79 research outputs found
Norbert Grob, Bernd Kiefer, Isabelle Louise Bastian (Hg.): Bruch der Weltenlinie: Zum Kino der Moderne. Essays â PortrĂ€ts â Hommagen
Kommunales Integrationsmanagement - deutsch-französische Konsultationen in Stuttgart: Tagungsbericht
Characterization and Purification of Polydisperse Reconstituted Lipoproteins and Nanolipoprotein Particles
Heterogeneity is a fact that plagues the characterization and application of many self-assembled biological constructs. The importance of obtaining particle homogeneity in biological assemblies is a critical goal, as bulk analysis tools often require identical species for reliable interpretation of the resultsâindeed, important tools of analysis such as x-ray diffraction typically require over 90% purity for effectiveness. This issue bears particular importance in the case of lipoproteins. Lipid-binding proteins known as apolipoproteins can self assemble with liposomes to form reconstituted high density lipoproteins (rHDLs) or nanolipoprotein particles (NLPs) when used for biotechnology applications such as the solubilization of membrane proteins. Typically, the apolipoprotein and phospholipids reactants are self assembled and even with careful assembly protocols the product often contains heterogeneous particles. In fact, size polydispersity in rHDLs and NLPs published in the literature are frequently observed, which may confound the accurate use of analytical methods. In this article, we demonstrate a procedure for producing a pure, monodisperse NLP subpopulation from a polydisperse self-assembly using size exclusion chromatography (SEC) coupled with high resolution particle imaging by atomic force microscopy (AFM). In addition, NLPs have been shown to self assemble both in the presence and absence of detergents such as cholate, yet the effects of cholate on NLP polydispersity and separation has not been systematically examined. Therefore, we examined the separation properties of NLPs assembled in both the absence and presence of cholate using SEC and native gel electrophoresis. From this analysis, NLPs prepared with and without cholate showed particles with well defined diameters spanning a similar size range. However, cholate was shown to have a dramatic affect on NLP separation by SEC and native gel electrophoresis. Furthermore, under conditions where different sized NLPs were not sufficiently separated or purified by SEC, AFM was used to deconvolute the elution pattern of different sized NLPs. From this analysis we were able to purify an NLP subpopulation to 90% size homogeneity by taking extremely fine elutions from the SEC. With this purity, we generate high quality NLP crystals that were over 100 ÎŒm in size with little precipitate, which could not be obtained utilizing the traditional size exclusion techniques. This purification procedure and the methods for validation are broadly applicable to other lipoprotein particles
Isolation, Characterization, and Stability of Discretely-Sized Nanolipoprotein Particles Assembled with Apolipophorin-III
Background: Nanolipoprotein particles (NLPs) are discoidal, nanometer-sized particles comprised of self-assembled phospholipid membranes and apolipoproteins. NLPs assembled with human apolipoproteins have been used for myriad biotechnology applications, including membrane protein solubilization, drug delivery, and diagnostic imaging. To expand the repertoire of lipoproteins for these applications, insect apolipophorin-III (apoLp-III) was evaluated for the ability to form discretely-sized, homogeneous, and stable NLPs. Methodology: Four NLP populations distinct with regards to particle diameters (ranging in size from 10 nm to.25 nm) and lipid-to-apoLp-III ratios were readily isolated to high purity by size exclusion chromatography. Remodeling of the purified NLP species over time at 4uC was monitored by native gel electrophoresis, size exclusion chromatography, and atomic force microscopy. Purified 20 nm NLPs displayed no remodeling and remained stable for over 1 year. Purified NLPs with 10 nm and 15 nm diameters ultimately remodeled into 20 nm NLPs over a period of months. Intra-particle chemical cross-linking of apoLp-III stabilized NLPs of all sizes. Conclusions: ApoLp-III-based NLPs can be readily prepared, purified, characterized, and stabilized, suggesting their utilit
The Conformation of Interfacially Adsorbed Ranaspumin-2 Is an Arrested State on the Unfolding Pathway
Ranaspumin-2 (Rsn-2) is a surfactant protein found in the foam nests of the
t\'{u}ngara frog. Previous experimental work has led to a proposed model of
adsorption which involves an unusual clam shell-like `unhinging' of the protein
at an interface. Interestingly, there is no concomitant denaturation of the
secondary structural elements of Rsn-2 with the large scale transformation of
its tertiary structure. In this work we use both experiment and simulation to
better understand the driving forces underpinning this unusual process. We
develop a modified G\={o}-model approach where we have included explicit
representation of the side-chains in order to realistically model the
interaction between the secondary structure elements of the protein and the
interface. Doing so allows for the study of the underlying energy landscape
which governs the mechanism of Rsn-2 interfacial adsorption. Experimentally, we
study targeted mutants of Rsn-2, using the Langmuir trough, pendant drop
tensiometry and circular dichroism, to demonstrate that the clam-shell model is
correct. We find that Rsn-2 adsorption is in fact a two-step process: the
hydrophobic N-terminal tail recruits the protein to the interface after which
Rsn-2 undergoes an unfolding transition which maintains its secondary
structure. Intriguingly, our simulations show that the conformation Rsn-2
adopts at an interface is an arrested state along the denaturation pathway.
More generally, our computational model should prove a useful, and
computationally efficient, tool in studying the dynamics and energetics of
protein-interface interactions.Comment: 8 figure
Physical Activity, Bone Health, and Obesity in Peri-/Pre- and Postmenopausal Women: Results from the EPIC-Potsdam Study
Prevalence of vasculare risk factors in different stages of prodromal Alzheimerâs disease and its influence on cognitive decline
peer reviewe
Körperliche AktivitĂ€t und das Risiko fĂŒr chronische Erkrankungen : Entwicklung und Evaluierung eines Indexes zur Messung körperlicher AktivitĂ€t und Baseline-datenkalibrierung in EPIC-Deutschland
Körperliche AktivitĂ€t (KA) ist ein Lifestyle Faktor, der vor chronischen Erkrankungen zu schĂŒtzen scheint. In der European Prospective Investigation into Cancer and Nutrition (EPIC)-Studie wurden zwei Fragebögen verwendet, mit denen anhand des Cambridge Indexes und des Total Physical Activity Indexes die KA der Teilnehmer kategorisiert werden kann. Die prĂ€zise Messung der PA ist essentiell, um die wahre AssoziationsstĂ€rke zwischen KA und chronischen Erkrankungen zu schĂ€tzen. Objektive Messmethoden, die heutzutage zum Einsatz kommen, vereinfachen die schwierige Aufgabe der prĂ€zisen Messung der KA. Basierend darauf, wurden die folgenden Ziele in dieser Doktorarbeit verfolgt: die Entwicklung und Evaluierung eines validen KA Indexesâ den Improved Physical Activity Index (IPAI), welcher in der Lage sein soll, die KA der Personen zu kategorisieren aber auch eine kontinuierliche Messung der KA zu liefern. Diese soll das BewegungsausmaĂ widerspiegeln. Des Weiteren wurde eine Kalibrierung der Basis Fragebogen-Daten zur PA und die SchĂ€tzung des Zusammenhangs zwischen nicht-kalibrierter und kalibrierter KA und chronischen Erkrankungen, Typ 2 Diabetes, Myokardinfarkt, Schlaganfall und Krebs, angestrebt.
Diese Ziele wurden in den deutschen EPIC-Zentren Potsdam und Heidelberg verfolgt. In einer Substudie wurden 1615 Teilnehmer rekrutiert. Diese Substudie beinhaltete ein breites Fragenspektrum zur ĂŒblichen KA, sowie eine objektive 7-Tage Messung der KA mittels eines Herzfrequenz und Bewegungsmessers - Actiheart. Die Baseline Fragebogenangaben zur KA wurden, mittels der aus der Substudie gewonnenen Daten, kalibriert. Das Risiko fĂŒr chronische Erkrankungen wurde in der Gesamtkohorte mit Hilfe von proportionaler Hazards-Regression nach Cox berechnet.
Der IPAI besteht aus den folgenden AktivitĂ€tsvariablen: Art der BerufstĂ€tigkeit, Fahrrad fahren (Stunden/Woche), dem SporthĂ€ufigkeitsscore, sowie Fernsehscore und Computernutzung (Stunden/Wochenende ab 18Uhr). Korrelationen zwischen dem IPAI und objektiv gemessener KA betrugen r=0.39-0.44 fĂŒr AktivitĂ€tscounts und r=0.32-0.40 fĂŒr AktivitĂ€tsenergieausgabe (PAEE). Der Cambridge Index und der Total Physical Activity Index waren schwĂ€cher mit objektiv gemessener KA korreliert als der IPAI. In nicht-berufstĂ€tigen Teilnehmern war der IPAI auch stĂ€rker mit objektiv gemessener KA korreliert, als die bisher genutzten Indizes.
StĂŒckweise Regression wurde zur Kalibrierung der Baseleine KA Daten angewandt.
In den kalibrierten Daten waren die Hazardraten (HR (95% Konfidenzeinterval)) zwischen der höchsten KA Kategorie, im Vergleich zur niedrigsten KA Kategorie, niedriger als in den nicht kalibrierten Daten fĂŒr die meisten chronischen Erkrankungen HR=0.40(0.35-0.46), Typ 2 Diabetes HR=0.08(0.06-0.10), Myokardinfarkt HR=0.40(0.24-0.67) und Schlaganfall HR=0.54(0.33-0.87). Die nicht-kalibrierten HRs betragen, in der gleichen Reihenfolge: HR=0.72(0.66-0.80), HR=0.57(0.48-0.67), HR=0.62(0.44-0.88), und HR=0.86(0.62-1.19), Es wurden keine ZusammenhĂ€nge zwischen KA und Krebs gefunden, auch nicht nach Ausschluss von Erkrankungen, die in den ersten 3 Jahren der Nachbeobachtung aufgetreten sind.
Zusammenfassend zeigen die vorliegenden Ergebnisse, dass der IPAI ein valides Instrument zur populationsbezogenen subjektiven Messung der ĂŒblichen KA/Bewegung (sowohl kontinuierlich als auch in Kategorien) ist und in Studien zu diesem Zweck angewandt werden kann.
Die Datenkalibrierung liefert eine prĂ€zisere SchĂ€tzung des Zusammenhangs zwischen KA und chronischen Erkrankungen. Die prĂ€ventiven Effekte von KA auf chronische Erkrankungen, Typ 2 Diabetes, Myokardinfarkt und Schlaganfall werden in Studien mit Fragebogenmessung der KA unterschĂ€tzt. Interventionen im Bereich Public Health, vor allem in der Gruppe von Typ 2 Diabetes GefĂ€hrdeten, sollten einen stĂ€rkeren Fokus auf KA legen.Physical activity (PA) is a lifestyle factor that has been shown to prevent chronic diseases. The accurate PA measurement is essential to estimate the true magnitude of the relationship between PA and disease risk. Nowadays, objective measurement methods for PA have been developed and improve the challenging task of measuring the individualsâ PA level. Therefore, the aims of this thesis were firstly, the development and evaluation of a valid physical activity index â the Improved Physical Activity Index (IPAI), which will be able to categorize people into activity categories but may also be used as a continuous measure that reflects oneâs activity amount (movement). Secondly, the calibration of the available baseline PA questionnaire measurement and finally, the estimation of the associations between calibrated and non-calibrated baseline PA data and risk of overall chronic diseases, type 2 diabetes, myocardial infarction, stroke and overall cancer.
These objectives were accomplished by applying an extensive physical activity questionnaire and a 7-day heart rate and acceleration sensor PA measurement to a sub-sample (n=1,615) of older adults from the European Prospective Investigation into Cancer and Nutrition (EPIC-Germany) study. Baseline self-reported PA was calibrated using statistical models based on the objective PA measurement in the sub-sample. The risk of chronic diseases was estimated using Cox proportional hazards regression in the whole EPIC-Germany cohort.
The IPAI consists of items covering five areas including PA at work, sport, cycling, television viewing, and computer use. The correlations between the IPAI and accelerometer counts in the training and validation sample ranged from r=0.39 to 0.44 and with physical activity energy expenditure from r=0.32 to 0.40 and were higher than between the Cambridge Index or the Total Physical Activity Index with the objective measures. In non-working participants the IPAI also showed higher correlations than the established indices.
For the baseline PA data calibration segmented regression analysis has been chosen.
The hazard rates (HR (95% confidence intervals)) for the risk reduction in the highest calibrated PA category compared to the lowest for overall chronic diseases HR=0.40(0.35-0.46), type 2 diabetes HR=0.08(0.06-0.10), myocardial infarction HR=0.40(0.24-0.67) and stroke HR=0.54(0.33-0.87) were lower than the non-calibrated results HR=0.72(0.66-0.80), HR=0.57(0.48-0.67), HR=0.62(0.44-0.88), and HR=0.86(0.62-1.19), respectively. There were no associations between the calibrated PA and risk of cancer, even after excluding the first 3 years of follow up.
In conclusion, a valid PA index which is able to express PA on a continuous scale as well as to categorize participants was developed. In populations with increasing rates of non-working people the performance of the IPAI is better than the established indices used in EPIC.
The PA data calibration provides more precise risk estimates. The preventive effects of PA on overall chronic disease risk, type 2 diabetes, myocardial infarction and stroke, has been shown to be underestimated. Public health interventions should persistently focus on PA, especially in type 2 diabetes jeopardized individuals
Protein kinase regulation of sarcoplasmic reticulum function in isolated adult rat ventricular myocytes
The sarcoplasmic reticulum (SR) is one of the major regulators of the
cytosolic Ca2+ concentration in cardiac ventricular muscle cells. In the
myocardium, relaxation results from a decrease in cytoplasmic free Ca2+ levels
mediated through an efflux of Ca2+ from the cell via the sarcolemmal Na+/Ca2+-
exchanger and by the sequestration of 0a2+ by the network SR membranes through
the actions of a calcium pump (a Mg2+dependent, Ca2+/K+activated adenosine
triphosphatase; Ca2+/K+-ATPase). During an action potential, the Ca2+ stored in
the SR is released to the cytoplasm via a Ca2+release channel present in the
junctional SR and Ca2+ also enters the cell through voltage-controlled Ca2+
channels in the sarcolemmal membrane. These two processes result in an increase
in cytoplasmic Ca2+ concentration which leads to contraction of the myocardium.
SR membrane function is regulated in part by the phosphorylation of
proteins present in this membrane. Subsequent to 3-adrenergic stimulation of the
heart by catecholamines, levels of cAMP are increased leading to the activation of
cAMP-dependent protein kinase (PK A). In isolated cardiac SR membrane vesicles
and perfused hearts, phosphorylation of an indigenous SR protein, phospholamban,
is mediated by PK A. Phosphorylated phospholamban acts as a modulator of the
Ca2+/K+-ATPase to stimulate active Ca2+ uptake by increasing the affinity of this
enzyme for Ca2+. This stimulation of Ca2+-uptake is the main mechanism by
which catecholamines accelerate relaxation in the heart. When phospholamban is
in the dephosphorylated state, a cytoplasmic portion of the molecule interacts near
the phosphorylation site of the Ca2+-pump to inhibit Ca2+-transport.
[More abstract follows]Pharmaceutical Sciences, Faculty ofGraduat
Entwicklung einer AdministratoroberflĂ€che fĂŒr Lehrveranstaltungen mit Photovoltaik-Anlagen in der virtuellen RealitĂ€t (Kurztitel: Leitstand fĂŒr die Lehre in VR)
Diese Arbeit beschĂ€ftigt sich mit der Erstellung einer AdministratoroberflĂ€che fĂŒr die Lehre bei Photovoltaik (PV)-Praktika in der virtuellen RealitĂ€t (VR). Die erstellte Umgebung bietet, mittels Bildschirmspiegelungen, Möglichkeiten zur didaktischen Anleitung und UnterstĂŒtzung der Studierenden. Das Thema wurde aufgrund einer bestehenden Lehranwendung in der VR bedeutungsvoll und zeigt deutliches Potenzial. Diese Lehranwendung wird bereits umfassend und verpflichtend in den Praktika eingesetzt. Sie bietet einen praxisnahen Aufbau von Solaranlagen und erhöht gefahrlos die Experimentierfreudigkeit. Mit ihr lassen sich die aufgebauten Anlagen technisch prĂŒfen, simulieren und bewerten. Zudem werden die beiden Möglichkeiten zur UnterstĂŒtzung der Studierenden beurteilt. Als Ergebnis wird die Umsetzung der nahezu automatisierten Administratorober-flĂ€che verdeutlicht und ein Usability-Test aus den Praktika evaluiert.
Schlagwörter: AdministratoroberflÀche, Bildschirmspiegelung, C, Didaktik, im-mersiv, Oculus Quest 2, Photovoltaik, Python, Tkinter, virtuelle RealitÀtThis work focuses on the creation of an administrator interface for teaching pho-tovoltaic (PV) practicals in the Virtual Reality (VR). The created environment pro-vides the possibilities for didactic guidance and support of the students by means of screen mirroring. The topic became significant due to an existing teaching ap-plication in VR and shows clear potential. The teaching application is already ex-tensively and mandatory used in the internships. It provides a hands-on construc-tion of solar systems and safely increases the willingness to experiment. It can be used to test, simulate and evaluate the constructed systems. In addition, the two existing possibilities for supporting the students are evaluated up on this ba-sis. As a result, the implementation of the almost automated administrator inter-face is clarified and a usability test from the practical courses is evaluated.
Keywords: administrator interface, C, didactic, immersive, Oculus Quest 2, photovoltaics, Python, screen mirroring, Tkinter, virtual realit
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