The IM30 (inner membrane-associated protein of 30 kDa), also known as the Vipp1 (vesicle-inducing protein in plastids 1), has a crucial role in thylakoid membrane biogenesis and maintenance. Recent ...results suggest that the protein binds peripherally to membranes containing negatively charged lipids. However, although IM30 monomers interact and assemble into large oligomeric ring complexes with different numbers of monomers, it is still an open question whether ring formation is crucial for membrane interaction. Here we show that binding of IM30 rings to negatively charged phosphatidylglycerol membrane surfaces results in a higher ordered membrane state, both in the head group and in the inner core region of the lipid bilayer. Furthermore, by using gold nanorods covered with phosphatidylglycerol layers and single particle spectroscopy, we show that not only IM30 rings but also lower oligomeric IM30 structures interact with membranes, although with higher affinity. Thus, ring formation is not crucial for, and even counteracts, membrane interaction of IM30.
IM30/Vipp1 proteins are crucial for thylakoid membrane biogenesis in chloroplasts and cyanobacteria. A characteristic C-terminal extension distinguishes these proteins from the homologous bacterial ...PspA proteins, and this extension has been discussed to be key for the IM30/Vipp1 activity. Here we report that the extension of the Synechocystis IM30 protein is indispensable, and argue that both, the N-terminal PspA-domain as well as the C-terminal extension are needed in order for the IM30 protein to conduct its in vivo function. In vitro, we show that the PspA-domain of IM30 is vital for stability/folding and oligomer formation of IM30 as well as for IM30-triggered membrane fusion. In contrast, the IM30 C-terminal domain is involved in and necessary to stabilize defined contacts to negatively charged membrane surfaces, and to modulate the IM30-induced membrane fusion activity. Although the two IM30 protein domains have distinct functional roles, only together they enable IM30 to work properly.
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•The IM30 C-terminus does not affect the structure and stability of IM30.•Membrane binding of IM30 involves the PspA- as well as the C-terminal domain.•The IM30 C-terminus interacts with negatively charged membrane surfaces.•Charged residues inside the IM30 C-terminus influence stability of membrane binding.•Membrane binding of the IM30 C-terminus correlates with the proteins fusogenic activity.
Cryo-EM is a powerful tool for understanding macromolecular structures, yet current methods for structure reconstruction are slow and computationally demanding. To accelerate research on pose ...estimation, we present CESPED, a data set specifically designed for supervised pose estimation in cryo-EM. Alongside CESPED, we provide a package to simplify cryo-EM data handling and model evaluation. We evaluate the performance of a baseline model, Image2Sphere, on CESPED, which shows promising results but also highlights the need for further improvements. Additionally, we illustrate the potential of deep learning-based pose estimators to generalize across different samples, suggesting a promising path toward more efficient processing strategies. Published by the American Physical Society 2024
The mechanisms that lead to the tegumentation of herpesviral particles are only poorly defined. The phosphoprotein 65 (pp65) is the most abundant constituent of the virion tegument of human ...cytomegalovirus (HCMV). It is, however, nonessential for virion formation. This seeming discrepancy has not met with a satisfactory explanation regarding the role of pp65 in HCMV particle morphogenesis. Here, we addressed the question of how the overall tegument composition of the HCMV virion depended on pp65 and how the lack of pp65 influenced the packaging of particular tegument proteins. To investigate this, we analyzed the proteomes of pp65-positive (pp65pos) and pp65-negative (pp65neg) virions by label-free quantitative mass spectrometry and determined the relative abundances of tegument proteins. Surprisingly, only pUL35 was elevated in pp65neg virions. As the abundance of pUL35 in the HCMV tegument is low, it is unlikely that it replaced pp65 as a structural component in pp65neg virions. A subset of proteins, including the third most abundant tegument protein, pUL25, as well as pUL43, pUL45, and pUL71, were reduced in pp65neg or pp65low virions, indicating that the packaging of these proteins was related to pp65. The levels of tegument components, like pp28 and the capsid-associated tegument proteins pp150, pUL48, and pUL47, were unaffected by the lack of pp65. Our analyses demonstrate that deletion of pp65 is not compensated for by other viral proteins in the process of virion tegumentation. The results are concordant with a model of pp65 serving as an optional scaffold protein that facilitates protein upload into the outer tegument of HCMV particles.
The assembly of the tegument of herpesviruses is only poorly understood. Particular proteins, like HCMV pp65, are abundant tegument constituents. pp65 is thus considered to play a major role in tegument assembly in the process of virion morphogenesis. We show here that deletion of the pp65 gene leads to reduced packaging of a subset of viral proteins, indicating that pp65 acts as an optional scaffold protein mediating protein upload into the tegument.
Deadbeat-direct torque and flux control is a flux-observer-based high-bandwidth digital closed-loop torque control law that achieves the commanded torque at the end of each switching period. When ...flux estimation is accurate, instantaneous torque is fed back and undesirable pulsating torque is inherently minimized. This study presents real-time flux-observer-based torque ripple estimation of a highly nonlinear synchronous reluctance machine. Saturation and cross saturation are incorporated directly into the control law, and are considered in each switching period. Deadbeat torque response and torque ripple reduction is demonstrated experimentally. Finite element method simulations validate the torque ripple minimization.
Oxygen transport in the hemolymph of many arthropods is mediated by hemocyanins, large copper‐containing proteins that are well‐studied in Chelicerata and Crustacea, but had long been considered ...unnecessary in the subphylum of Myriapoda. Only recently has it become evident that hemocyanins are present in Scutigeromorpha (Chilopoda) and Spirostreptida (Diplopoda). Here we present evidence for a more widespread occurrence of hemocyanin in the myriapods. By means of RT‐PCR, western blotting and database searches, hemocyanins were identified in the symphylans Hanseniella audax and Symphylella vulgaris, the chilopod Scolopendra subspinipes dehaani and the diplopod Polydesmus angustus. No hemocyanins were found in the diplopods Polyxenus lagurus, Cylindroiulus punctatus, Glomeris marginata, Glomeris pustulata and Arthrosphaera brandtii, or the chilopods Lithobius forficatus, Geophilus flavus and Strigamia maritima. This suggests multiple independent losses in myriapod taxa. Two independent hemocyanin subunits were found that were already present in the myriapod stem line. We specifically investigated the structure of the hemocyanin of P. angustus, which consists of three distinct subunits that occur in an approximately equimolar ratio. As deduced by 3D electron microscopy, the quaternary structure is a 3 × 6‐mer that resembles the half structure of the 6 × 6‐mer hemocyanin from Scutigera coleoptrata. It was analyzed more closely by homology modeling of 1 × 6‐mers and their rigid‐body fitting to the electron density map of the 3 × 6‐mer. In addition, we obtained the cDNA sequence of a putative myriapod phenoloxidase. Phenoloxidases are related to the arthropod hemocyanins, but diverged before radiation of the arthropod subphyla.
Hemocyanin transports O2 in many arthropods. We demonstrate a widespread occurrence hemocyanin in centipedes, millipedes and symphylans. Some species lack hemocyanin. Two distinct subunit types evolved early in myriapod evolution. We further show the presence of hemocyanin‐like phenoloxidases in this taxon. Specifically, the structure of the 3 × 6‐mer hemocyanin of Polydesmus angustus was revealed by sequencing, 3D electron microscopy and homology modelling.
Materiale Textkulturen Thomas Meier, Michael R. Ott, Rebecca Sauer / Thomas Meier, Michael R. Ott, Rebecca Sauer
2015, 2015-02-13, 2015-12-18, Letnik:
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Was ist und wie erforscht man die 'Materialität und Präsenz des Geschriebenen in non-typographischen Gesellschaften'? In diesem ersten Band der Reihe Materiale Textkulturen setzt sich der ...Sonderforschungsbereich 933 mit den Konzepten, Materialien und Praktiken auseinander, zu denen seit dem Jahr 2011 an der Universität Heidelberg geforscht wird. In mehr als 50 Beiträgen, die fast 60 Autorinnen und Autoren aus zahlreichen Disziplinen gemeinsam diskutiert und verfasst haben, werden theoretische Grundlagen vorgestellt, texttragende Materialien und Gegenstände präsentiert und die vielfältigen Praktiken analysiert, in die solche texttragenden Artefakte eingebunden sind. Durch eine Vielzahl von Beispielen und Abbildungen aus einem Zeitraum von mehr als 4500 Jahren werden die Analysen vertieft und Analogien und Differenzen deutlich, die nur im disziplinübergreifenden Gespräch sichtbar werden. Der Sammelband richtet sich an alle Forscherinnen und Forscher, die mit texttragenden Artefakten zu tun haben und sich für die Position ihrer Gegenstände innerhalb weitreichender textkultureller Netzwerke interessieren. Nicht zuletzt wendet sich das Buch an eine breitere Öffentlichkeit, der sich der SFB 933 mit diesem Handbuch vorstellt.
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