring of the sole His residue. Through the use of competitive chelators, the affinity of the first equivalent of bound Cu(II) was accurately determined to be in the submicromolar range for both AS and BS. Our results prove that Cu(II) binding in the C-terminal region of synucleins represents a nonspecific, very low affinity process. These new insights into the bioinorganic chemistry of PD are central to an understanding of the role of Cu(II) in the fibrillization process of AS and have implications for the molecular mechanism by which BS might inhibit AS amyloid assembly.