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Sequence-specific response of collagen-mimetic peptides to osmotic pressure
Full text linkNative collagen molecules usually contract upon dehydration, but the details of their interaction with water are poorly understood. Previous molecular modeling studies indicated a spatially inhomogeneous response, with a combination of local axial expansion and contraction. Such sequence-dependent effects are difficult to study with native collagen. In this article, we use collagen-mimetic peptides (CMPs) to investigate the effect of osmotic pressure on several collagen-mimetic sequences. Synchrotron x-ray diffraction combined with molecular dynamics simulations shows that CMPs pack differently depending on osmotic pressure and exhibit changes in the helical rise per residue of individual molecules. Infrared spectroscopy reveals that osmotic pressure affects the stability of the triple helix through changes in triple helix-stabilizing hydrogen bonds. Surprisingly, CMPs with the canonical collagen sequence glycine–proline–hydroxyproline are found to elongate upon dehydration, while sequence modifications are able to reverse this tendency. This strongly suggests that the overall contraction of native collagen molecules is not programmed into the canonical sequence but is specific to local amino acids that substitute for proline or hydroxyproline along the protein chain. Collagen is an essential protein in mammalian extracellular tissues and a better understanding of its mechanical function is important both from a materials science and from a biomedical viewpoint. Recently, collagen has been shown to contract along the fibre direction when subjected to osmotic stress, a process that could play important roles in strengthening bone and in developing tissue tension during extracellular matrix development. The present work uses collagen-like short peptides to show that the canonical collagen sequence is not responsible for this contraction. The conclusion is that the collagen amino acid sequence must have evolved to include guest sequences within the canonical glycine-proline-hydroxyproline repeat that provide the observed contractility.
Impact statement: Collagen is an essential protein in mammalian extracellular tissues and a better understanding of its mechanical function is important both from a materials science and from a biomedical viewpoint. Recently, collagen has been shown to contract along the fibre direction when subjected to osmotic stress, a process that could play important roles in strengthening bone and in developing tissue tension during extracellular matrix development. The present work uses collagen-like short peptides to show that the canonical collagen sequence is not responsible for this contraction. The conclusion is that the collagen amino acid sequence must have evolved to include guest sequences within the canonical glycine-proline-hydroxyproline that provide the observed contractility
The home bias and the local bias: A survey
Full text linkThe home bias like the disposition effect is a well-researched economic phenomenon in investor behaviour which has been examined in finance journal articles for decades. While there is little doubt about the existence of the bias, its magnitude varies across countries and investor groups. The home bias has to be regarded as a multifactorial phenomenon, a combination of numerous causes which all synergistically contribute. In contrast to other biases the home bias can at least partially be explained by reasons beyond irrational investor behaviour. While institutional restrictions play a minor role, informational asymmetries and superior information of domestic investors are important factors. Thus, the performance of investments may well benefit from a home bias, and the bias then no longer would be a puzzle but rather rational behaviour as a lower diversification level may lead to higher returns. The contemporary understanding of the home bias gains in relevance as the ongoing political debate in Germany has to clarify an institutional framework for long-run retirement savings plans of private households based on equity investments
Characterization of the FK506 binding protein 51-Glucocorticoid Receptor-p23 complex in living cells
Full text linkFKBP51 is a co-chaperone known for its inhibitory role in GR signaling. Besides FKBP51, there are multiple other co-chaperones known to interact with the GR, such as the small protein p23 which ensures the correct folding and activity of the GR. Understanding the complex formation and protein-protein interactions in these complexes are crucial for comprehending the regulation and activity of proteins.
In this work I analyzed the interaction of FKBP51 and p23 as well as the interaction of the GR and p23.
I utilized photoinducible crosslinking which relies on the incorporation of an unnatural amino acid at the surface of a protein. This unnatural amino acid can form a covalent bond with another protein in the vicinity upon UV irradiation. The covalent bond formation can then be analyzed by e.g. Western Blotting and deliver information about interaction sites between two proteins.
I used this method in this work to show that (i) FKBP51 and p23 interact in dependence of Hsp90; (ii) the FKBP51-p23-Hsp90 complex can exist independently of the GR; (iii) the interaction of FKBP51 and p23 can be partially abolished by addition of the GR; (iv) FKBP51, GR and p23 can participate in a Hsp90 associated complex; and (v) FKBP51 enhances the GR p23 interaction while FKBP52 does not.
Taken together, I showed the existence of a FKBP51-GR-p23-complex for the first time in cellulo.
In addition to the data about FKBP51-p23 interactions, I also provided novel insights into the GR p23 interaction: I showed that (i) the GR carries two distinct interaction sites with p23; (ii) each interaction hotspot binds to a different region of the C-terminal domain of p23; and (iii) the C-terminal tail of p23 is necessary to enhance the GR activity, even though it is not as relevant for the interaction of the probed GR residues with p23.
Overall, the data about the FKBP51-p23 and FKBP51-GR-p23 interaction I gathered provide additional information about the structures of the mentioned complexes in cellulo. This will prove useful in the future for further understanding of the GR regulation and ultimately may result in novel ways to manipulate the GR.
In addition to the FKBP51-GR interaction, I also investigated the role of FKBP51 in NF κB signaling. I investigated multiple cell lines, different readouts and stimuli and was not able to determine an effect of FKBP51 degradation or inhibition on the NF κB activity in the chosen context
Impact of S-sulfocysteine on Chinese hamster ovary cells
Full text linkIn the biopharmaceutical industry, Chinese hamster ovary (CHO) cells are the primary workhorse for therapeutic protein production (monoclonal antibodies, fusion proteins) in large-scale processes — constantly undergoing optimization to increase the product yield. In this context, optimized cell culture media and feeds are critical and often rely on cysteine (Cys) analogues to overcome the Cys-related stability and solubility issues at neutral pH. Previously, the Cys analogue, S-sulfocysteine (SSC), was successfully applied in cell culture, implying the SSC uptake, metabolization and biological activity, though the latter have not yet been characterized thoroughly. Furthermore, SSC also demonstrated a concentration-dependent toxicity in cells. Therefore, the present study sought to investigate the SSC metabolism and activity in more detail to provide an enhanced understanding and knowledge surrounding its beneficial, non-toxic and toxic characteristics.
In the first part of the study, the CHO-K1 model clone was investigated in the presence of SSC at a non-toxic concentration to understand its metabolism/activity, and underlying mechanism(s) of action. Therefore, a CHO-K1-based fed-batch process (+ pH control), multi-omics and mechanistic approaches were performed, pointing out the following hallmarks: (1) SSC/glutathione (GSH) mixed disulfide formation, (2) glutaredoxin 1- and glutathione reductase (GR)-conveyed reduction and (3) sulfur species liberation, followed by their diversion towards catabolic pathways. The SSC-derived molecules, GSH, taurine and sulfite, were identified as those that are most likely conferring the favorable activity via their antioxidant and protective roles.
In the second part of the study, the CHO-K1 clone was examined using SSC at toxic concentrations to understand the mechanism(s) of toxicity. The CHO-K1 fed-batch process (± pH control) pointed to the SSC toxicity in the pH- surrounding that correlated with an extracellular pH decrease — likely reflecting an intracellular pH drop. The sulfo-glutathione (GS-SO3) mixed disulfide reduction, particularly the GR-conveyed GSH recycling, was strongly hampered at lower pH levels in vitro. Complementary to these findings, CHO-K1 cells experienced an aggravated SSC toxicity upon a GSH decrease triggered by buthionine sulfoximine, while control cells (without SSC treatment) remained vital. These results underline the importance of GSH, though its deficit appeared not to be solely driving the toxicity. Intensified SSC toxicity further correlated with a hindered GS-SO3 reduction and export. Mitigation strategies to prevent the SSC toxicity through the addition of sodium carbonate, sodium glutamate and copper (II) sulfate to the feed were successful.
In the third part of the study, the CHO-K1, CHOZN and CHO-DG44 clones, exhibiting phenotypic heterogeneity, were explored using SSC at wide-ranging doses to provide complementary data. The CHO-DG44 clone showed the lowest SSC sensitivity that was linked to a putative lower SSC uptake capacity, as well as a higher reactive sulfur and oxygen species scavenging capacity. This SSC-related knowledge regarding its dual nature paves the way for further research, and may be transferred to other clones that may respond differently to this modified amino acid
Integrated Approach for the Adjustment of Disruption Programs to the Actual Infrastructural Availability
Full text linkSo-called disruption programs (DRP) are used to deal with suddenly occurring disruptions in commuter railway systems. Since the creation of DRPs is a complicated task, they regularly to support disruptions that result due to the failure of a given set of infrastructural elements (e.g. signal boxes). In case a disruption affects a combination of infrastructural elements that are not explicitly covered by available DRPs, their benefits would be highly affected. Therefore, it becomes necessary to derive approaches that allow an ad-hoc adjustment of the DRPs to the actual infrastructure availability.
This article proposes an approach supporting the adjustment of a manually chosen DRP to the actual infrastructural availability. The approach performs the adjustments by means of a heuristic method. The heuristic method uses a local search to derive the necessary adjustments on the operational measures in the operating concept of the chosen DRP that are conflicting with the actual infrastructural availability. The local search method allows to minimize the extent of the adjustments, as it is considered that the optimal solution resides in the vicinity of the chosen DRP. Ultimately, the adjusted operating concept alternatives generated by the heuristic are assessed to ensure their operational and passenger-transport feasibilities
Cones of orthogonal Shimura subvarieties and equidistribution
Full text linkLet X be an orthogonal Shimura variety, and let Cᵣᵒʳᵗ(X) be the cone generated by the cohomology classes of orthogonal Shimura subvarieties in X of dimension r. We investigate the asymptotic properties of the generating rays of Cᵣᵒʳᵗ(X) for large values of r. They accumulate towards rays generated by wedge products of the Kähler class of X and the fundamental class of an orthogonal Shimura subvariety. We also compare Cᵣᵒʳᵗ(X) with the cone generated by the special cycles of dimension r. The main ingredient to achieve the results above is the equidistribution of orthogonal Shimura subvarieties
A Note on Alexandrov Immersed Mean Curvature Flow
Full text linkWe demonstrate that the property of being Alexandrov immersed is preserved along mean curvature flow. Furthermore, we demonstrate that mean curvature flow techniques for mean convex embedded flows such as noncollapsing and gradient estimates also hold in this setting. We also indicate the necessary modifications to the work of Brendle–Huisken to allow for mean curvature flow with surgery in the Alexandrov immersed, 2-dimensional setting
Der explizite Leser : Kognitionswissenschaftliche Herausforderungen für die Literaturwissenschaft zwischen Korpus- und Rezeptionsanalyse
Full text linkProduktion, Text und Rezeption sind die Gegenstandsfelder der klassischen, hermeneutischen Literaturwissenschaft. Der Beitrag argumentiert, dass diese Felder auch für die Digitale Literaturwissenschaft zusammengehören. Ergebnisse von Untersuchungen im data rich approach, der Korpusanalysen mit der Analyse von Daten aus der Institutionengeschichte der Literatur verbindet, erhärten die Notwendigkeit, sich stärker mit der Rezeptionsseite der Literatur zu befassen. Dazu gehört die Durchführung von Experimenten, deren empirische Daten in die Daten-Systematik der Literaturwissenschaft integriert werden müssen. Für diese Perspektive greift die rezeptionsästhetische Vorstellung vom ›impliziten Leser‹, der auf Textstrukturen nur reagiert, zu kurz. Vor dem Hintergrund neuer Evidenz aus der Kognitionswissenschaft wird gezeigt, dass Lesen immer auch der Eigengesetzlichkeit der dabei beteiligten kognitiven Prozesse unterliegt. Der ›explizite Leser‹ bietet die Möglichkeit, zentrale interdisziplinäre Fragen wie die nach der Funktionsweise von Aufmerksamkeit, nach der Wirkung von Narrativen oder nach den speziellen Wahrscheinlichkeitskalkülen der Literatur neu zu fassen und dies bei der Analyse zu berücksichtigen
Executing Ad-Hoc Queries on Large Geospatial Data Sets Without Acceleration Structures
Full text linkIn this case study, we investigate if it is possible to harness the capabilities of modern commodity hardware to perform ad-hoc queries on large raw geospatial data sets. Normally, this requires building an index structure, which is a time-consuming process. We aim to provide means to individual users who receive a new or updated geospatial data set and want to directly start working with it without having to build such an index structure first. To this end, we conduct various experiments on two distinct types of data: 3D building models and point clouds. For the former, we demonstrate that well-known algorithms such as fast string search allow a wide range of queries to be answered in at most a few seconds on data sets with over a million buildings. The usage of progressive indexing additionally improves query run time by more than a factor of two. Regarding point clouds, we achieve similar run times using the popular LAS file format and a query throughput of up to a billion points per second when using a columnar memory layout. The run time of ad-hoc queries is often on par with that of database-driven solutions, sometimes even outperforming them. Considering that ad-hoc queries require no preprocessing, our results show that they are a viable alternative to acceleration structures when working with geospatial data
Enzymatic synthesis of semi-IPNs within hydrogel-based microfluidics
Full text linkWith the goal of achieving environmentally friendly polymer synthesis strategies, enzyme-promoted polymerisation has gradually attracted people's attention. The development of hydrogel-based microfluidics provides a new carrier system for enzymatic catalysis. Here, we report a new technique for enzyme-promoted free radical polymerisation, supported on hydrogel microdots (μHDs) within a microfluidic chip. Free radical polymerisation initiated by free horseradish peroxidase (HRP) in vials confirmed the formation of poly(N-isopropyl acrylamide) (PNiPAAm), achieving high molecular weight (500 000 Da) in 5 min. For polymerisation in microfluidics, disulphide-bearing μHDs were mounted on a PDMS-on-glass chip. Utilising a disulphide-thiol exchange reaction, modified HRP was then captured “from the flow” through the chip, which was confirmed by fluorescence microscopy. Various polymerisation parameters were studied in the microfluidic chip, and the successful polymer formation was confirmed by copolymerisation with a fluorescent comonomer. The physical entanglement fixed the formed polymer on the μHDs, forming a structure similar to a semi-interpenetrating network (semi-IPN). Thus, this technique provides a new direct approach to achieving semi-IPNs within microfluidic chips, showcasing the versatility in which microfluidic systems can be utilised