TY - JOUR A1 - Sperber, Hannah Sabeth A1 - Welke, Robert-William A1 - Petazzi, Roberto Arturo A1 - Bergmann, Ronny A1 - Schade, Matthias A1 - Shai, Yechiel A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Self-association and subcellular localization of Puumala hantavirus envelope proteins JF - Scientific reports N2 - Hantavirus assembly and budding are governed by the surface glycoproteins Gn and Gc. In this study, we investigated the glycoproteins of Puumala, the most abundant Hantavirus species in Europe, using fluorescently labeled wild-type constructs and cytoplasmic tail (CT) mutants. We analyzed their intracellular distribution, co-localization and oligomerization, applying comprehensive live, single-cell fluorescence techniques, including confocal microscopy, imaging flow cytometry, anisotropy imaging and Number&Brightness analysis. We demonstrate that Gc is significantly enriched in the Golgi apparatus in absence of other viral components, while Gn is mainly restricted to the endoplasmic reticulum (ER). Importantly, upon co-expression both glycoproteins were found in the Golgi apparatus. Furthermore, we show that an intact CT of Gc is necessary for efficient Golgi localization, while the CT of Gn influences protein stability. Finally, we found that Gn assembles into higher-order homo-oligomers, mainly dimers and tetramers, in the ER while Gc was present as mixture of monomers and dimers within the Golgi apparatus. Our findings suggest that PUUV Gc is the driving factor of the targeting of Gc and Gn to the Golgi region, while Gn possesses a significantly stronger self-association potential. Y1 - 2019 U6 - https://doi.org/10.1038/s41598-018-36879-y SN - 2045-2322 VL - 9 PB - Nature Publ. Group CY - London ER - TY - CHAP A1 - Haralampiev, Ivan A1 - Mertens, Monique A1 - Schwarzer, Roland A1 - Herrmann, Andreas A1 - Volkmer, Rudolf A1 - Wessig, Pablo A1 - Müller, Peter T1 - A palmitic acid functionalized with a maleimide group is used to recruit SH-containing peptides to lipid and biological membranes T2 - The FEBS journal Y1 - 2015 SN - 1742-464X SN - 1742-4658 VL - 282 SP - 204 EP - 204 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Wawrzinek, Robert A1 - Wessig, Pablo A1 - Möllnitz, Kristian A1 - Nikolaus, Joerg A1 - Schwarzer, Roland A1 - Müller, Peter A1 - Herrmann, Andreas T1 - DBD dyes as fluorescent probes for sensing lipophilic environments JF - Bioorganic & medicinal chemistry letters : a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry and related disciplines N2 - Small fluorescent organic molecules based on [1,3]dioxolo[4,5-f][1,3]benzodioxole (DBD) could be used as probes for lipophillic microenvironments in aqueous solutions by indicating the critical micelles concentration of detergents and staining cell organelles. Their fluorescence lifetime decreases drastically by the amount of water in their direct environment. Therefore they are potential probes for fluorescence lifetime imaging microscopy (FLIM). KW - Fluorescence lifetime probes KW - FLIM KW - Cell staining KW - Lysotrackers KW - Detergents Y1 - 2012 U6 - https://doi.org/10.1016/j.bmcl.2012.07.056 SN - 0960-894X VL - 22 IS - 17 SP - 5367 EP - 5371 PB - Elsevier CY - Oxford ER - TY - GEN A1 - Sperber, Hannah Sabeth A1 - Welke, Robert-William A1 - Petazzi, Roberto Arturo A1 - Bergmann, Ronny A1 - Schade, Matthias A1 - Shai, Yechiel A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Self-association and subcellular localization of Puumala hantavirus envelope proteins T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Hantavirus assembly and budding are governed by the surface glycoproteins Gn and Gc. In this study, we investigated the glycoproteins of Puumala, the most abundant Hantavirus species in Europe, using fluorescently labeled wild-type constructs and cytoplasmic tail (CT) mutants. We analyzed their intracellular distribution, co-localization and oligomerization, applying comprehensive live, single-cell fluorescence techniques, including confocal microscopy, imaging flow cytometry, anisotropy imaging and Number&Brightness analysis. We demonstrate that Gc is significantly enriched in the Golgi apparatus in absence of other viral components, while Gn is mainly restricted to the endoplasmic reticulum (ER). Importantly, upon co-expression both glycoproteins were found in the Golgi apparatus. Furthermore, we show that an intact CT of Gc is necessary for efficient Golgi localization, while the CT of Gn influences protein stability. Finally, we found that Gn assembles into higher-order homo-oligomers, mainly dimers and tetramers, in the ER while Gc was present as mixture of monomers and dimers within the Golgi apparatus. Our findings suggest that PUUV Gc is the driving factor of the targeting of Gc and Gn to the Golgi region, while Gn possesses a significantly stronger self-association potential. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 648 KW - Sin-Nombre-Virus KW - nucleocapsid protein KW - cytoplasmic tails KW - electron cryotomography KW - autophagic clearance KW - glycoprotein KW - Gn KW - G1 KW - brightness KW - fever Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-425040 SN - 1866-8372 IS - 648 ER - TY - JOUR A1 - Haralampiev, Ivan A1 - Mertens, Monique A1 - Schwarzer, Roland A1 - Herrmann, Andreas A1 - Volkmer, Rudolf A1 - Wessig, Pablo A1 - Mueller, Peter T1 - Recruitment of SH-Containing peptides to lipid and biological membranes through the use of a palmitic acid functionalized with a Maleimide Group JF - Angewandte Chemie : a journal of the Gesellschaft Deutscher Chemiker ; International edition N2 - This study presents a novel and easily applicable approach to recruit sulfhydryl-containing biomolecules to membranes by using a palmitic acid which is functionalized with a maleimide group. Notably, this strategy can also be employed with preformed (biological) membranes. The applicability of the assay is demonstrated by characterizing the binding of a Rhodamine-labeled peptide to lipid and cellular membranes using methods of fluorescence spectroscopy, lifetime measurement, and microscopy. Our approach offers new possibilities for preparing biologically active liposomes and manipulating living cells. KW - liposomes KW - maleimide KW - membranes KW - palmitic acid KW - palmitoylation KW - peptides Y1 - 2015 U6 - https://doi.org/10.1002/anie.201408089 SN - 1433-7851 SN - 1521-3773 VL - 54 IS - 1 SP - 323 EP - 326 PB - Wiley-VCH CY - Weinheim ER - TY - BOOK A1 - Schwarzer, Ingo A1 - Weiß-Saoumi, Said A1 - Kittel, Roland A1 - Friedrich, Tobias A1 - Kaynak, Koraltan A1 - Durak, Cemil A1 - Isbarn, Andreas A1 - Diestel, Jörg A1 - Knittel, Jens A1 - Franz, Marquart A1 - Morra, Carlos A1 - Stahnke, Susanne A1 - Braband, Jens A1 - Dittmann, Johannes A1 - Griebel, Stephan A1 - Krampf, Andreas A1 - Link, Martin A1 - Müller, Matthias A1 - Radestock, Jens A1 - Strub, Leo A1 - Bleeke, Kai A1 - Jehl, Leander A1 - Kapitza, Rüdiger A1 - Messadi, Ines A1 - Schmidt, Stefan A1 - Schwarz-Rüsch, Signe A1 - Pirl, Lukas A1 - Schmid, Robert A1 - Friedenberger, Dirk A1 - Beilharz, Jossekin Jakob A1 - Boockmeyer, Arne A1 - Polze, Andreas A1 - Röhrig, Ralf A1 - Schäbe, Hendrik A1 - Thiermann, Ricky T1 - RailChain BT - Abschlussbericht N2 - The RailChain project designed, implemented, and experimentally evaluated a juridical recorder that is based on a distributed consensus protocol. That juridical blockchain recorder has been realized as distributed ledger on board the advanced TrainLab (ICE-TD 605 017) of Deutsche Bahn. For the project, a consortium consisting of DB Systel, Siemens, Siemens Mobility, the Hasso Plattner Institute for Digital Engineering, Technische Universität Braunschweig, TÜV Rheinland InterTraffic, and Spherity has been formed. These partners not only concentrated competencies in railway operation, computer science, regulation, and approval, but also combined experiences from industry, research from academia, and enthusiasm from startups. Distributed ledger technologies (DLTs) define distributed databases and express a digital protocol for transactions between business partners without the need for a trusted intermediary. The implementation of a blockchain with real-time requirements for the local network of a railway system (e.g., interlocking or train) allows to log data in the distributed system verifiably in real-time. For this, railway-specific assumptions can be leveraged to make modifications to standard blockchains protocols. EULYNX and OCORA (Open CCS On-board Reference Architecture) are parts of a future European reference architecture for control command and signalling (CCS, Reference CCS Architecture – RCA). Both architectural concepts outline heterogeneous IT systems with components from multiple manufacturers. Such systems introduce novel challenges for the approved and safety-relevant CCS of railways which were considered neither for road-side nor for on-board systems so far. Logging implementations, such as the common juridical recorder on vehicles, can no longer be realized as a central component of a single manufacturer. All centralized approaches are in question. The research project RailChain is funded by the mFUND program and gives practical evidence that distributed consensus protocols are a proper means to immutably (for legal purposes) store state information of many system components from multiple manufacturers. The results of RailChain have been published, prototypically implemented, and experimentally evaluated in large-scale field tests on the advanced TrainLab. At the same time, the project showed how RailChain can be integrated into the road-side and on-board architecture given by OCORA and EULYNX. Logged data can now be analysed sooner and also their trustworthiness is being increased. This enables, e.g., auditable predictive maintenance, because it is ensured that data is authentic and unmodified at any point in time. N2 - Das Projekt RailChain hat einen verteilten Juridical Recorder entworfen, implementiert und experimentell evaluiert, der auf einem echtzeitfähigen verteilten Konsensprotokoll basiert. Dieser Juridical Blockchain Recorder wurde als distributed ledger an Bord des advanced TrainLabs der Deutschen Bahn (ICE-TD 605 017) umgesetzt. Für das Projekt hat sich ein Konsortium aus DB Systel, Siemens, Siemens Mobility, dem Hasso-Plattner-Institut für Digital Engineering, der Technischen Universität Braunschweig, sowie TÜV Rheinland InterTraffic und Spherity formiert und dabei Kompetenzen aus den Bereichen Bahnbetrieb, Informatik und Zulassungswesen gebündelt. Die Partner kombinieren Erfahrungen aus der Industrie und die akademische Forschung mit der Aufbruchstimmung aus dem Start-Up-Umfeld. Distributed-Ledger-Technologien (DLTs) definieren verteilte Datenbanken und stellen ein digitales Protokoll für Transaktionen zwischen Geschäftspartnern dar, ohne dass ein Mittelsmann beteiligt sein müsste. Die Implementierung einer Blockchain mit Echtzeitanforderungen für das lokale Netzwerk einer Eisenbahnanlage (z. B. Stellwerk oder Zug) erlaubt es, die im verteilten System entstehenden Daten nachweislich in Echtzeit zu protokollieren. Dabei können eisenbahnspezifische Randbedingungen ausgenutzt werden, um Standard-Blockchain-Protokolle anzupassen. EULYNX und OCORA (Open CCS On-board Reference Architecture) sind Bestandteile einer zukünftigen europäischen Referenzarchitektur für das Leit- und Sicherungssystem (Reference CCS Architecture – RCA, Control Command and Signalling – CCS). Beide Architekturkonzepte skizzieren herstellerübergreifende, komponentenbasierende heterogene IT-Systeme. Solche Systeme bergen neue Herausforderungen, die bislang im Kontext der zugelassenen, sicherheitsrelevanten Leit- und Sicherungstechnik der Bahn weder strecken- noch fahrzeugseitig adressiert werden mussten. Logbuch-Implementierungen, wie der gängige Juridical Recorder auf Fahrzeugen, können nun nicht mehr als zentrale Systemkomponente eines einzelnen Herstellers umgesetzt werden. Alle zentralisierten Lösungsansätze sind in Frage gestellt. Das mFUND-geförderte Forschungsprojekt erbringt den praktischen Nachweis, dass Zustandsinformationen über eine Vielzahl von Systemkomponenten herstellerübergreifend und gerichtsfest mittels verteilten Konsensprotokollen gespeichert werden können. Ergebnisse von RailChain wurden publiziert, prototypisch implementiert und in großen Feldtests auf dem advanced TrainLab experimentell evaluiert. Gleichzeitig wurde aufgezeigt, wie sich RailChain in den mit OCORA und EULYNX vorgegebenen fahrzeug- und streckenseitigen Architekturentwurf integrieren lässt. Daten können dadurch zeitnaher ausgewertet werden und gleichzeitig wird ihre Vertrauenswürdigkeit erhöht. Dies ermöglicht u. a. nachvollziehbare zustandsorientierte Wartung, denn es kann jederzeit sichergestellt werden, dass die Daten authentisch sind und auch nicht verändert wurden. T3 - Technische Berichte des Hasso-Plattner-Instituts für Digital Engineering an der Universität Potsdam - 152 KW - Distributed-Ledger-Technologie (DLT) KW - juridical recording KW - Konsensprotokolle KW - consensus protocols KW - Digitalisierung KW - digitalization KW - Bahnwesen KW - railways KW - Blockchain KW - asset management KW - selbstbestimmte Identitäten KW - self-sovereign identity KW - dezentrale Identitäten KW - decentral identities KW - überprüfbare Nachweise KW - verifiable credentials KW - Echtzeit KW - real-time KW - Standardisierung KW - standardization KW - Verlässlichkeit KW - dependability KW - Fehlertoleranz KW - fault tolerance Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-577409 SN - 978-3-86956-550-7 SN - 1613-5652 SN - 2191-1665 IS - 152 PB - Universitätsverlag Potsdam CY - Potsdam ER - TY - JOUR A1 - Welke, Robert-William A1 - Sperber, Hannah Sabeth A1 - Bergmann, Ronny A1 - Koikkarah, Amit A1 - Menke, Laura A1 - Sieben, Christian A1 - Krüger, Detlev H. A1 - Chiantia, Salvatore A1 - Herrmann, Andreas A1 - Schwarzer, Roland T1 - Characterization of hantavirus N protein intracellular dynamics and localization JF - Viruses N2 - Hantaviruses are enveloped viruses that possess a tri-segmented, negative-sense RNA genome. The viral S-segment encodes the multifunctional nucleocapsid protein (N), which is involved in genome packaging, intracellular protein transport, immunoregulation, and several other crucial processes during hantavirus infection. In this study, we generated fluorescently tagged N protein constructs derived from Puumalavirus (PUUV), the dominant hantavirus species in Central, Northern, and Eastern Europe. We comprehensively characterized this protein in the rodent cell line CHO-K1, monitoring the dynamics of N protein complex formation and investigating co-localization with host proteins as well as the viral glycoproteins Gc and Gn. We observed formation of large, fibrillar PUUV N protein aggregates, rapidly coalescing from early punctate and spike-like assemblies. Moreover, we found significant spatial correlation of N with vimentin, actin, and P-bodies but not with microtubules. N constructs also co-localized with Gn and Gc albeit not as strongly as the glycoproteins associated with each other. Finally, we assessed oligomerization of N constructs, observing efficient and concentration-dependent multimerization, with complexes comprising more than 10 individual proteins. KW - hantavirus KW - N protein KW - oligomerization KW - actin KW - P-bodies KW - vimentin KW - Number and Brightness KW - Puumalavirus KW - macromolecular assemblies Y1 - 2022 U6 - https://doi.org/10.3390/v14030457 SN - 1999-4915 VL - 14 IS - 3 PB - MDPI CY - Basel ER -