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Auswahl der wissenschaftlichen Literatur zum Thema „Type IV filaments“
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Zeitschriftenartikel zum Thema "Type IV filaments"
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Ching, GY, und RK Liem. „Assembly of type IV neuronal intermediate filaments in nonneuronal cells in the absence of preexisting cytoplasmic intermediate filaments“. Journal of Cell Biology 122, Nr. 6 (15.09.1993): 1323–35. http://dx.doi.org/10.1083/jcb.122.6.1323.
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We report here on the in vivo assembly of alpha-internexin, a type IV neuronal intermediate filament protein, in transfected cultured cells, comparing its assembly properties with those of the neurofilament triplet proteins (NF-L, NF-M, and NF-H). Like the neurofilament triplet proteins, alpha-internexin coassembles with vimentin into filaments. To study the assembly characteristics of these proteins in the absence of a preexisting filament network, transient transfection experiments were performed with a non-neuronal cell line lacking cytoplasmic intermediate filaments. The results showed that only alpha-internexin was able to self-assemble into extensive filamentous networks. In contrast, the neurofilament triplet proteins were incapable of homopolymeric assembly into filamentous arrays in vivo. NF-L coassembled with either NF-M or NF-H into filamentous structures in the transfected cells, but NF-M could not form filaments with NF-H. alpha-internexin could coassemble with each of the neurofilament triplet proteins in the transfected cells to form filaments. When all but 2 and 10 amino acid residues were removed from the tail domains of NF-L and NF-M, respectively, the resulting NF-L and NF-M deletion mutants retained the ability to coassemble with alpha-internexin into filamentous networks. These mutants were also capable of forming filaments with other wild-type neurofilament triplet protein subunits. These results suggest that the tail domains of NF-L and NF-M are dispensable for normal coassembly of each of these proteins with other type IV intermediate filament proteins to form filaments.
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Goosens, Vivianne J., Andreas Busch, Michaella Georgiadou, Marta Castagnini, Katrina T. Forest, Gabriel Waksman und Vladimir Pelicic. „Reconstitution of a minimal machinery capable of assembling periplasmic type IV pili“. Proceedings of the National Academy of Sciences 114, Nr. 25 (06.06.2017): E4978—E4986. http://dx.doi.org/10.1073/pnas.1618539114.
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Type IV pili (Tfp), which are key virulence factors in many bacterial pathogens, define a large group of multipurpose filamentous nanomachines widespread in Bacteria and Archaea. Tfp biogenesis is a complex multistep process, which relies on macromolecular assemblies composed of 15 conserved proteins in model gram-negative species. To improve our limited understanding of the molecular mechanisms of filament assembly, we have used a synthetic biology approach to reconstitute, in a nonnative heterologous host, a minimal machinery capable of building Tfp. Here we show that eight synthetic genes are sufficient to promote filament assembly and that the corresponding proteins form a macromolecular complex at the cytoplasmic membrane, which we have purified and characterized biochemically. Our results contribute to a better mechanistic understanding of the assembly of remarkable dynamic filaments nearly ubiquitous in prokaryotes.
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Couturier, Marc Roger, und Markus Stein. „Helicobacter pylori produces unique filaments upon host contact in vitro“. Canadian Journal of Microbiology 54, Nr. 7 (Juli 2008): 537–48. http://dx.doi.org/10.1139/w08-042.
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Helicobacter pylori exists in 2 distinct morphological states, helicoid and coccoid. Both have been observed in in vitro culture and in gastric biopsies. We visualized H. pylori during AGS cell infections using immunofluorescence microscopy. Anti-H. pylori mouse serum as well as human serum from H. pylori-positive patients recognized long, thin bacterial filaments, which formed on helicoids and more frequently on coccoids. These filaments reached lengths of 59 μm and often connected bacteria. Periodate oxidation abolished antibody recognition, suggesting that carbohydrates compose a major antigenic component of the filaments. Similar to results obtained using immunofluorescence microscopy, scanning electron microscopy imaging revealed thin filamentous structures, which were absent on uninfected cells. Both coccoid conversion and filament development increased over the time course of infection with peak filament formation at 4 h. The number of visible filaments then decreased as bacteria clustered on the apical surface of AGS cells. Since the observed filaments were clearly distinct from previously described surface structures, including flagella and the cag type IV secretion system, our results demonstrate that these filaments represent a unique, previously unrecognized, organelle.
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Kraljic, Katarina, Christopher Duckworth, Rita Tojeiro, Shadab Alam, Dmitry Bizyaev, Anne-Marie Weijmans, Nicholas Fraser Boardman und Richard R. Lane. „SDSS-IV MaNGA: 3D spin alignment of spiral and S0 galaxies“. Monthly Notices of the Royal Astronomical Society 504, Nr. 3 (21.04.2021): 4626–33. http://dx.doi.org/10.1093/mnras/stab1109.
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ABSTRACT We investigate the 3D spin alignment of galaxies with respect to the large-scale filaments using the MaNGA survey. The cosmic web is reconstructed from the Sloan Digital Sky Survey using disperse and the 3D spins of MaNGA galaxies are estimated using the thin disc approximation with integral field spectroscopy kinematics. Late-type spiral galaxies are found to have their spins parallel to the closest filament’s axis. The alignment signal is found to be dominated by low-mass spirals. Spins of S0-type galaxies tend to be oriented preferentially in perpendicular direction with respect to the filament’s axis. This orthogonal orientation is found to be dominated by S0s that show a notable misalignment between their kinematic components of stellar and ionized gas velocity fields and/or by low-mass S0s with lower rotation support compared to their high-mass counterparts. Qualitatively similar results are obtained when splitting galaxies based on the degree of ordered stellar rotation, such that galaxies with high spin magnitude have their spin aligned, and those with low spin magnitude in perpendicular direction to the filaments. In the context of conditional tidal torque theory, these findings suggest that galaxies’ spins retain memory of their larger scale environment. In agreement with measurements from hydrodynamical cosmological simulations, the measured signal at low redshift is weak, yet statistically significant. The dependence of the spin-filament orientation of galaxies on their stellar mass, morphology, and kinematics highlights the importance of sample selection to detect the signal.
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Braun, Tatjana, Matthijn R. Vos, Nir Kalisman, Nicholas E. Sherman, Reinhard Rachel, Reinhard Wirth, Gunnar F. Schröder und Edward H. Egelman. „Archaeal flagellin combines a bacterial type IV pilin domain with an Ig-like domain“. Proceedings of the National Academy of Sciences 113, Nr. 37 (30.08.2016): 10352–57. http://dx.doi.org/10.1073/pnas.1607756113.
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The bacterial flagellar apparatus, which involves ∼40 different proteins, has been a model system for understanding motility and chemotaxis. The bacterial flagellar filament, largely composed of a single protein, flagellin, has been a model for understanding protein assembly. This system has no homology to the eukaryotic flagellum, in which the filament alone, composed of a microtubule-based axoneme, contains more than 400 different proteins. The archaeal flagellar system is simpler still, in some cases having ∼13 different proteins with a single flagellar filament protein. The archaeal flagellar system has no homology to the bacterial one and must have arisen by convergent evolution. However, it has been understood that the N-terminal domain of the archaeal flagellin is a homolog of the N-terminal domain of bacterial type IV pilin, showing once again how proteins can be repurposed in evolution for different functions. Using cryo-EM, we have been able to generate a nearly complete atomic model for a flagellar-like filament of the archaeon Ignicoccus hospitalis from a reconstruction at ∼4-Å resolution. We can now show that the archaeal flagellar filament contains a β-sandwich, previously seen in the FlaF protein that forms the anchor for the archaeal flagellar filament. In contrast to the bacterial flagellar filament, where the outer globular domains make no contact with each other and are not necessary for either assembly or motility, the archaeal flagellin outer domains make extensive contacts with each other that largely determine the interesting mechanical properties of these filaments, allowing these filaments to flex.
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Sheppard, Devon, Jamie-Lee Berry, Rémi Denise, Eduardo P. C. Rocha, Steve Matthews und Vladimir Pelicic. „The major subunit of widespread competence pili exhibits a novel and conserved type IV pilin fold“. Journal of Biological Chemistry 295, Nr. 19 (09.04.2020): 6594–604. http://dx.doi.org/10.1074/jbc.ra120.013316.
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Type IV filaments (T4F), which are helical assemblies of type IV pilins, constitute a superfamily of filamentous nanomachines virtually ubiquitous in prokaryotes that mediate a wide variety of functions. The competence (Com) pilus is a widespread T4F, mediating DNA uptake (the first step in natural transformation) in bacteria with one membrane (monoderms), an important mechanism of horizontal gene transfer. Here, we report the results of genomic, phylogenetic, and structural analyses of ComGC, the major pilin subunit of Com pili. By performing a global comparative analysis, we show that Com pili genes are virtually ubiquitous in Bacilli, a major monoderm class of Firmicutes. This also revealed that ComGC displays extensive sequence conservation, defining a monophyletic group among type IV pilins. We further report ComGC solution structures from two naturally competent human pathogens, Streptococcus sanguinis (ComGCSS) and Streptococcus pneumoniae (ComGCSP), revealing that this pilin displays extensive structural conservation. Strikingly, ComGCSS and ComGCSP exhibit a novel type IV pilin fold that is purely helical. Results from homology modeling analyses suggest that the unusual structure of ComGC is compatible with helical filament assembly. Because ComGC displays such a widespread distribution, these results have implications for hundreds of monoderm species.
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Daehnel, Katrin, Robin Harris, Lucinda Maddera und Philip Silverman. „Fluorescence assays for F-pili and their application“. Microbiology 151, Nr. 11 (01.11.2005): 3541–48. http://dx.doi.org/10.1099/mic.0.28159-0.
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Conjugative pili are extracellular filaments elaborated by Gram-negative bacteria expressing certain type IV secretion systems. They are required at the earliest stages of conjugal DNA transfer to establish specific and secure cell–cell contacts. Conjugative pili also serve as adsorption organelles for both RNA and DNA bacteriophages. Beyond these facts, the structure, formation and function of these filaments are poorly understood. This paper describes a rapid, quantitative assay for F-pili encoded by the F plasmid type IV secretion system. The assay is based on the specific lateral adsorption of icosahedral RNA bacteriophage R17 by F-pili. Bacteriophage particles conjugated with a fluorescent dye, Alexa 488, and bound to F-pili defined filaments visible by immunofluorescence microscopy. F-pili attached to F+ cells and free F-pili were both visible by this method. For quantification, cell-bound bacteriophage were separated from free bacteriophage particles by sedimentation and released by suspending cell pellets in 0·1 % SDS. Fluorescence in cell-free supernatant fractions was measured by fluorometry. The authors present a characterization of this assay and its application to F-pilus formation by cells carrying mutations in the gene for the F-pilus subunit F-pilin. Each mutation introduced a cysteine, which F-pilin normally lacks, at a different position in its primary structure. Cysteine residues in the N-terminal domain I abolished filament formation as measured by fluorescent R17 binding. This was confirmed by measurements of DNA donor activity and filamentous DNA bacteriophage infection. With one exception (G53C), cysteines elsewhere in the F-pilin primary structure did not abolish filament formation, although some mutations differentially affected F-pilus functions.
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Ching, G. Y., und R. K. Liem. „Analysis of the roles of the head domains of type IV rat neuronal intermediate filament proteins in filament assembly using domain-swapped chimeric proteins“. Journal of Cell Science 112, Nr. 13 (01.07.1999): 2233–40. http://dx.doi.org/10.1242/jcs.112.13.2233.
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Type IV neuronal intermediate filament proteins consist of alpha-internexin, which can self-assemble into filaments and the neurofilament triplet proteins, which are obligate heteropolymers, at least in rodents. These IF proteins therefore provide good systems for elucidating the mechanism of intermediate filament assembly. To analyze the roles of the head domains of these proteins in contributing to their differential assembly properties, we generated chimeric proteins by swapping the head domains between rat alpha-internexin and either rat NF-L or NF-M and examined their assembly properties in transfected cells that lack their own cytoplasmic intermediate filament network. Lalphaalpha and Malphaalpha, the chimeric proteins generated by replacing the head domain of alpha-internexin with those of NF-L and NF-M, respectively, were unable to self-assemble into filaments. In contrast, alphaLL, a chimeric NF-L protein generated by replacing the head domain of NF-L with that of alpha-internexin, was able to self-assemble into filaments, whereas MLL, a chimeric NF-L protein containing the NF-M head domain, was unable to do so. These results demonstrate that the alpha-internexin head domain is essential for alpha-internexin's ability to self-assemble. While coassembly of Lalphaalpha with NF-M and coassembly of Malphaalpha with NF-L resulted in formation of filaments, coassembly of Lalphaalpha with NF-L and coassembly of Malphaalpha with NF-M yielded punctate patterns. These coassembly results show that heteropolymeric filament formation requires that one partner has the NF-L head domain and the other partner has the NF-M head domain. Thus, the head domains of rat NF-L and NF-M play important roles in determining the obligate heteropolymeric nature of filament formation. The data obtained from these self-assembly and coassembly studies provide some new insights into the mechanism of intermediate filament assembly.
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Lamparter, Tilman, Jennifer Babian, Katrin Fröhlich, Marion Mielke, Nora Weber, Nadja Wunsch, Finn Zais et al. „The involvement of type IV pili and the phytochrome CphA in gliding motility, lateral motility and photophobotaxis of the cyanobacterium Phormidium lacuna“. PLOS ONE 17, Nr. 1 (27.01.2022): e0249509. http://dx.doi.org/10.1371/journal.pone.0249509.
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Phormidium lacuna is a naturally competent, filamentous cyanobacterium that belongs to the order Oscillatoriales. The filaments are motile on agar and other surfaces and display rapid lateral movements in liquid culture. Furthermore, they exhibit a photophobotactic response, a phototactic response towards light that is projected vertically onto the area covered by the culture. However, the molecular mechanisms underlying these phenomena are unclear. We performed the first molecular studies on the motility of an Oscillatoriales member. We generated mutants in which a kanamycin resistance cassette (KanR) was integrated in the phytochrome gene cphA and in various genes of the type IV pilin apparatus. pilM, pilN, pilQ and pilT mutants were defective in gliding motility, lateral movements and photophobotaxis, indicating that type IV pili are involved in all three kinds of motility. pilB mutants were only partially blocked in terms of their responses. pilB is the proposed ATPase for expelling of the filament in type IV pili. The genome reveals proteins sharing weak pilB homology in the ATPase region, these might explain the incomplete phenotype. The cphA mutant revealed a significantly reduced photophobotactic response towards red light. Therefore, our results imply that CphA acts as one of several photophobotaxis photoreceptors or that it could modulate the photophobotaxis response.
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Steinert, Peter M., Lyuben N. Marekov und David A. D. Parry. „Molecular Parameters of Type IV α-Internexin and Type IV-Type III α-Internexin-Vimentin Copolymer Intermediate Filaments“. Journal of Biological Chemistry 274, Nr. 3 (15.01.1999): 1657–66. http://dx.doi.org/10.1074/jbc.274.3.1657.
Der volle Inhalt der QuelleDissertationen zum Thema "Type IV filaments"
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Jacobsen, Theis. „Structure and assembly of bacterial type IV filaments unravelled by an integrative approach“. Electronic Thesis or Diss., Sorbonne université, 2022. http://www.theses.fr/2022SORUS146.
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La superfamille des filaments de type IV (TFF) est un groupe de machines moléculaires localisées dans la membrane des bactéries et des archées. Ces machines associent des polymères protéiques de manière non-covalente appelés des pili, qui s’étendent depuis la cellule pour réaliser plusieurs fonctions qui ont évolué spécifiquement pour s’adapter à des organismes hôtes différents. La superfamille TFF comprend le système de sécrétion de type II (T2SS) et les pili de type IVa (T4aP). Le T2SS induit la sécrétion de substrats chez les bactéries Gram-négatives. Ces substrats sont en général des enzymes qui dégradent les complexes carbohydrates, le peptidoglycane, les lipides, ce qui à terme entraîne la libération de nutriments. Les T4aP sont de longues fibres flexibles qui sont ancrées dans la membrane et permettent de nombreuses fonctions. Le mécanisme par lequel le T2SS et les T4aP remplissent ces différentes fonctions n’est toujours pas entièrement compris. Pour comprendre le mécanisme de sécrétion du T2SS, nous avons étudié par RMN la structure de la pseudopiline OutG, le composant majeur du pseudopilus chez Dickeya dadantii. Dans une seconde partie, nous avions pour objectif d’aborder la structure et l’assemblage des pilines mineures, des protéines qui composent le T4P d’Escherichia coli entérohémorragique. Nous avons optimisé la surexpression, la purification et le marquage de les pilines mineures pour leur étude par RMN. De plus, la modélisation des pilines et le cross-linking ont été réalisés sur les échantillons de pseudopili T4P et T2SS purifiés en tant que méthodologie pour déterminer la structure et les interactions des pilines et pseudopilines au sein du pilus natifThe type IV filament (TFF) superfamily is a group of molecular machineries located in the membrane of bacteria and archaea. These machineries assemble non-covalent protein polymers called pili extending away from the cell to perform multiple functions which have evolved specifically to adapt to different host organisms. The TFF superfamily includes the type II secretion system (T2SS) and the type IVa pili (T4aP). The T2SS promotes the secretion of substrates in Gram-negative bacteria. These substrates are in general enzymes degrading complex carbohydrates, peptidoglycan, and lipids, resulting in the release of nutrients. The T4aP are long flexible fibres anchored in the membrane and enable various functions such as twitching motility, DNA uptake and biofilm formation. The mechanism by which the T2SS and T4aP pilus fulfil their different functions is still not completely understood. To understand the mechanism of secretion by T2SS, we studied the structure of the pseudopilin OutG, the major component of the pseudopilus in Dickeya dadantii by Nuclear Magnetic Resonance (NMR). In a second part, we aimed to address the structure and the assembly of minor pilins, protein components of Enterohemorrhagic Escherichia coli T4aP. We optimised the overexpression, purification and labelling of the minor pilins for their structural study by NMR. Furthermore, molecular modelling of the minor pilins and crosslinking mass spectrometry were performed on whole T4aP and T2SS pseudopili purified samples as a methodology to determine the structure and the interactions of pilins and pseudopilins within the native pilus
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Leh, David. „Optimisation du dimensionnement d'un réservoir composite type IV pour stockage très haute pression d'hydrogène“. Phd thesis, Université de Grenoble, 2013. http://tel.archives-ouvertes.fr/tel-00942731.
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Ce travail a pour but de proposer une nouvelle approche du dimensionnement optimisé des réservoirs de stockage d'hydrogène de type IV visant à mieux répondre aux enjeux industriels. Les objectifs scientifiques et techniques consistent à disposer de modèles qualifiés pour la simulation du comportement de ces réservoirs, associés à des méthodologies de dimensionnement et d'optimisation fiables. La démarche s'appuie sur trois axes principaux :- proposer une démarche de conception prédictive en intégrant (i) un premier aspect lié à la ruine de la structure qui est la conséquence de mécanismes complexes et multiples d'endommagement s'initiant, s'accumulant et se développant dans un milieu anisotrope et (ii) des modèles de simulation de la structuration composite spécifique au procédé d'enroulement filamentaire, technologie employée largement dans la fabrication des réservoirs de stockage à haute pression. Leurs implémentations constituent une première avancée face à l'existant ;- choisir et évaluer les paramètres structuraux par une démarche d'optimisation où nous sommes amenés à utiliser (i) des méthodes de métamodélisation permettant de répondre aux contraintes de coûts, (ii) des méthodes spécifiques de tri et (iii) des méthodes à spectres larges qui recherchent des solutions sur une large population telles que des méthodes génétiques ;- qualifier la démarche dans sa globalité par une comparaison entre calculs et essais. Ainsi, la finalité de ce travail est de développer et valider des modèles et méthodes pour permettre de mieux concevoir, tester et fabriquer à moindre coût un réservoir avec une structure calculée optimisée.
Buchteile zum Thema "Type IV filaments"
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Weber, Klaus. „Evolutionary aspects of IF proteins“. In Guidebook to the Cytoskeletal and Motor Proteins, 291–95. Oxford University PressOxford, 1999. http://dx.doi.org/10.1093/oso/9780198599579.003.0094.
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Abstract Extensive work on cytoplasmic intermediate filament (IF) proteins of vertebrates has established a very complex multigene family with close to 50 members in a mammal. These proteins include IF types I to IV as well as additional species. Variability arises primarily from the N- and C-terminal head and tail domains but also from the central α-helical rod domain, which, except for its ends, preserves sequence principles rather than actual sequences. In spite of such differences the helix 1B subdomain keeps a constant length in all cytoplasmic IF proteins so far characterized from vertebrates. In contrast, in nuclear lamins, a special subtype (type V) of IF proteins, from both vertebrates and invertebrates, the length of helix 1B is increased by six heptads or 42 residues. Recent evidence shows that the long helix 1B form is also characteristic of the cytoplasmic IF proteins of various protostomic animals.H Thus in metazoan evolution the long helix 1B form was changed by a deletion to the short helix 1B version prior to the diversification into types I to IV. Current results suggest that this deletion occurred either with the deuterostomic branch or prior to the chordates. Sequence comparisons further suggest that cytoplasmic IF proteins arose possibly from a mutated lamin early in evolution.
Konferenzberichte zum Thema "Type IV filaments"
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Farhood, Naseer H., Saravanan Karuppanan, H. H. Ya und Mohamad Ariff Baharom. „Burst pressure investigation of filament wound type IV composite pressure vessel“. In ADVANCED MATERIALS FOR SUSTAINABILITY AND GROWTH: Proceedings of the 3rd Advanced Materials Conference 2016 (3rd AMC 2016). Author(s), 2017. http://dx.doi.org/10.1063/1.5010482.
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BIANCHI, I. „Process and structural simulation for the development of a pressure vessel through filament winding technology“. In Material Forming. Materials Research Forum LLC, 2023. http://dx.doi.org/10.21741/9781644902479-38.
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Abstract. Recently, Università Politecnica delle Marche and COMEC Innovative srl are involved in the research project “Smart Tow Winding” funded by MIUR (Ministry of Education, University and Research), concerning the development of an innovative process for the realization of a pressure vessel through Filament Winding (FW) technology. In this context, a design procedure for type IV composite pressure vessel is proposed. To design the component, the dedicated simulation software CADWIND was used to virtually generate the pressure vessel through the definition of the desired geometry, the type of prepreg, the number of layers and the bandwidth. The generated file was imported in the FEM simulation software Siemens NX with the aim of evaluating the structural resistance under an internal pressure of 70 MPa. Different external configurations of mandrels and stratification were tested in order to optimize the geometry of the vessel and the resistance to weight ratio. A high performance and low weight vessel configuration was finally identified.
Berichte der Organisationen zum Thema "Type IV filaments"
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Elbaum, Michael, und Peter J. Christie. Type IV Secretion System of Agrobacterium tumefaciens: Components and Structures. United States Department of Agriculture, März 2013. http://dx.doi.org/10.32747/2013.7699848.bard.
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Objectives: The overall goal of the project was to build an ultrastructural model of the Agrobacterium tumefaciens type IV secretion system (T4SS) based on electron microscopy, genetics, and immunolocalization of its components. There were four original aims: Aim 1: Define the contributions of contact-dependent and -independent plant signals to formation of novel morphological changes at the A. tumefaciens polar membrane. Aim 2: Genetic basis for morphological changes at the A. tumefaciens polar membrane. Aim 3: Immuno-localization of VirB proteins Aim 4: Structural definition of the substrate translocation route. There were no major revisions to the aims, and the work focused on the above questions. Background: Agrobacterium presents a unique example of inter-kingdom gene transfer. The process involves cell to cell transfer of both protein and DNA substrates via a contact-dependent mechanism akin to bacterial conjugation. Transfer is mediated by a T4SS. Intensive study of the Agrobacterium T4SS has made it an archetypal model for the genetics and biochemistry. The channel is assembled from eleven protein components encoded on the B operon in the virulence region of the tumor-inducing plasmid, plus an additional coupling protein, VirD4. During the course of our project two structural studies were published presenting X-ray crystallography and three-dimensional reconstruction from electron microscopy of a core complex of the channel assembled in vitro from homologous proteins of E. coli, representing VirB7, VirB9, and VirB10. Another study was published claiming that the secretion channels in Agrobacterium appear on helical arrays around the membrane perimeter and along the entire length of the bacterium. Helical arrangements in bacterial membranes have since fallen from favor however, and that finding was partially retracted in a second publication. Overall, the localization of the T4SS within the bacterial membranes remains enigmatic in the literature, and we believe that our results from this project make a significant advance. Summary of achievements : We found that polar inflations and other membrane disturbances relate to the activation conditions rather than to virulence protein expression. Activation requires low pH and nutrient-poor medium. These stress conditions are also reflected in DNA condensation to varying degrees. Nonetheless, they must be considered in modeling the T4SS as they represent the relevant conditions for its expression and activity. We identified the T4SS core component VirB7 at native expression levels using state of the art super-resolution light microscopy. This marker of the secretion system was found almost exclusively at the cell poles, and typically one pole. Immuno-electron microscopy identified the protein at the inner membrane, rather than at bridges across the inner and outer membranes. This suggests a rare or transient assembly of the secretion-competent channel, or alternatively a two-step secretion involving an intermediate step in the periplasmic space. We followed the expression of the major secreted effector, VirE2. This is a single-stranded DNA binding protein that forms a capsid around the transferred oligonucleotide, adapting the bacterial conjugation to the eukaryotic host. We found that over-expressed VirE2 forms filamentous complexes in the bacterial cytoplasm that could be observed both by conventional fluorescence microscopy and by correlative electron cryo-tomography. Using a non-retentive mutant we observed secretion of VirE2 from bacterial poles. We labeled the secreted substrates in vivo in order detect their secretion and appearance in the plant cells. However the low transfer efficiency and significant background signal have so far hampered this approach.