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Small molecules that bind to oligomeric protein species such as membrane proteins and fibrils are of clinical interest for development of therapeutics and diagnostics. Definition of the binding site at atomic resolution via NMR is often challenging due to low binding stoichiometry of the small molecule. For fibrils and aggregation intermediates grown in the presence of lipids, we report atomic-resolution contacts to the small molecule at sub nm distance via solid-state NMR using dynamic nuclear polarization (DNP) and orthogonally labelled samples of the protein and the small molecule. We apply this approach to α-synuclein (αS) aggregates in complex with the small molecule anle138b, which is a clinical drug candidate for disease modifying therapy. The small central pyrazole moiety of anle138b is detected in close proximity to the protein backbone and differences in the contacts between fibrils and early intermediates are observed. For intermediate species, the 100 K condition for DNP helps to preserve the aggregation state, while for both fibrils and oligomers, the DNP enhancement is essential to obtain sufficient sensitivity.

The publication contains the documents previously not introduced into scientific circulation, revealing both the view on the police guard of the Vyatka province of its auditor, the Russian statesman, Lieutenant General Baron G. P. von Medem, and the changes in police guard organization that occurred over the three years of the institute. In the first document, the gendarme describes the police guard of the Vyatka provinces, as the auditor of which he acted during the First Russian Revolution. The second document presents the results of the revision of the Vyatka guard in October-December 1909. The introductory article highlights the historical, source study and archaeographic issues associated with the published documents. The sources are published in accordance with the current «Rules for the Publication of Historical Documents» (1990).

Introduction. The object of this study is the police reform of 1862. The subject is the course of its implementation in the Ural region. The purpose of the article is to analyze the transformations carried out in the general police of the Ural provinces in the 1860s. The geographical scope of the study is limited to Vyatka, Orenburg, and Perm provinces. Methods and materials. The modernization theory was chosen as an average level methodology. The methods of the research are historical-genetic, historical-comparative and historical-systemic. Besides, the discursive approach was chosen to understand the sources of the research better. Analysis. By the law of 1862, city police was united with county police under the authority of a single leader called county police officer. Only Vyatka, Orenburg, Perm, Ufa and Yekaterinburg retained independent city police. The police staff introduced by the law of 1862 was insufficient, primarily due to the liquidation of the police of the Ministry of Finance and the police of the Military Ministry in factory districts and mines. The real process of strengthening the Ural police lasted for years or even decades. The Mining Department tried to maintain some degree of control over the police in the region. Results. It was concluded that in the confrontation with the Mining Department for the full control over the police of the region, the provincial authorities managed to win an almost complete victory. At the same time the provincial authorities underestimated the need to professionalize the police, often giving preference to Cossack units.

The article examines the lifepath, teaching and scientific activities of A. V. Shilov, Candidate of historical sciences, associate professor of the Department of Ancient and Modern History of Russia at the Perm State National Research University. The article observes an assessment of his contribution to the study of the Ural region and to historical science in general. A complete list of his scientific works is provided as an appendix to the article.

There is an urgent clinical need for imaging of α‐synuclein (αSyn) fibrils, the hallmark biomarker for Parkinson's disease, in neurodegenerative disorders. Despite immense efforts, promising tracer candidates for nuclear imaging of αSyn are rare. Diphenyl pyrazoles are known modulators of αSyn aggregation and thus bear potential for non‐invasive detection of this biomarker in vivo. Here we demonstrate high‐affinity binding of the family member anle253b to fibrillar αSyn and present a high‐yielding site‐selective radiosynthesis route for 11C radiolabeling using in‐situ generated [11C]formaldehyde and reductive methylation. Radio‐HPLC of the tracer after incubation with rat serum in vitro shows excellent stability of the molecule. Positron emission tomography in healthy animals is used to assess the pharmacokinetics and biodistribution of the tracer, showing good penetration of the blood–brain barrier and low background binding to the non‐pathological brain., High binding, low background: Development of radiotracers for α‐synuclein has proven challenging. Herein we describe a radiolabeling strategy for the aggregation modulator anle253b (2) and demonstrate its successful penetration of the blood‐brain barrier in healthy rats as well as its high‐affinity binding to aggregated α‐synuclein in vitro.

Purpose Deposition of misfolded alpha-synuclein (αSYN) aggregates in the human brain is one of the major hallmarks of synucleinopathies. However, a target-specific tracer to detect pathological aggregates of αSYN remains lacking. Here, we report the development of a positron emission tomography (PET) tracer based on anle138b, a compound shown to have therapeutic activity in animal models of neurodegenerative diseases. Methods Specificity and selectivity of [3H]MODAG-001 were tested in in vitro binding assays using recombinant fibrils. After carbon-11 radiolabeling, the pharmacokinetic and metabolic profile was determined in mice. Specific binding was quantified in rats, inoculated with αSYN fibrils and using in vitro autoradiography in human brain sections of Lewy body dementia (LBD) cases provided by the Neurobiobank Munich (NBM). Results [3H]MODAG-001 revealed a very high affinity towards pure αSYN fibrils (Kd = 0.6 ± 0.1 nM) and only a moderate affinity to hTau46 fibrils (Kd = 19 ± 6.4 nM) as well as amyloid-β1–42 fibrils (Kd = 20 ± 10 nM). [11C]MODAG-001 showed an excellent ability to penetrate the mouse brain. Metabolic degradation was present, but the stability of the parent compound improved after selective deuteration of the precursor. (d3)-[11C]MODAG-001 binding was confirmed in fibril-inoculated rat striata using in vivo PET imaging. In vitro autoradiography showed no detectable binding to aggregated αSYN in human brain sections of LBD cases, most likely, because of the low abundance of aggregated αSYN against background protein. Conclusion MODAG-001 provides a promising lead structure for future compound development as it combines a high affinity and good selectivity in fibril-binding assays with suitable pharmacokinetics and biodistribution properties.

Parkinson’s disease (PD) represents an increasing problem in society. The oligomerization of alpha-synuclein (αSyn) is a suggested key event in its pathogenesis, yet the pathological modes of action remain to be fully elucidated. To identify potential disease-modifying therapeutics and to study αSyn-mediated toxic mechanisms, we established cell lines with inducible overexpression of different αSyn constructs: αSyn, αSyn coupled to the fluorescence protein Venus (αSyn-Venus), and αSyn coupled to the N-terminal or C-terminal part of Venus (V1S and SV2, respectively) for a bimolecular fluorescence complementation assay (BiFC). Inducibility was achieved by applying modified GAL4-UAS or Cre-loxP systems and addition of tebufenozide or 4-OH-tamoxifen, respectively. Expression constructs were stably integrated into the host genome of H4 neuroglioma cells by lentiviral transduction. We here demonstrate a detailed investigation of the expression characteristics of inducible H4 cells showing low background expression and high inducibility. We observed increased protein load and aggregation of αSyn upon incubation with DMSO and FeCl3 along with an increase in cytotoxicity. In summary, we present a system for the creation of inducibly αSyn-overexpressing cell lines holding high potential for the screening for modulators of αSyn aggregation and αSyn-mediated toxicity.

Parkinson's disease (PD) and Alzheimer's disease (AD) are common neurodegenerative disorders of the elderly and, therefore, affect a growing number of patients worldwide. Both diseases share, as a common hallmark, the accumulation of characteristic protein aggregates, known as Lewy bodies (LB) in PD, and neurofibrillary tangles in AD. LBs are primarily composed of misfolded α-synuclein (aSyn), and neurofibrillary tangles are primarily composed of tau protein. Importantly, upon pathological evaluation, most AD and PD/Lewy body dementia cases exhibit mixed pathology, with the co-occurrence of both LB and neurofibrillary tangles, among other protein inclusions. Recent studies suggest that both aSyn and tau pathology can spread and propagate through neuronal connections. Therefore, it is important to investigate the mechanisms underlying aggregation and propagation of these proteins for the development of novel therapeutic strategies. Here, we assessed the effects of different pharmacological interventions on the aggregation and internalization of tau and aSyn. We found that anle138b and fulvic acid decrease aSyn and tau aggregation, that epigallocatechin gallate decreases aSyn aggregation, and that dynasore reduces tau internalization. Establishing the effects of small molecules with different chemical properties on the aggregation and spreading of aSyn and tau will be important for the development of future therapeutic interventions.

Parkinson’s disease (PD) and Alzheimer’s disease (AD) are common neurodegenerative disorders of the elderly and, therefore, affect a growing number of patients worldwide. Both diseases share, as a common hallmark, the accumulation of characteristic protein aggregates, known as Lewy bodies (LB) in PD, and neurofibrillary tangles in AD. LBs are primarily composed of misfolded α-synuclein (aSyn), and neurofibrillary tangles are primarily composed of tau protein. Importantly, upon pathological evaluation, most AD and PD/Lewy body dementia cases exhibit mixed pathology, with the co-occurrence of both Lewy bodies and neurofibrillary tangles, among other protein inclusions. Recent studies suggest that both aSyn and tau pathology can spread and propagate through neuronal connections. Therefore, it is important to investigate the mechanisms underlying aggregation and propagation of these proteins for the development of novel therapeutic strategies. Here, we assessed the effects of different pharmacological interventions on the aggregation and internalization of tau and aSyn. We found that anle138b and epigallocatechin gallate decrease aSyn aggregation, that fulvic acid attenuates aggregation of the repeat domain of tau, and that dynasore reduces tau internalization. Establishing the effects of small molecules with different chemical properties on the aggregation and spreading of aSyn and tau will be important for the development of future therapeutic interventions.

BACKGROUND: MSA is a fatal neurodegenerative disease characterized by autonomic failure and severe motor impairment. Its main pathological hallmark is the accumulation of alpha-synuclein in oligodendrocytes, leading to glial and neuronal dysfunction and neurodegeneration. These features are recapitulated in the PLP-halphaSyn mouse model expressing human alpha-synuclein in oligodendrocytes. At present, there is no effective disease-modifying therapy. Previous experiments have shown that the aggregation inhibitor, anle138b, reduces neurodegeneration and behavioral deficits in mouse models of other proteinopathies. OBJECTIVES: To test the therapeutic potential of anle138b in a mouse model of MSA. METHODS: Two-month-old PLP-halphaSyn mice were fed over a period of 4 months with pellets containing anle138b at two different doses (0.6 and 2 g/kg) and compared to healthy controls and PLP-halphaSyn mice fed with placebo pellets. At the end of the treatment, behavioral and histological analyses were performed. RESULTS: We observed a reversal of motor function to healthy control levels when PLP-halphaSyn mice were treated with both doses of anle138b. Histological and molecular analyses showed a significant reduction in alpha-synuclein oligomers and glial cytoplasmic inclusions in animals fed with anle138b compared to nontreated mice. These animals also present preservation of dopaminergic neurons and reduction in microglial activation in SN correlating with the alpha-synuclein reduction observed. CONCLUSIONS: Anle138b reduces alpha-synuclein accumulation in PLP-halphaSyn mice, leading to neuroprotection, reduction of microglial activation, and preservation of motor function supporting the use of anle138b in a future clinical trial for MSA. (c) 2018 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

Background Recently diphenyl-pyrazole (DPP) compounds and especially anle138b were found to reduce the aggregation of α-synuclein or Tau protein in vitro as well as in a mouse model of neurodegenerative diseases [1,2]. Direct interaction of the DPPs with the fibrillar structure was identified by fluorescence spectroscopy. Thereby a strong dependence of the fluorescence on the surroundings could be identified [3]. Methods Stationary and time-resolved emission experiments were performed on DPP compounds substituted by different halogens. Results The compounds reveal a pronounced dependence of the fluorescence on the surrounding solvent. In non-polar solvents they show strong emission in the blue part of the spectrum while in polar and proton donating solvents, such as water or acetic acid a dual fluorescence can be observed where a red-shifted emission points to a charge transfer in the excited state with large dipole moment. Non-radiative processes including photochemical reactions are observed for DPP substituted with heavy halogens. Upon binding of anle138b and its derivatives to protein fibrils in aqueous buffer, strong enhancement of the fluorescence at short wavelengths is found. Conclusion The investigations of the DPPs in different surroundings lead to a detailed model of the fluorescence characteristics. We propose a model for the binding in fibrils of different proteins, where the DPP is located in a hydrophobic groove independent of the specific sequence of the amino acids. General significance These investigations characterize the binding site of the DPP anle138b in protein aggregates and contribute to the understanding of the therapeutic mode of action of this compound.

Parkinson’s disease (PD) is characterized by the presence of α-synuclein aggregates known as Lewy bodies and Lewy neurites, whose formation is linked to disease development. The causal relation between α-synuclein aggregates and PD is not well understood. We generated a new transgenic mouse line (MI2) expressing human, aggregation-prone truncated 1–120 α-synuclein under the control of the tyrosine hydroxylase promoter. MI2 mice exhibit progressive aggregation of α-synuclein in dopaminergic neurons of the substantia nigra pars compacta and their striatal terminals. This is associated with a progressive reduction of striatal dopamine release, reduced striatal innervation and significant nigral dopaminergic nerve cell death starting from 6 and 12 months of age, respectively. In the MI2 mice, alterations in gait impairment can be detected by the DigiGait test from 9 months of age, while gross motor deficit was detected by rotarod test at 20 months of age when 50% of dopaminergic neurons in the substantia nigra pars compacta are lost. These changes were associated with an increase in the number and density of 20–500 nm α-synuclein species as shown by dSTORM. Treatment with the oligomer modulator anle138b, from 9 to 12 months of age, restored striatal dopamine release, prevented dopaminergic cell death and gait impairment. These effects were associated with a reduction of the inner density of large α-synuclein aggregates and an increase in dispersed small α-synuclein species as revealed by dSTORM. The MI2 mouse model recapitulates the progressive dopaminergic deficit observed in PD, showing that early synaptic dysfunction is associated to fine behavioral motor alterations, precedes dopaminergic axonal loss and neuronal death that become associated with a more consistent motor deficit upon reaching a certain threshold. Our data also provide new mechanistic insight for the effect of anle138b’s function in vivo supporting that targeting α-synuclein aggregation is a promising therapeutic approach for PD. Electronic supplementary material The online version of this article (10.1007/s00401-019-02023-x) contains supplementary material, which is available to authorized users.

Background Augmenting the brain clearance of toxic oligomers with small molecule modulators constitutes a promising therapeutic concept against tau deposition. However, there has been no test of this concept in animal models of Alzheimer’s disease (AD) with initiation at a late disease stage. Thus, we aimed to investigate the effects of interventional late-stage Anle138b treatment, which previously indicated great potential to inhibit oligomer accumulation by binding of pathological aggregates, on the metabolic decline in transgenic mice with established tauopathy in a longitudinal 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) study. Methods Twelve transgenic mice expressing all six human tau isoforms (hTau) and ten controls were imaged by FDG-PET at baseline (14.5 months), followed by randomization into Anle138b treatment and vehicle groups for 3 months. FDG-PET was repeated after treatment for 3 months, and brains were analyzed by tau immunohistochemistry. Longitudinal changes of glucose metabolism were compared between study groups, and the end point tau load was correlated with individual FDG-PET findings. Results Tau pathology was significantly ameliorated by late-stage Anle138b treatment when compared to vehicle (frontal cortex − 53%, p

Misfolding and aggregate formation by the tau protein has been closely related with neurotoxicity in a large group of human neurodegenerative disorders, which includes Alzheimer's disease. Here, we investigate the membrane-active properties of tau oligomers on mitochondrial membranes, using minimalist in vitro model systems. Thus, exposure of isolated mitochondria to oligomeric tau evoked a disruption of mitochondrial membrane integrity, as evidenced by a combination of organelle swelling, efflux of cytochrome c and loss of the mitochondrial membrane potential. Tau-induced mitochondrial dysfunction occurred independently of the mitochondrial permeability transition (mPT) pore complex. Notably, mitochondria were rescued by pre-incubation with 10-N-nonyl acridine orange (NAO), a molecule that specifically binds cardiolipin (CL), the signature phospholipid of mitochondrial membranes. Additionally, NAO prevented direct binding of tau oligomers to isolated mitochondria. At the same time, tau proteins exhibited high affinity to CL-enriched membranes, whilst permeabilisation of lipid vesicles also strongly correlated with CL content. Intriguingly, using single-channel electrophysiology, we could demonstrate the formation of non-selective ion-conducting tau nanopores exhibiting multilevel conductances in mito-mimetic bilayers. Taken together, the data presented here advances a scenario in which toxic cytosolic entities of tau protein would target mitochondrial organelles by associating with their CL-rich membrane domains, leading to membrane poration and compromised mitochondrial structural integrity.

SUMMARYParkinson’s Disease (PD) is characterized by the presence of α-synuclein aggregates known as Lewy bodies and Lewy neurites, whose formation is linked to disease development. The causal relation between α-synuclein aggregates and PD is not well understood. We generated a new transgenic mouse line (MI2) expressing human, aggregation-prone truncated 1-120 α-synuclein under the control of the tyrosine hydroxylase promoter. MI2 mice exhibit progressive aggregation of α-synuclein in dopaminergic neurons of the substantia nigra pars compacta and their striatal terminals. This is associated with a progressive reduction of striatal dopamine release, reduced striatal innervation and significant nigral dopaminergic nerve cell death starting from 6 and 12 months of age, respectively. Overt impairment in motor behavior was found in MI2 mice at 20 months of age, when 50% of dopaminergic neurons are lost. These changes were associated with an increase in the number and density of 20-500nm α-synuclein species as shown by dSTORM. Treatment with the oligomer modulator anle138b, from 9-12 months of age, restored striatal dopamine release and prevented dopaminergic cell death. These effects were associated with a reduction of the inner density of α-synuclein aggregates and an increase in dispersed small α-synuclein species as revealed by dSTORM. The MI2 mouse model recapitulates the progressive dopaminergic deficit observed in PD, showing that early synaptic dysfunction precedes dopaminergic axonal loss and neuronal death that become associated with a motor deficit upon reaching a certain threshold. Our data also provide new mechanistic insight for the effect of anle138b’s function in vivo supporting that targeting α-synuclein aggregation is a promising therapeutic approach for PD.

Alzheimer’s disease is a devastating neurodegenerative disease eventually leading to dementia. An effective treatment does not yet exist. Here we show that oral application of the compound anle138b restores hippocampal synaptic and transcriptional plasticity as well as spatial memory in a mouse model for Alzheimer’s disease, when given orally before or after the onset of pathology. At the mechanistic level, we provide evidence that anle138b blocks the activity of conducting Ab pores without changing the membrane embedded Ab-oligomer structure. In conclusion, our data suggest that anle138b is a novel and promising compound to treat AD-related pathology that should be investigated further., peer-reviewed

Our skin, our belongings, the world surrounding us, and the environment we live in are covered with molecular traces. Detecting and characterizing these molecular traces is necessary to understand the environmental impact on human health and disease, and to decipher complex molecular interactions between humans and other species, particularly microbiota. We recently introduced 3D molecular cartography for mapping small organic molecules (including metabolites, lipids, and environmental molecules) found on various surfaces, including the human body. Here, we provide a protocol and open-source software for 3D molecular cartography. The protocol includes step-by-step procedures for sample collection and processing, liquid chromatography-mass spectrometry (LC-MS)-based metabolomics, quality control (QC), molecular identification using MS/MS, data processing, and visualization with 3D models of the sampled environment. The LC-MS method was optimized for a broad range of small organic molecules. We enable scientists to reproduce our previously obtained results, and illustrate the broad utility of our approach with molecular maps of a rosemary plant and an ATM keypad after a PIN code was entered. To promote reproducibility, we introduce cartographical snapshots: files that describe a particular map and visualization settings, and that can be shared and loaded to reproduce the visualization. The protocol enables molecular cartography to be performed in any mass spectrometry laboratory and, in principle, for any spatially mapped data. We anticipate applications, in particular, in medicine, ecology, agriculture, biotechnology, and forensics. The protocol takes 78 h for a molecular map of 100 spots, excluding the reagent setup.

Pathological tau aggregation leads to filamentous tau inclusions and characterizes neurodegenerative tauopathies such as Alzheimer’s disease and frontotemporal dementia and parkinsonism linked to chromosome 17. Tau aggregation coincides with clinical symptoms and is thought to mediate neurodegeneration. Transgenic mice overexpressing mutant human P301S tau exhibit many neuropathological features of human tauopathies including behavioral deficits and increased mortality. Here, we show that the di-phenyl-pyrazole anle138b binds to aggregated tau and inhibits tau aggregation in vitro and in vivo. Furthermore, anle138b treatment effectively ameliorates disease symptoms, increases survival time and improves cognition of tau transgenic PS19 mice. In addition, we found decreased synapse and neuron loss accompanied by a decreased gliosis in the hippocampus. Our results suggest that reducing tau aggregates with anle138b may represent an effective and promising approach for the treatment of human tauopathies. Electronic supplementary material The online version of this article (doi:10.1007/s00401-015-1483-3) contains supplementary material, which is available to authorized users.

Recent epidemiological and clinical studies have reported a significantly increased risk for melanoma in people with Parkinson's disease. Because no evidence could be obtained that genetic factors are the reason for the association between these two diseases, we hypothesized that of the three major Parkinson's disease-related proteins-α-synuclein, LRRK2, and Parkin-α-synuclein might be a major link. Our data, presented here, demonstrate that α-synuclein promotes the survival of primary and metastatic melanoma cells, which is the exact opposite of the effect that α-synuclein has on dopaminergic neurons, where its accumulation causes neuronal dysfunction and death. Because this detrimental effect of α-synuclein on neurons can be rescued by the small molecule anle138b, we explored its effect on melanoma cells. We found that treatment with anle138b leads to massive melanoma cell death due to a major dysregulation of autophagy, suggesting that α-synuclein is highly beneficial to advanced melanoma because it ensures that autophagy is maintained at a homeostatic level that promotes and ensures the cell's survival.

Natural product screening programs have uncovered molecules from diverse natural sources with various biological activities and unique structures. However, much is yet underexplored and additional information is hidden in these exceptional collections. We applied untargeted mass spectrometry approaches to capture the chemical space and dispersal patterns of metabolites from an in-house library of marine cyanobacterial and algal collections. Remarkably, 86% of the metabolomics signals detected were not found in other available datasets of similar nature, supporting the hypothesis that marine cyanobacteria and algae possess distinctive metabolomes. The data were plotted onto a world map representing eight major sampling sites, and revealed potential geographic locations with high chemical diversity. We demonstrate the use of these inventories as a tool to explore the diversity and distribution of natural products. Finally, we utilized this tool to guide the isolation of a new cyclic lipopeptide, yuvalamide A, from a marine cyanobacterium. DOI: http://dx.doi.org/10.7554/eLife.24214.001, eLife digest Cyanobacteria and algae are found in all oceans around the globe. Like plants, they can use sunlight as a source of energy in a process called photosynthesis. As a result, these organisms are important sources of oxygen and another vital nutrient called nitrogen for other marine organisms. Many of these organisms also produce a variety of other chemicals known as “natural products” to help them to survive in their environments. Some of these natural products have shown potential as medicinal drugs. The search for new chemicals with useful medicinal properties has led researchers to collect samples of algae and cyanobacteria from various locations around the world. An approach called mass spectrometry is often used to identify new chemicals because it can provide information about the structure of a molecule based on how much its fragments weigh. Luzzatto-Knaan et al. used mass spectrometry to search for new chemicals in samples of algae and cyanobacteria that had been collected by diving and snorkeling in a wide variety of tropical marine environments over several decades. The experiments reveal that the organisms in these samples produce a diverse range of chemicals, most of which were previously unknown and have not been found in other similar environmental collections. The data were grouped together into eight major collection areas covering different parts of the tropics. The samples from some areas contained a wider variety of chemicals than others. Within each collection area, some molecules were found to be very common whereas others were only present at specific locations. To highlight the distribution of these natural products, Luzzatto-Knaan et al. display the data on a world map. Further experiments used this approach as a guide to extract a previously unknown chemical called yuvalamide A from a marine cyanobacterium. The next challenge would be to associate the geographical patterns of chemicals to their potential ecological roles. This approach offers a new way to explore large-scale collections of environmental samples to discover and study new natural products. DOI: http://dx.doi.org/10.7554/eLife.24214.002

Proteins are inserted into the bacterial plasma membrane cotranslationally after translating ribosomes are targeted to the translocon in the membrane via the signal recognition particle (SRP) pathway. The targeting pathway involves an interaction between SRP and the SRP receptor, FtsY. Here we focus on the role of FtsY and its interaction with the translocon in controlling targeting. We show that in unbound FtsY the NG and A domains interact with one another. The interaction involves the membrane-targeting region at the junction between A and N domain. The closed form of FtsY is impaired in binding to SRP. Upon binding to the phospholipid-embedded translocon the domains of FtsY move apart. This enhances the docking of the FtsY NG domain to the homologous NG domain of the SRP protein Ffh. Thus, FtsY binding to the translocon has a central role in orchestrating the formation of a quaternary transfer complex in which the nascent peptide is transferred to the translocon. We propose that FtsY activation at the translocon ensures that ribosome-SRP complexes are directed to available translocons. This way sequestering SRP in futile complexes with unbound FtsY can be avoided and efficient targeting to the translocon achieved.

Prion diseases are fatal neurodegenerative diseases characterized by accumulation of the pathogenic prion protein PrP in the brain. We established quantitative real-time quaking-induced conversion for the measurement of minute amounts of PrP in body fluids such as urine. Using this approach, we monitored the efficacy of antiprion therapy by quantifying the seeding activity of PrP from the brain and urine of mice after prion infection. We found that the aggregation inhibitor anle138b decreased the levels of PrP in the brain and urine. Importantly, variations of PrP levels in the urine closely corresponded to those in the brain. Our findings indicate that quantification of urinary PrP enables measurement of prion disease progression in body fluids and can substitute for immunodetection in brain tissue. We expect PrP quantification biologic fluids (such as urine and cerebrospinal fluid) with quantitative real-time quaking-induced conversion to emerge as a valuable noninvasive diagnostic tool for monitoring disease progression and the efficacy of therapeutic approaches in animal studies and human clinical trials of prion diseases. Moreover, highly sensitive methods for quantifying pathologic aggregate seeds might provide novel molecular biomarkers for other neurodegenerative diseases that may involve prion-like mechanisms (protein aggregation and spreading), such as Alzheimer disease and Parkinson disease.

Background Special diphenyl-pyrazole compounds and in particular anle138b were found to reduce the progression of prion and Parkinson's disease in animal models. The therapeutic impact of these compounds was attributed to the modulation of α-synuclein and prion-protein aggregation related to these diseases. Methods Photophysical and photochemical properties of the diphenyl-pyrazole compounds anle138b, anle186b and sery313b and their interaction with monomeric and aggregated α-synuclein were studied by fluorescence techniques. Results The fluorescence emission of diphenyl-pyrazole is strongly increased upon incubation with α-synuclein fibrils, while no change in fluorescence emission is found when brought in contact with monomeric α-synuclein. This points to a distinct interaction between diphenyl-pyrazole and the fibrillar structure with a high binding affinity (Kd = 190 ± 120 nM) for anle138b. Several α-synuclein proteins form a hydrophobic binding pocket for the diphenyl-pyrazole compound. A UV-induced dehalogenation reaction was observed for anle138b which is modulated by the hydrophobic environment of the fibrils. Conclusion Fluorescence of the investigated diphenyl-pyrazole compounds strongly increases upon binding to fibrillar α-synuclein structures. Binding at high affinity occurs to hydrophobic pockets in the fibrils. General significance The observed particular fluorescence properties of the diphenyl-pyrazole molecules open new possibilities for the investigation of the mode of action of these compounds in neurodegenerative diseases. The high binding affinity to aggregates and the strong increase in fluorescence upon binding make the compounds promising fluorescence markers for the analysis of aggregation-dependent epitopes.

]. parkinson’s disease (pD) is characterized by the deposition of aggregated alpha-synuclein (asyn). recent evidence suggests that oligomers formed in the aggregation process constitute the main toxic species causing neurodegeneration. recently, we reported that the oligomer modulator “anle138b” [3-(1,3-benzodioxol-5-yl)-5-(3-bromophenyl)-1H-pyrazole] is capable of prolonging the survival of prion-infected mice and of various animal models of pD [5blocked formation and accumulation of asyn oligomers in the brain, reduced disease-associated motor deficits, and led to prolonged disease-free survival [5 ]. These findings support the following

In neurodegenerative diseases such as Alzheimer’s disease (AD), Parkinson’s disease (PD) and prion diseases, deposits of aggregated disease-specific proteins are found. Oligomeric aggregates are presumed to be the key neurotoxic agent. Here we describe the novel oligomer modulator anle138b [3-(1,3-benzodioxol-5-yl)-5-(3-bromophenyl)-1H-pyrazole], an aggregation inhibitor we developed based on a systematic high-throughput screening campaign combined with medicinal chemistry optimization. In vitro, anle138b blocked the formation of pathological aggregates of prion protein (PrPSc) and of α-synuclein (α-syn), which is deposited in PD and other synucleinopathies such as dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). Notably, anle138b strongly inhibited all prion strains tested including BSE-derived and human prions. Anle138b showed structure-dependent binding to pathological aggregates and strongly inhibited formation of pathological oligomers in vitro and in vivo both for prion protein and α-synuclein. Both in mouse models of prion disease and in three different PD mouse models, anle138b strongly inhibited oligomer accumulation, neuronal degeneration, and disease progression in vivo. Anle138b had no detectable toxicity at therapeutic doses and an excellent oral bioavailability and blood–brain-barrier penetration. Our findings indicate that oligomer modulators provide a new approach for disease-modifying therapy in these diseases, for which only symptomatic treatment is available so far. Moreover, our findings suggest that pathological oligomers in neurodegenerative diseases share structural features, although the main protein component is disease-specific, indicating that compounds such as anle138b that modulate oligomer formation by targeting structure-dependent epitopes can have a broad spectrum of activity in the treatment of different protein aggregation diseases. Electronic supplementary material The online version of this article (doi:10.1007/s00401-013-1114-9) contains supplementary material, which is available to authorized users.

Aggregation of islet amyloid polypeptide (IAPP) in the human pancreas is linked to the pathogenesis of type-II diabetes. Interactions of IAPP with membranes may play an important role in the disease, because membranes may catalyze IAPP aggregation and because amyloid formation of IAPP results in membrane permeabilization, and hence cytotoxicity [1]. In order to control this cytotoxicity, there is an intense focus on the development of amyloid inhibitors. Here, we studied the interaction of a novel amyloid inhibitor “anle145c” [2,3,4] on IAPP aggregation in the absence and presence of model membranes using a range of biophysical techniques. Our results show that in both cases the compound acts as an efficient inhibitor, but that it has a distinctly different mode of action in the presence of membranes. Importantly, similar results were obtained with the structurally related natural inhibitor EGCG but with less efficiency. Our complementary experimental approaches reveal that the inhibitors target a different species in the absence and presence of membranes. We propose a model to explain our findings and we discuss its implications for in-vivo studies on these inhibitors.[1] Engel M et al., PNAS. (2008) - 105:6033.[2] Wagner J et al., Acta Neuropathologica (2013) - 125:795.[3] Levin J et al., Acta Neuropathologica (2014) - 127:779.[4] Wagner J et al., Acta Neuropathologica in press (2015) “Reducing tau aggregates with anle138b delays disease progression in a mouse model of tauopathies”.

Amyloid formation in the pancreas by islet amyloid polypeptide (IAPP) is closely associated with type-2 diabetes. Compelling evidence indicates that membranes play a crucial role in contributing to IAPP amyloid formation and that IAPP amyloid formation leads to cell membrane disruption [1]. Since both IAPP amyloid formation and membrane damage are considered perilous to the pancreatic beta-cells, their inhibition may be an effective strategy for the prevention and/ or treatment of the disease. Here, we studied the interaction between a novel amyloid inhibitor “anle145c” [2] and IAPP in the absence and presence of model membranes. Our results suggest that addition of anle145c to IAPP inhibits IAPP fibril growth even at sub-stoichiometric concentrations, both in the absence and presence of membranes. Anle145c also reduces membrane damage induced by IAPP but does not induce membrane leakage by itself. Most notably, anle145c shows a strong membrane interaction, resulting in a distinctly different mode of inhibition than in the absence of membranes. We present a model in which anle145c interacts with oligomeric IAPP species in solution, but with monomeric or early oligomeric IAPP species in the presence of membranes.[1] Engel M et al., PNAS. (2008) - 105 :6033[2] Wagner J et al., Acta Neuropathologica (2013) - 125: 795