Glutaminyl cyclase-mediated toxicity of pyroglutamate-beta amyloid induces striatal neurodegeneration

Background Posttranslational modifications of beta amyloid (Aβ) have been shown to affect its biophysical and neurophysiological properties. One of these modifications is N-terminal pyroglutamate (pE) formation. Enzymatic glutaminyl cyclase (QC) activity catalyzes cyclization of truncated Aβ(3-x), generating pE3-Aβ. Compared to unmodified Aβ, pE3-Aβ is more hydrophobic and neurotoxic. In addition, it accelerates aggregation of other Aβ species. To directly investigate pE3-Aβ formation and toxicity in vivo, transgenic (tg) ETNA (E at the truncated N-terminus of Aβ) mice expressing truncated human Aβ(3–42) were generated and comprehensively characterized. To further investigate the role of QC in pE3-Aβ formation in vivo, ETNA mice were intercrossed with tg mice overexpressing human QC (hQC) to generate double tg ETNA-hQC mice. Results Expression of truncated Aβ(3–42) was detected mainly in the lateral striatum of ETNA mice, leading to progressive accumulation of pE3-Aβ. This ultimately resulted in astrocytosis, loss of DARPP-32 immunoreactivity, and neuronal loss at the sites of pE3-Aβ formation. Neuropathology in ETNA mice was associated with behavioral alterations. In particular, hyperactivity and impaired acoustic sensorimotor gating were detected. Double tg ETNA-hQC mice showed similar Aβ levels and expression sites, while pE3-Aβ were significantly increased, entailing increased astrocytosis and neuronal loss. Conclusions ETNA and ETNA-hQC mice represent novel mouse models for QC-mediated toxicity of truncated and pE-modified Aβ. Due to their significant striatal neurodegeneration these mice can also be used for analysis of striatal regulation of basal locomotor activity and sensorimotor gating, and possibly for DARPP-32-dependent neurophysiology and neuropathology. The spatio-temporal correlation of pE3-Aβ and neuropathology strongly argues for an important role of this Aβ species in neurodegenerative processes in these models

Optimizing the composition of a synthetic cellulosome complex for the hydrolysis of softwood pulp: identification of the enzymatic core functions and biochemical complex characterization

Abstract Background The development of efficient cellulase blends is a key factor for cost-effectively valorizing biomass in a new bio-economy. Today, the enzymatic hydrolysis of plant-derived polysaccharides is mainly accomplished with fungal cellulases, whereas potentially equally effective cellulose-degrading systems from bacteria have not been developed. Particularly, a thermostable multi-enzyme cellulase complex, the cellulosome from the anaerobic cellulolytic bacterium Clostridium thermocellum is promising of being applied as cellulolytic nano-machinery for the production of fermentable sugars from cellulosic biomass. Results In this study, 60 cellulosomal components were recombinantly produced in E. coli and systematically permuted in synthetic complexes to study the function–activity relationship of all available enzymes on Kraft pulp from pine wood as the substrate. Starting from a basic exo/endoglucanase complex, we were able to identify additional functional classes such as mannanase and xylanase for optimal activity on the substrate. Based on these results, we predicted a synthetic cellulosome complex consisting of seven single components (including the scaffoldin protein and a β-glucosidase) and characterized it biochemically. We obtained a highly thermostable complex with optimal activity around 60–65 °C and an optimal pH in agreement with the optimum of the native cellulosome (pH 5.8). Remarkably, a fully synthetic complex containing 47 single cellulosomal components showed comparable activity with a commercially available fungal enzyme cocktail on the softwood pulp substrate. Conclusions Our results show that synthetic bacterial multi-enzyme complexes based on the cellulosome of C. thermocellum can be applied as a versatile platform for the quick adaptation and efficient degradation of a substrate of interest

Glutaminyl cyclase-mediated toxicity of pyroglutamate-beta amyloid induces striatal neurodegeneration

Background Posttranslational modifications of beta amyloid (Aβ) have been shown to affect its biophysical and neurophysiological properties. One of these modifications is N-terminal pyroglutamate (pE) formation. Enzymatic glutaminyl cyclase (QC) activity catalyzes cyclization of truncated Aβ(3-x), generating pE3-Aβ. Compared to unmodified Aβ, pE3-Aβ is more hydrophobic and neurotoxic. In addition, it accelerates aggregation of other Aβ species. To directly investigate pE3-Aβ formation and toxicity in vivo, transgenic (tg) ETNA (E at the truncated N-terminus of Aβ) mice expressing truncated human Aβ(3–42) were generated and comprehensively characterized. To further investigate the role of QC in pE3-Aβ formation in vivo, ETNA mice were intercrossed with tg mice overexpressing human QC (hQC) to generate double tg ETNA-hQC mice. Results Expression of truncated Aβ(3–42) was detected mainly in the lateral striatum of ETNA mice, leading to progressive accumulation of pE3-Aβ. This ultimately resulted in astrocytosis, loss of DARPP-32 immunoreactivity, and neuronal loss at the sites of pE3-Aβ formation. Neuropathology in ETNA mice was associated with behavioral alterations. In particular, hyperactivity and impaired acoustic sensorimotor gating were detected. Double tg ETNA-hQC mice showed similar Aβ levels and expression sites, while pE3-Aβ were significantly increased, entailing increased astrocytosis and neuronal loss. Conclusions ETNA and ETNA-hQC mice represent novel mouse models for QC-mediated toxicity of truncated and pE-modified Aβ. Due to their significant striatal neurodegeneration these mice can also be used for analysis of striatal regulation of basal locomotor activity and sensorimotor gating, and possibly for DARPP-32-dependent neurophysiology and neuropathology. The spatio-temporal correlation of pE3-Aβ and neuropathology strongly argues for an important role of this Aβ species in neurodegenerative processes in these models

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Additional file 4. Overview of 47 recombinant proteins. All proteins were mixed in equimolar amounts before adding them to the pentavalent SKLMY complex

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Additional file 3. Effect of inhibitors on the cellulosomalcomplexes. The glucose concentration was measured using the Glu-HK determination kit (Megazyme). The residual complex activity was assessed using 0.5 mL standard reaction mixture containing 0.25% (w/v) of the substrate for one to 2 days of incubation at 60 °C. The initial activity at time point 0 corresponds to 100% relative activity

MOESM1 of Optimizing the composition of a synthetic cellulosome complex for the hydrolysis of softwood pulp: identification of the enzymatic core functions and biochemical complex characterization

Additional file 1. Oligonucleotides used in this study

MOESM2 of Optimizing the composition of a synthetic cellulosome complex for the hydrolysis of softwood pulp: identification of the enzymatic core functions and biochemical complex characterization

Additional file 2. SDS-PAGE summary of all recombinantly expressed dockerin type I containing proteins used in this study (60 different components of C. thermocellum cellulosome). Each protein is shown after the purification process, including the final heat precipitation step. Enzyme names and Clo1313 numbers are shown above the corresponding SDS-PAGE picture. Molecular weight standards are depicted in kDa. Proteins marked with asterisks are truncated versions of the original protein

Massacre de la rue Transnonain : 15 avril 1834, lithographie publiée dans L'Association mensuelle de juillet&nbsp1834

Épreuve sur blanc provenant du dépôt légal, 29 x 44,5 cm. Delteil 135.Les mouvements populaires se succèdent depuis l’avènement de Louis-Philippe, roi des Français, en août 1830. Le malaise économique et les mesures anti-ouvrières prises par Thiers, ministre de l‘Intérieur, contribuent à les provoquer. L’agitation est également entretenue par des associations républicaines comme la Société des Droits de l’homme. En février 1834, afin d’affaiblir la propagande républicaine, le gouvernement fait voter une série de lois qui réglementent de manière drastique l’activité des crieurs publics et des vendeurs de journaux ambulants et interdisent les associations politiques. Le 9 avril 1834, la Société des Droits de l’homme organise une manifestation à Lyon. Au même moment se tient le procès de canuts accusés de coalition et de grève. Des coups de feu sont tirés sur la foule des manifestants. S’en suit une semaine de combats meurtriers (plus de 300 morts, près de 600 blessés). Le 13 avril, l’insurrection s’étend à Paris, notamment dans le quartier du Marais. Le 14 au matin, elle est réprimée dans le sang. Les soldats envahissent un immeuble situé au 12 rue Transnonain (l’actuelle rue Beaubourg) et massacrent tous les habitants qui pourtant n’avaient pas pris part aux émeutes. De cette bavure policière que Philipon nomme "la boucherie de la rue Transnonain", Daumier tire un chef-d'œuvre de l’estampe, simple constat de la vérité, aussi sobre que dramatique. De caricaturiste, Daumier se hissait au rang de peintre d'histoire en noir et blanc et devançait le courant réaliste en peinture.téléchargeabl

Glutaminyl cyclases as novel targets for the treatment of septic arthritis

Background. Septic arthritis is a severe and rapidly debilitating disease mainly caused by Staphylococcus aureus. Here, we assess the antiarthritic efficiency of glutaminyl cyclase (QC) inhibitors. Methods. Mice were inoculated with an arthritogenic amount of S. aureus intravenously or by local administration into the knee joint. Animals were treated with QC inhibitors (PBD155 and PQ529) via chow during the experiment. QC and isoQC knockout mice were also analyzed for arthritis symptoms after local administration of bacteria. Results. Both QC inhibitors significantly delayed the onset of clinical signs of arthritis, and inhibitors significantly decreased weight loss in treated animals. Following intraarticular injection of S. aureus, PBD155-treated mice had lower levels of synovitis and bone erosion, as well as less myeloperoxidase in synovial tissue. Fluorescence-activated cell sorter analysis revealed that PBD155 treatment affected the expression pattern of adhesion molecules, preventing the upregulation of cells expressing CD11b/CD18. Conclusion. The compounds investigated here represent a novel class of small molecular antiarthritic inhibitors. In our studies, they exerted strong antiinflammatory actions, and therefore they might be suited for disease-modifying treatment of infectious arthritis

Overexpression of Glutaminyl Cyclase, the Enzyme Responsible for Pyroglutamate Aβ Formation, Induces Behavioral Deficits, and Glutaminyl Cyclase Knock-out Rescues the Behavioral Phenotype in 5XFAD Mice*

Pyroglutamate-modified Aβ (AβpE3–42) peptides are gaining considerable attention as potential key players in the pathology of Alzheimer disease (AD) due to their abundance in AD brain, high aggregation propensity, stability, and cellular toxicity. Overexpressing AβpE3–42 induced a severe neuron loss and neurological phenotype in TBA2 mice. In vitro and in vivo experiments have recently proven that the enzyme glutaminyl cyclase (QC) catalyzes the formation of AβpE3–42. The aim of the present work was to analyze the role of QC in an AD mouse model with abundant AβpE3–42 formation. 5XFAD mice were crossed with transgenic mice expressing human QC (hQC) under the control of the Thy1 promoter. 5XFAD/hQC bigenic mice showed significant elevation in TBS, SDS, and formic acid-soluble AβpE3–42 peptides and aggregation in plaques. In 6-month-old 5XFAD/hQC mice, a significant motor and working memory impairment developed compared with 5XFAD. The contribution of endogenous QC was studied by generating 5XFAD/QC-KO mice (mouse QC knock-out). 5XFAD/QC-KO mice showed a significant rescue of the wild-type mice behavioral phenotype, demonstrating the important contribution of endogenous mouse QC and transgenic overexpressed QC. These data clearly demonstrate that QC is crucial for modulating AβpE3–42 levels in vivo and prove on a genetic base the concept that reduction of QC activity is a promising new therapeutic approach for AD