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The development of photosynthesis was a highlight in the progression of bacteria. In addition to the photosystems with their structural proteins, the photosynthesis apparatus consists of different cofactors including essential carotenoids. Thus, the evolution of the carotenoid pathways in relation to the functionality of the resulting structures in photosynthesis is the focus of this review. Analysis of carotenoid pathway genes indicates early evolutionary roots in prokaryotes. The pathway complexity leading to a multitude of structures is a result of gene acquisition, including their functional modifications, emergence of novel genes and gene exchange between species. Along with the progression of photosynthesis, carotenoid pathways coevolved with photosynthesis according to their advancing functionality. Cyanobacteria, with their oxygenic photosynthesis, became a landmark for evolutionary events including carotenogenesis. Concurrent with endosymbiosis, the cyanobacterial carotenoid pathways were inherited into algal plastids. In the lineage leading to Chlorophyta and plants, carotenoids evolved to their prominent role in protection and regulation of light energy input as constituents of a highly efficient light-harvesting complex.
Chondrules are thought to play a crucial role in planet formation, but the mechanisms leading to their formation are still a matter of unresolved discussion. So far, experiments designed to understand chondrule formation conditions have been carried out only under the influence of terrestrial gravity. In order to introduce more realistic conditions, we developed a chondrule formation experiment, which was carried out at long-term microgravity aboard the International Space Station. In this experiment, freely levitating forsterite (Mg2SiO4) dust particles were exposed to electric arc discharges, thus simulating chondrule formation via nebular lightning. The arc discharges were able to melt single dust particles completely, which then crystallized with very high cooling rates of >105 K h−1. The crystals in the spherules show a crystallographic preferred orientation of the [010] axes perpendicular to the spherule surface, similar to the preferred orientation observed in some natural chondrules. This microstructure is probably the result of crystallization under microgravity conditions. Furthermore, the spherules interacted with the surrounding gas during crystallization. We show that this type of experiment is able to form spherules, which show some similarities with the morphology of chondrules despite very short heating pulses and high cooling rates.
Redirection of the transcription factor SP1 to AT rich binding sites by a synthetic adaptor molecule
(2021)
The ubiquitous transcription factor SP1 binds to a GC rich consensus sequence. Here we describe an adaptor molecule that mediates binding of SP1 to a non-cognate DNA site rich in AT. The adaptor is comprised of a Dervan-type hairpin polyamide with high affinity to an AT rich hexamer duplex. It also carries a 27mer DNA that contains the SP1 consensus sequence. The synthesis and purification of the polyamide-DNA conjugate is reported. Pulldown experiments and western blot analysis demonstrate adaptor mediated binding of SP1 to the hexamer duplex TTGTTA.
Photoactivatable compounds for example photoswitches or photolabile protecting groups (PPGs, photocages) for spatiotemporal light control, play a crucial role in different areas of research. For each application, parameters such as the absorption spectrum, solubility in the respective media and/or photochemical quantum yields for several competing processes need to be optimized. The design of new photochemical tools therefore remains an important task. In this study, we exploited the concept of excited-state-aromaticity, first described by N. Colin Baird in 1971, to investigate a new class of photocages, based on cyclic, ground-state-antiaromatic systems. Several thio- and nitrogen-functionalized compounds were synthesized, photochemically characterized and further optimized, supported by quantum chemical calculations. After choosing the optimal scaffold, which shows an excellent uncaging quantum yield of 28 %, we achieved a bathochromic shift of over 100 nm, resulting in a robust, well accessible, visible light absorbing, compact new photocage with a clean photoreaction and a high quantum product (ϵ⋅Φ) of 893 M−1 cm−1 at 405 nm.
Background: To test the effect of urological primary cancers (bladder, kidney, testis, upper tract, penile, urethral) on overall mortality (OM) after secondary prostate cancer (PCa). Methods: Within the Surveillance, Epidemiology and End Results (SEER) database, patients with urological primary cancers and concomitant secondary PCa (diagnosed 2004-2016) were identified and were matched in 1:4 fashion with primary PCa controls. OM was compared between secondary and primary PCa patients and stratified according to primary urological cancer type, as well as to time interval between primary urological cancer versus secondary PCa diagnoses. Results: We identified 5,987 patients with primary urological and secondary PCa (bladder, n = 3,287; kidney, n = 2,127; testis, n = 391; upper tract, n = 125; penile, n = 47; urethral, n = 10) versus 531,732 primary PCa patients. Except for small proportions of Gleason grade group and age at diagnosis, PCa characteristics between secondary and primary PCa were comparable. Conversely, proportions of secondary PCa patients which received radical prostatectomy were smaller (29.0 vs. 33.5%), while no local treatment rates were higher (34.2 vs. 26.3%). After 1:4 matching, secondary PCa patients exhibited worse OM than primary PCa patients, except for primary testis cancer. Here, no OM differences were recorded. Finally, subgroup analyses showed that the survival disadvantage of secondary PCa patients decreased with longer time interval since primary cancer diagnosis. Conclusions: After detailed matching for PCa characteristics, secondary PCa patients exhibit worse survival, except for testis cancer patients. The survival disadvantage is attenuated, when secondary PCa diagnosis is made after longer time interval, since primary urological cancer diagnosis.
Two species, namely Russula adwanitekae A.Ghosh, K.Das & Buyck sp. nov. and Russula purpureozonata K.Das, A.Ghosh & Buyck sp. nov. are proposed herein as new mushroom taxa from the Indian Himalaya based on their morphological features and ITS-based phylogenetic inferences. Both species belong to the crown clade of Russula subgenus Russula but with affinities to different subsections, viz. subsect. Laricinae Romagn. and subsect. Decolorantes Maire, respectively. Russula adwanitekae sp. nov. was collected in mixed temperate forests where it is most likely associated with conifers. It is distinct from several similarly looking, small, mild species with dark spore print and reddish lilac, orchid purple or greyish to deep magenta colored pileus in subsect. Laricinae by its sequence data (nrITS) or geographic distribution. Russula purpureozonata sp. nov. associates with Abies densa Griff., and possesses all typical features of Decolorantes, viz. the amyloid suprahilar spot on the spores, presence of pileogloeocystidia, the reddening then blackening context, equal lamellae and colored spore print. It reminds of North American R. californiensis Burl. and R. magna Beardslee under the microscope but has distinctly smaller spores and differs further in the unique coloration and concentrically zonated pileus margin. Macro- and micromorphological features are illustrated for both species. Their habitats, distributions and relationships with allied species are discussed.
This paper is the conclusion of the “Spaghetti Project” aiming to revise French species of Terebellidae sensu lato (s.l.) belonging to the five families: Polycirridae, Telothelepodidae, Terebellidae sensu stricto (s.s.), Thelepodidae and Trichobranchidae. During this project, 41 species were observed, 31 of them new for science: eight species of Polycirridae, eleven species of Terebellidae s.s., three species of Thelepodidae and nine species of Trichobranchidae. We provide a comprehensive key for all European species of terebellids with a focus on the important diagnostic characters for each family. Finally, we discuss issues on taxonomy, biodiversity and cryptic and pseudo-cryptic species of polychaetes in European waters, based on results obtained during this project.
The problem of unconstrained or constrained optimization occurs in many branches of mathematics and various fields of application. It is, however, an NP-hard problem in general. In this thesis, we examine an approximation approach based on the class of SAGE exponentials, which are nonnegative exponential sums. We examine this SAGE-cone, its geometry, and generalizations. The thesis consists of three main parts:
1. In the first part, we focus purely on the cone of sums of globally nonnegative exponential sums with at most one negative term, the SAGE-cone. We ex- amine the duality theory, extreme rays of the cone, and provide two efficient optimization approaches over the SAGE-cone and its dual.
2. In the second part, we introduce and study the so-called S-cone, which pro- vides a uniform framework for SAGE exponentials and SONC polynomials. In particular, we focus on second-order representations of the S-cone and its dual using extremality results from the first part.
3. In the third and last part of this thesis, we turn towards examining the con- ditional SAGE-cone. We develop a notion of sublinear circuits leading to new duality results and a partial characterization of extremality. In the case of poly- hedral constraint sets, this examination is simplified and allows us to classify sublinear circuits and extremality for some cases completely. For constraint sets with certain conditions such as sets with symmetries, conic, or polyhedral sets, various optimization and representation results from the unconstrained setting can be applied to the constrained case.
My PhD work employed genetic and pharmacological manipulations, coupled with highresolution live imaging, to understand intercellular communications during zebrafish cardiovascular development. The heart is the first organ to form, and it is composed of several tissues, among which interactions are crucial. I identified two important interactions between muscular and non-muscular tissues in poorly characterized contexts, and the molecules required for the signalling. First, I discovered an important cellular and molecular crosstalk orchestrating the development of the cardiac outflow tract (i.e., the aortic root in mammals).
Endothelial-derived TGF-beta signalling controls the generation of the local extracellular matrix (ECM). The ECM in turn affects endothelial proliferation as well as smooth muscle cell organization (Boezio et al, 2020; Bensimon-Brito*, Boezio* et al, 2020). In my second project, I investigated the crosstalk between the epicardial layer and the myocardial wall. By generating epicardial-impairment models, I identified a novel role for the epicardium in regulating cardiomyocyte volume during heart development (Boezio et al, 2021). Ultimately, this research contributed to our understanding of how paracrine signalling controls the multicellular interactions integral to organogenesis.
Die Bildung von Blutgefäßen ist essentiell für die Entwicklung und Homöostase von Wirbeltieren und die Endothelzellspezifikation ist ein wichtiger erster Schritt in diesem Prozess. Das früheste bekannte Ereignis bei der Endothelzellspezifikation im Zebrafisch ist die Expression des bHLH-PAS-Transkriptionsfaktor-Gens npas4l. Ich habe eine transgene V5-Linie zum Nachweis des markierten Npas4l auf Proteinebene und eine Gal4-VP16-Reporterlinie zur Visualisierung und Verfolgung von npas4l exprimierenden Zellen in vivo generiert. Beide Linien können bereits in frühen Entwicklungsstadien nachgewiesen werden und komplementieren auch starke npas4l-Mutanten Allele. Um npas4l Reporter exprimierende Zellen in npas4l Mutanten zu verfolgen, habe ich anschließend eine mutierte Variante der Gal4-Reporterlinie erzeugt. Diese Mutante trägt eine Insertion in der Region, die die DNA-Bindedomäne kodiert. Dadurch stört sie die Npas4l-Funktion, aber nicht die Reporterexpression. Dieses mutierte Reporterallel komplementiert nicht die npas4l-Mutanten und zeigt einen starken Phänotyp, was darauf hindeutet, dass es sich um ein funktionelles Nullallel handelt. Phänotypische Analysen zeigten, dass npas4l-Reporter positive Zellen in npas4l-Mutanten nicht spezifizieren oder zur Mittelachse wandern. Stattdessen tragen sie zu den vom intermediären Mesoderm abgeleiteten pronephrischen Tubuli und dem vom paraxialen Mesoderm abgeleiteten Skelettmuskel bei. Ich habe diese Phänotypen durch Einzelzell-RNAseq an den npas4l-Reporter positiven Zellen in npas4l+/- und npas4l-/- Embryonen bestätigt. Zusammen erklären diese beiden alternativen Zellschicksale den Großteil der beobachteten Veränderungen zwischen den Genotypen. Npas4l ist dafür bekannt die Expression der drei Transkriptionsfaktorgene etsrp, tal1 und lmo2 zu fördern. Ich stellte die Hypothese auf, dass das Fehlen jedes dieser Transkriptionsfaktoren in npas4l-Mutanten verschiedene Aspekte des npas4l-Phänotyps verursacht. Daher habe ich Mutantenlinien für alle drei Gene generiert und sie sowohl in vaskulären Reporterlinien als auch im npas4l-Reporterhintergrund analysiert. Die Daten legen nahe, dass verschiedene Gene unterschiedliche Prozesse während der frühen Endothelentwicklung regulieren. In npas4l-/- und etsrp-/- Embryonen differenzieren npas4l-Reporter exprimierende Zellen nicht zu Endothelzellen und tragen stattdessen zur Skelettmuskelzellpopulation bei. In npas4l-/- und tal1-/- Embryonen können npas4l-Reporter exprimierende Zellen nicht migrieren und tragen stattdessen zu der Bildung der pronephrischen Tubuli bei. Um die Beziehung zwischen diesen Faktoren besser zu verstehen, habe ich getestet, ob die Injektion von etsrp-, tal1- oder lmo2-mRNA verschiedene Aspekte des npas4l-Phänotyps retten würde. npas4l-, etsrp- und tal1-Mutanten zeigen alle schwere vaskuläre Phänotypen. Einige Endothelzellen und vaskuläre Strukturen bleiben jedoch in jeder Mutante erhalten. Der Phänotyp ist am stärksten in npas4l-/- Embryonen, aber selbst in diesen Embryonen können einige fli1a-positive Endothelzellen in der Schwanzregion beobachtet werden. Es war unklar, ob sich diese Population von Endothelzellen unabhängig von der Npas4l-, Tal1- und Etsrp-Funktion entwickelt oder als Folge einer restlichen tal1- oder etsrp-Expression unabhängig von Npas4l. Um diese Frage zu untersuchen, habe ich Doppelmutanten generiert und nach dem Vorhandensein von fli1a-positiven Endothelzellen in diesen Mutanten gesucht. Während fli1a-positive Endothelzellen in npas4l-/- und npas4l-/-;tal1-/- Embryonen deutlich vorhanden sind, können keine solchen Zellen in npas4l-/-;etsrp-/- oder etsrp-/-;tal1-/- Embryonen beobachtet werden. Diese Daten deuten darauf hin, dass sich im Zebrafisch keine Endothelzellen entwickeln können, wenn zugleich npas4l und etsrp oder etsrp und tal1 gestört sind. Während der Verlust von etsrp zu stärkeren Defekten in npas4l-Mutanten führt, gibt es keinen zusätzlichen Phänotyp, der durch den Verlust von tal1verursacht wird, was darauf hindeutet, dass die Expression von etsrp, aber nicht die von tal1, unabhängig von Npas4l auftreten kann. Diese Idee wird durch die Beobachtung unterstützt, dass etsrp, aber nicht tal1-Expression in den meisten fli1a-exprimierenden Zellen in npas4l-/- Embryonen beobachtet wird. Dennoch wird der Großteil -Expression durch Npas4l reguliert. tal1-mRNA-Injektionen reichten aus, um eine Wildtyp-ähnliche vaskuläre Musterbildung im Bauchbereich der npas4l-/- Embryonen wiederherzustellen, einschließlich der Rettung sowohl der Zellmigration als auch der Differenzierung. Da Npas4l mehrere unterschiedliche transkriptionelle Effektoren hat, war eine so starke Rettung durch nur einen dieser Effektoren unerwartet. In den geretteten Mutanten wurde die bilaterale Population von npas4l-Reporter-positiven pronephrischen Tubuluszellen nicht entdeckt, aber die Anzahl der ektopischen npas4l-Reporter exprimierenden Muskelzellen war im Vergleich zu nicht injizierten npas4l-Mutanten gleichbleibend.
...
It is increasingly recognized that neuroscience has not delivered the revolutionary clinical possibilities for psychiatry that had been promised. Explanations differ, however: some proponents emphasize the divide between biopsychosocial psychiatry and mechanistic neurology. Others rely on further basic experimental neuroscience as only the most elementary level of explanation will allow us to fully understand and treat mental disorders. From a clinical-neuropsychological perspective, I shall argue that both views are mistaken. Diagnosis and treatment of neurological diseases demands a biopsychosocial perspective similar to psychiatry. Acknowledging this might help to bring both disciplines together and improve clinical outcome.
Multi-view microscopy techniques are used to increase the resolution along the optical axis for 3D imaging. Without this, the resolution is insufficient to resolve subcellular events. In addition, parts of the images of opaque specimens are often highly degraded or masked. Both problems motivate scientists to record the same specimen from multiple directions. The images, then have to be digitally fused into a single high-quality image. Selective-plane illumination microscopy has proven to be a powerful imaging technique due to its unsurpassed acquisition speed and gentle optical sectioning. However, even in the case of multi view imaging techniques that illuminate and image the sample from multiple directions, light scattering inside tissues often severely impairs image contrast.
Here we show that for c-elegans embryos multi view registration can be achieved based on segmented nuclei. However, segmentation of nuclei in high density distribution like c-elegans embryo is challenging. We propose a method which uses 3D Mexican hat filter for preprocessing and 3D Gaussian curvature for the post-processing step to separate nuclei. We used this method successfully on 3 data sets of c-elegans embryos in 3 different views. The result of segmentation outperforms previous methods. Moreover, we provide a simple GUI for manual correction and adjusting the parameters for different data.
We then proposed a method that combines point and voxel registration for an accurate multi view reg- istration of c-elegans embryo, which does not need any special experimental preparation. We demonstrate the performance of our approach on data acquired from fixed embryos of c-elegans worms. This multi step approach is successfully evaluated by comparison to different methods and also by using synthetic data. The proposed method could overcome the typically low resolution along the optical axis and enable stitching to- gether the different parts of the embryo available through the different views. A tool for running the code and analyzing the results is developed.
Neurons are cells with a highly complex morphology; their dendritic arbor spans up to thousands of micrometers. This extended arbor poses a challenge for the logistics of neuronal processes: mRNA, proteins, and organelles have to be transported to dendrites, hundreds of micrometers away from the soma. This thesis aims to calculate the minimum number of proteins needed to populate the dendritic trees for different scenarios.
In chapter 2, I analyzed the ability of different mechanisms to populate the dendritic arbor. I started from the solution of the diffusion equation in Sec. 2.1, then I included the contribution of active transport in Sec. 2.2 and showed how it could have either the effect of increasing the effective diffusion coefficient or of introducing a bias in the diffusion process. In Sec. 2.3 I studied the spatial distribution of locally synthesized protein, accordingly with actively and passively transported mRNA. In Sec. 2.5, I derived the boundary condition for branches showing a qualitatively different behavior of surface and cytoplasmic proteins induced by the medium’s dimensionality in which they diffuse.
In chapter 3, I introduced the concept of protein requirement, defined as the minimum number of proteins that the neuron needs to produce to provide at least one protein to each micrometer of the dendritic arbor. In Sec. 3.1, I derived the protein requirement for diffusive proteins for somatic translation and constant translation in the dendritic arbor. In Sec. 3.2, I analyzed numerically the protein requirement in the case of actively transported protein synthesized in the soma, and, in Sec. 3.3, in the case of actively transported proteins synthesized in the dendritic arbor. In Sec. 3.4, I analyzed the protein requirement of protein synthesized in the dendrite accordingly with the distribution of mRNA described in Sec. 3.3 and 3.2. In Sec. 3.5, I derived the protein requirement for a single branch and purely diffusive proteins.
In chapter 4, I analyzed the relation between the radii of the three afferent dendrites in a branch, their length, and the diffusion length of a protein. In Sec. 4.1 I derived the optimal ratio between the radii of the daughter dendrites that minimizes the protein requirement. In Sec. 4.3 I introduced the 3/2− Rall Rule and in Sec. 4.5 its generalization. Finally, I used those rules to estimate the fraction of proteins diffusing away from and toward the soma.
In chapter 5, I analyzed the radii distribution for three categories of neurons: cultured hippocampal neurons in Sec. 5.1, stomatogastric ganglia neuron in Sec. 5.2, and 3DEM reconstructed prefrontal pyramidal neurons in Sec. 5.3. For each of these three classes, I analyzed the distribution of radii, Rall exponents, and the probability ratio. For most of them, I found that the probability of a protein diffusing away from the soma is higher for surface proteins than for cytoplasmic ones. I quantified this with a parameter called surface bias.
In Chapter 6, I analyzed the fluorescent ratio imaged by our collaborators Anne-Sophie Hafner, for a surface protein, GFP::Nlg, and a soluble one, GFP, in cultured hippocampal neurons, and I compared the fluorescent ratio with the probability ratio obtained in 5.1, finding that they are in good agreement.
In chapter 7, I compared the real dendritic morphologies imaged by one of our collaborators Ali Karimi with the optimal branching rule obtained in Sec. 4.1 and I calculated the cost for not having optimal branching radii.
Finally, in Chapter 8, I used the knowledge of the branching statistics gathered in 5.3 to simulate the protein profile on three different classes of neurons: pyramidal neurons, granule neuron, and Purkinje neurons. I compared the protein profile for surface and cytoplasmic neurons for each morphology for two different values of the diffusion length: λ = 109µm and λ = 473µm, both for optimized radii and symmetrical radii. I showed how the radii optimization reduces the protein requirement of a factor 10 4 for pyramidal neurons.
Sampling of remote inland aquatic habitats in South Africa has constantly been yielding novel endemic freshwater crab species (Potamonautes MacLeay,1838). During the present study, we report on the discovery and description of two new freshwater crab species (Potamonautes baziya sp. nov., and P. mariepskoppie sp. nov.) from Afrotemperate forested mountain regions in the Eastern Cape and Mpumalanga provinces of South Africa, respectively. Phylogenetic evidence derived from DNA sequence data of three partial mitochondrial loci (12S rRNA, 16S rRNA and cytochrome oxidase subunit one, COI) corroborates the evolutionary distinction of the two novel species. In addition, morphological and ecological data for the two new species further delineate their evolutionary distinction from congeneric sister species. A comparison of the taxonomically important gonopods 1 and 2 and carapace features among the sister species and other known freshwater crabs of South Africa was further used to provide evidence for the distinction of the two novel species. The discovery of two new species suggest that remote mountainous areas or unsampled regions in South Africa likely harbor several novel species, reiterating a call to document aquatic inland biodiversity in forested and mountainous regions of the country.
Diese Arbeit ist ein detaillierter Bericht über die Forschungsaktivitäten, die ich während meiner Promotion am Max-Planck-Institut für Biophysik durchgeführt habe. Mit dem Aufkommen der direkten Elektronendetektoren erlebte die Transmissionselektronenmikroskopie von gefrorenen hydratisierten Proben (Kryo-EM) einen epochalen Wandel, die sogenannte “Auflösungsrevolution”. Ab den 2010er Jahren ermöglichte die Kommerzialisierung der ersten direkten Detektoren die Erforschung biologischer Phänomene in beispiellosem Detail und machte Kryo-EM zu einer der leistungsstärksten (und gefragtesten) Forschungsmethoden in den Biowissenschaften. Meine Forschung konzentrierte sich auf die Verwendung der Elektronen-Kryotomographie, um zwei herausfordernde Ziele zu erreichen. Das erste bestand darin, die Denaturierung von Proteinen an der Luft-Wasser-Grenzfläche zu untersuchen, und das zweite die molekulare Landschaft eines lichtempfindlichen Chloroplastenvorläufers, des Etioplasten, zu beschreiben. Um die Relevanz, Herausforderungen und Auswirkungen meiner Arbeit zu vermitteln, habe ich diese Arbeit in drei Kapitel unterteilt.
Kapitel eins enthält eine Einführung in die Transmission-Elektronenmikroskopie.
Nach einer kurzen Zusammenfassung der historischen Meilensteine in der Disziplin beschreibe ich die wesentlichen Komponenten des TEM und deren Funktionsweise. Hier lege ich besonderen Wert auf die Struktur elektromagnetischer Linsensysteme, wie sie den Weg der Elektronen beim Durchlaufen der Säule beeinflussen und wie Bilder entstehen. Der hardwarebezogene Teil der Einführung wird durch eine vereinfachte Beschreibung der Elektronendetektoren abgeschlossen, in der ich die revolutionären Aspekte der direkten Elektronendetektoren, mit der Struktur und Funktion von CCD-Detektoren (Charge Coupled Device detector) vergleiche. Als nächstes konzentriere ich mich auf die theoretischen Prinzipien der Bilderzeugung. Um die Hauptphänomene im Zusammenhang mit der Bildqualität in TEM hervorzuheben, stelle ich grundlegende Konzepte wie den Einfluss von Elektronenenergie und optischen Aberrationen vor, gefolgt von einer ausführlicheren Beschreibung des Ursprungs von Kontrast und Rauschen. Der Unterabschnitt schließt mit einigen Überlegungen darüber, wie - und vor allem wie effizient - Detektoren kontinuierliche Elektronenwellen in diskrete Bereiche (Pixel) abtasten. Der folgende Unterabschnitt ist der Erfassung und Verarbeitung tomografischer Daten gewidmet. Hier gebe ich eine vereinfachte Beschreibung, wie Kippserien mit dem Mikroskop erfasst werden und wie die Rohdaten zu einer dreidimensionalen Darstellung der Probe verarbeitet werden. Der Einfluss der Neigungsgeometrie und der Dosisverteilung auf die Rekonstruktionsqualität wird ebenfalls diskutiert. Der zweite Teil des Unterabschnitts befasst sich mit der Strukturbestimmung durch Subtomogramm-Mittelung und der Errechnung der Auflösung von Kryo-EM-Rekonstruktion. Zuletzt schließe ich das Kapitel mit einer Beschreibung der Vorbereitung biologischer Proben für die Kryo-EM-Bildgebung mit einigen abschließenden Bemerkungen zur Dynamik und den Grenzen der Vitrifizierung ab.
Kapitel zwei folgt dem Thema der Kryo-Präparation biologischer Proben mit der Untersuchung der Denaturierung von Proteinen an der Luft-Wasser-Grenzfläche.
Im Einführungsabschnitt skizziere ich die wichtigsten Aspekte dieses Phänomens. Frühe Experimente zum Verhalten von Proteinen in Lösung zeigten ihre Neigung, aus der Lösung zu ihrer Grenzfläche mit der Atmosphäre zu diffundieren. Hier bilden sie meist unlösliche Schichten denaturierter Fibrillen Es wurde vorgeschlagen, dass die Korrelation zwischen Proteindenaturierung und Kontakt mit der Grenzfläche auf einen allmählichen Entfaltungsprozess zurückzuführen ist, bei dem Tausende von Wechselwirkungen pro Sekunde zu einer immer größeren strukturellen Schädigung führen würden. Ein direkter Beweis für diesen Mechanismus wurde jedoch nie dokumentiert. Um einen tieferen Einblick in die Dynamik an der Luft-Wasser-Grenzfläche zu erhalten, sammelte ich Kryotomogramme vitrifizierter Präparate der Fettsäuresynthase (FAS, Fatty Acid Synthase) aus Hefe. Im ersten Unterabschnitt der Ergebnisse beschreibe ich, wie die biochemische und Negativkontrastierung-TEM-Analyse von FAS-Fraktionen zeigte, dass der Komplex während des gesamten Reinigungsverfahrens intakt und katalytisch aktiv blieb. Nach der Vitrifizierung ergab die Einzelpartikelanalyse jedoch, dass 90% aller Komplexe stark beschädigt waren. Die tomographische Rekonstruktion derselben Proben zeigte, dass alle FAS-Komplexe an die Luft-Wasser-Grenzfläche gebunden waren. Die Seite des Moleküls, die der Grenzfläche ausgesetzt war, schien abgeflacht zu sein, während die Seite, in der wässrigen Phase, ihre native Struktur beibehielt. Die Mittelung der Subtomogramme bestätigte, dass eine Seite von fast 90% der Partikel stark beschädigt war. Durch den Vergleich der Ausrichtung dieser beschädigten Seite mit der Position eines Rechenmodells der Luft-Wasser-Grenzfläche konnte ich nachweisen, dass sie perfekt übereinstimmen, was den ersten direkten Beweis dafür liefert, dass die Wechselwirkung mit der Luft-Wasser-Grenzfläche die lokale Denaturierung großer Proteinkomplexe herbeiführt.
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The US Tax Cuts and Jobs Act (TCJA) led to a drastic reduction in the corporate tax and improved the treatment of C corporations compared to S corporations. We study the differential effect of the TCJA on these types of corporations using key economic variables of US banks, such as the number of employees, average salaries and benefits, profit/loss before taxes, and net income. Our analysis suggests that the TCJA increased the net-of-tax profits of C corporation banks compared to S corporations and, to a lesser extent, their pre-tax profits. At the same time, the reform triggered no significantly differential effect on the employment and average wages.
Historically Central Bank Independence (CBI) was anything but the norm. CBI seems to contradict core principles of democracy. Most economists were also against CBI. After the Great Inflation of the 1970ies many empirical studies demonstrated that there is a strong negative correlation between the degree of CBI and the rate of inflation. In 1990 most major countries had endowed their central bank with the status of independence. Overburdening with elevated expectations and additional competences are threatening the reputation of central banks and undermining the case for CBI.