METHODS DEVELOPMENT IN BIOLOGICAL MASS SPECTROMETRY: APPLICATION IN GLYCOPROTEOMICS

Proteomics refers to global characterization of the full set of proteins present in a biological sample. Various analytical disciplines contribute to proteomics but mass spectrometry became method of choice for analysis of complex protein samples. Mass spectrometry allows for high throughput analysis of the proteome but, moreover, it has the ability to acquire higher-order information such as post-translational modifications (PTM). Glycosylation is the most abundant PTM on eukaryotic proteins. This dissertation will focus on method development for structural proteomics that will be utilized to explain the glycoproteome of obligate intracellular protozoan parasite Toxoplasma gondii as a model system. Optimization of sample preparation is addressed in the first part of this dissertation. Sample preparation for mass spectrometry analysis is a critical step in the proteomics workflow because the quality and reproducibility of sample extraction and preparation significantly impacts the separation and identification capabilities of mass spectrometers. Also, there are problems unique to intracellular parasites as limited amount, host cell impurity and choice of the host. The additional obstacle is to extract only glycosylated proteins for which there is no one standard method. Here we report the optimal sample preparation method utilizing agarose bound Concanavalin A (Con A) beads to efficiently pull down glycoproteins, dialyze and analyze them using MuDPIT. This method was further enhanced by passing the non-retained protein fraction (first flow-through) through a second Con A column and then passing the second non-retained protein fraction (second flow-through) through the third Con A column (3 sequential pull-downs) yielding 394 benchmark proteins. Glycoproteome of Toxoplasma gondii is not yet fully understood. However, evidence suggests that glycosylation could be essential for cyst formation and maintenance which is characteristic of chronic stage of disease. The focus of the second part of dissertation is to better understand the differences in glycoproteomes of tachizoites and tissue cysts. Cyst proteins pulled down using optimized sample preparation method that do not appear in the tachyzoites pulldowns could be critical elements in the structural stability of the tissue cyst

Digital Cultural Heritage – an Overview of the Field

Der Sammelband Digital Cultural Heritage besteht aus dreißig Beiträgen, die mit einem breiten geographischen und zeitlichen Rahmen einen umfassenden Überblick über die aktuellen Entwicklungen in diesem Forschungsfeld anbieten. Einige der Themen, denen im Band besondere Aufmerksamkeit gewidmet wurden, sind die Fragen der Konservierung und Präsentation von (digitalem) Kulturerbe, sowie Möglichkeiten zu dessen Anreicherung mit Hilfe digitaler Werkzeuge. Obwohl der Band reich an Fallbeispielen ist, fehlt es ihm jedoch an Leserführung und einem umfassenden konzeptuellen Ansatz.The edited volume Digital Cultural Heritage is a collection of thirty contributions dealing with cultural heritage in the digital context. Covering a broad geographical and chronological scope, it presents the reader with an extensive overview of the current work in the field. Some of the topics that were given special attention in the volume are questions of preservation and presentation of (digital) cultural heritage, as well as possibilities for its enrichment with the help of digital tools. While rich in examples, the volume, however, lacks reader guidance and a rounded approach to the topic

On the robustness of ultra-high voltage 4H-SiC IGBTs with an optimized retrograde p-well

The robustness of ultra-high voltage (>10kV) SiC IGBTs comprising of an optimized retrograde p-well is investigated. Under extensive TCAD simulations, we show that in addition to offering a robust control on threshold voltage and eliminating punch-through, the retrograde is highly effective in terms of reducing the stress on the gate oxide of ultra-high voltage SiC IGBTs. We show that a 10 kV SiC IGBT comprising of the retrograde p-well exhibits a much-reduced peak electric field in the gate oxide when compared with the counterpart comprising of a conventional p-well. Using an optimized retrograde p-well with depth as shallow as 1 μm, the peak electric field in the gate oxide of a 10kV rated SiC IGBT can be reduced to below 2 MV.cm -1 , a prerequisite to achieve a high-degree of reliability in high-voltage power devices. We therefore propose that the retrograde p-well is highly promising for the development of>10kV SiC IGBTs

Optimal edge termination for high oxide reliability aiming 10kV SiC n-IGBTs

The edge termination design strongly affects the ability of a power device to support the desired voltage and its reliable operation. In this paper we present three appropriate termination designs for 10kV n-IGBTs which achieve the desired blocking requirement without the need for deep and expensive implantations. Thus, they improve the ability to fabricate, minimise the cost and reduce the lattice damage due to the high implantation energy. The edge terminations presented are optimised both for achieving the widest immunity to dopant activation and to minimise the electric field at the oxide. Thus, they ensure the long-term reliability of the device. This work has shown that the optimum design for blocking voltage and widest dose window does not necessarily give the best design for reliability. Further, it has been shown that Hybrid Junction Termination Extension structure with Space Modulated Floating Field Rings can give the best result of very high termination efficiency, as high as 99%, the widest doping variation immunity and the lowest electric field in the oxide

Production of Hydroxy Acids:Selective Double Oxidation of Diols by Flavoprotein Alcohol Oxidase

Flavoprotein oxidases can catalyze oxidations of alcohols and amines by merely using molecular oxygen as the oxidant, making this class of enzymes appealing for biocatalysis. The FAD-containing (FAD=flavin adenine dinucleotide) alcohol oxidase from P. chrysosporium facilitated double and triple oxidations for a range of aliphatic diols. Interestingly, depending on the diol substrate, these reactions result in formation of either lactones or hydroxy acids. For example, diethylene glycol could be selectively and fully converted into 2-(2-hydroxyethoxy)acetic acid. Such a facile cofactor-independent biocatalytic route towards hydroxy acids opens up new avenues for the preparation of polyester building blocks

Estimating and validating the interbeat intervals of the heart using near-infrared spectroscopy on the human forehead

In studies with near-infrared spectroscopy, the recorded signals contain information on the temporal interbeat intervals of the heart. If this cardiac information is needed exclusively and could directly be extracted, an additional electrocardiography device would be unnecessary. The aim was to estimate these intervals from signals measured with near-infrared spectroscopy with two novel approaches. In one approach, we model the heartbeat oscillations in these signals with a Fourier series where the coefficients and the fundamental frequency can continuously change over time. The time-dependent model parameters are estimated and used to calculate the interbeat intervals. The second approach uses empirical mode decomposition. The signal measured with near-infrared spectroscopy is empirically decomposed into a set of oscillatory components. The sum of a specific subset of them is an estimate of the pure heartbeat signal in which the diastolic peaks and consequential interbeat intervals are detected. We show in simultaneous electrocardiography and near-infrared spectroscopy measurements on 11 subjects (8 men and 3 woman with mean age 32.8 ± 8.1 yr), that the interbeat intervals (and the consequential pulse rate variability measures), estimated using the proposed approaches, are in high agreement with their correspondents from electrocardiography

Means and methods for selective double diol oxidation.

The invention relates to the field of enzyme engineering and biocatalysis, in particular to alcohol oxidases and their application in the selective oxidation of diols for the production of industrially useful oxidation products thereof, such as precursors for biodegradable polyesters. Provided is a method for the selective oxidation of a diol substrate, comprising subjecting a diol substrate of the formula HO-CH2-CH2-X-CH2-CH2-OH, wherein X = O, S or CH2, to a FAD-containing alcohol oxidase (AOX) enzyme of the class EC 1.1.3.13 or EC 1.1.3.17. or a mutant thereof, under conditions allowing lor the double oxidation of one hydroxyl moiety of the diol substrate into an oxidation product