Three Sides of the Same Coin: Combining Microbial,Enzymatic, and Organometallic Catalysis for Integrated Conversion of Renewable Carbon Sources

All catalysts have unique abilities. This is especially true for microbial, enzymatic, and organometallic catalysis, which are often seen as competitive approaches preventing the exploitation of their complementarity. An increasing number of examples show, how using the complete catalytic spectrum can open roads from new substrates to new products. C1-compounds such as formate, formaldehyde, methanol, or methane from CO₂ in combination with green H₂ are likely to be future sources of carbon feedstock. This short review highlights how combinations of different catalyst types can facilitate integrated reaction sequences with biogenic substrates to form “bio-hybrid” fuels and products.     

Harmonization of OF LC-MS oxylipin analysis to investigate lipid mediator formation and signaling in immune cells

The oxidation products of polyunsaturated fatty acids (PUFA) - oxylipins - are formed via enzymatic conversion by cyclooxygenases, lipoxygenases (LOX) and cytochrome P450 monooxygenases or by autoxidation. Together with multiple PUFA substrates this leads to a large variety of oxylipins, whose physiological functions are - besides those of well-studied prostaglandins and leukotrienes - little understood. The interplay of several PUFA-oxidizing enzymes, as in the case of multihydroxylated oxylipins, additionally complicates the investigation of their biosynthesis. Due to regulatory crosstalk and complex interactions of different oxylipin classes during physiological processes, such as inflammation, a study of the whole oxylipin profile rather than individual compounds is necessary. Therefore, in order to understand the functions of these lipid mediators potent and reliable biochemical methods and (molecular) biological models are required, in combination with sensitive and sound preanalytical, analytical and post-analytical procedures for their investigation. The first part of this thesis investigated the relevance of harmonizing such analytical methods for the quantitative determination of oxylipins in biological samples. Based on the outcome of an international laboratory comparison, comparable and reproducible results were obtained across independent laboratories when these harmonized methods were used for sample preparation and analysis by means of liquid chromatography coupled with tandem mass spectrometry. The examination of the effect of different storage times and temperatures as a parameter of pre-analytics demonstrated that oxylipin levels in plasma samples are robust to long-term storage and, furthermore, to minor variations during plasma generation. As accurate quantification of oxylipin concentrations rises and falls with the quality of analytical standards, a strategy for the verification of oxylipin standard concentrations with mass- spectrometry and UV spectroscopy based approaches using certified standard compounds was presented. In the second part of the thesis the established methods were applied to study the biosynthesis of oxylipins in immune cells with a special attention to the involved LOX. Stimulation of primary human M2-like macrophages with the liver X receptor (LXR) agonist T09 led to a massive induction of ALOX15 gene expression and thus, 15-LOX abundance and activity. Cholesterol derivatives were identified as potent endogenous ligands of LXR yielding increased 15-LOX abundance and activity. The examination of 5-LOX induced synthesis of multihydroxylated metabolites through supplementation of neutrophils with 15-HETE, 18- HEPE and 17-HDHA primary leads to formation of double oxygenated oxylipins. In intact cells 5-LOX preferred DHA-derived 17-HDHA as substrate, whereas upon cell integrity destruction the formation of 15-HETE derived multihydroxylated oxylipins increased. With the combination of harmonized protocols for sampling and analysis this thesis sets the basis for the reliable quantification of oxylipins. The use of the optimized methodologies allowed to further characterize regulatory pathways of the ARA cascade in human immune cells, contributing to a more thorough understanding of inflammation regulation.     

Competitive Registration Fields for The Development of Complex Block Copolymer Structures by A Layer-by-Layer Approach

Block copolymer (BCP) self-assembly in thin films is an elegant method to generate nanometric features with tunable geometrical configurations. By combining directed assembly and hybridization methods, advances in nano-manufacturing have been attested over the past decades with flagship applications in lithography and optics. Nevertheless, the range of geometrical configurations is limited by the accessible morphologies inherent to the energy minimization process involved in BCP self-assembly. Layering of nanostructured BCP thin films has been recently proposed in order to enrich the span of nanostructures derived from BCP self-assembly with the formation of non-native heterostructures such as double-layered arrays of nanowires or dots-on-line and dots-in-hole hierarchical structures. In this work, the layer-by-layer method is further exploited for the generation of nano-mesh arrays using nanostructured BCP thin films. In particular, a subtle combination of chemical and topographical fields is leveraged in order to demonstrate design rules for the controlled registration of a BCP layer on top of an underneath immobilized one by the precise tuning of the interfacial chemical field between the two BCP layers.     

Design and evaluation of a collaborative UML modeling environment in virtual reality

Software and Systems Modeling - Modeling is a key activity in conceptual design and system design. Through collaborative modeling, end-users, stakeholders, experts, and entrepreneurs are able to...     

Visible-Light Removable Photocaging Groups Accepted by MjMAT Variant: Structural Basis and Compatibility with DNA and RNA Methyltransferases

Methylation and demethylation of DNA, RNA and proteins constitutes a major regulatory mechanism in epigenetic processes. Investigations would benefit from the ability to install photo-cleavable groups at methyltransferase target sites that block interactions with reader proteins until removed by non-damaging light in the visible spectrum. Engineered methionine adenosyltransferases (MATs) have been exploited in cascade reactions with methyltransferases (MTases) to modify biomolecules with non-natural groups, including first evidence for accepting photo-cleavable groups. We show that an engineered MAT from Methanocaldococcus jannaschii (PC-MjMAT) is 308-fold more efficient at converting ortho-nitrobenzyl-(ONB)-homocysteine than the wildtype enzyme. PC-MjMAT is active over a broad range of temperatures and compatible with MTases from mesophilic organisms. We solved the crystal structures of wildtype and PC-MjMAT in complex with AdoONB and a red-shifted derivative thereof. These structures reveal that aromatic stacking interactions within the ligands are key to accommodating the photocaging groups in PC-MjMAT. The enlargement of the binding pocket eliminates steric clashes to enable AdoMet analogue binding. Importantly, PC-MjMAT exhibits remarkable activity on methionine analogues with red-shifted ONB-derivatives enabling photo-deprotection of modified DNA by visible light.     

SBIR-BYOL: a self-supervised sketch-based image retrieval model

Neural Computing and Applications - Sketch-based image retrieval is demanding interest in the computer vision community due to its relevance in the visual perception system and its potential...     

Development of analytical methods fort he quantitative analysis of the enzyme abundance and activity in the arachidonic acid cascade

The arachidonic acid (ARA) cascade is a key regulatory pathway where eicosanoids and other oxylipins are formed via the oxidation of polyunsaturated fatty acids. These oxylipins are involved in controlling physiological functions such as vascular tone, blood clotting and immune regulation. Changes in the oxylipin pattern can be caused by direct enzyme inhibition or modulated gene expression. In order to understand the effect mechanisms modulating the ARA cascade it is essential to not only investigate the metabolite, i.e. oxylipin, concentrations but also the abundances of the enzymes involved in their formation. For this reason, a quantitative targeted proteomics method for the ARA cascade was developed within this thesis. A detailed workflow for method development was established where enzyme/protein levels are measured via sequence-specific, representative peptides chosen on the basis of several criteria. With liquid chromatography-tandem mass spectrometry-based analysis, the cyclooxygenase (COX) pathway of the ARA cascade was investigated in human colon carcinoma cell lines and primary macrophages where strong correlations were found between the oxylipins formed via the COX pathway and the COX-2 abundance. The modulating effects of polyphenols on the 5-lipoxygenase (LOX) pathway of the ARA cascade were investigated in cell-free enzyme assays and human neutrophils. Resveratrol, its dimer ɛ-viniferin and a resveratrol imine analogue directly inhibited the 5-LOX with micromolar potencies and modulated the total oxylipin pattern in the neutrophils with remarkably different effects. Genistein did not act as a 5-LOX inhibitor in the cell-free assay but the modulated oxylipin pattern in the cells suggests a global cellular interference. The enzymes of the LOX pathway were included in the targeted proteomics method which together with an extended targeted oxylipin metabolomics platform enables a sensitive multi- omics analysis of the ARA cascade in a single sample. The comparison of the multiple reaction monitoring (MRM) detection and the MRM^-based approach with an additional fragmentation step demonstrated that MRM is more favorable in targeted proteomics due to its higher sensitivity, greater linear range and higher multiplexing capacities. The analysis with the multi- omics approach showed that the 5 LOX protein is induced together with its product formation during the differentiation of human monocytic THP 1 cells to macrophages. M1-like human primary macrophages were also characterized by the abundance of 5-LOX and its activating protein (FLAP), while 15-LOX and 15-LOX-2 dominated the protein pattern of the M2 like macrophages accompanied by high levels of the oxylipins formed via these enzymes. The developed methodology allows mechanistic investigation of a modulation of the ARA cascade as demonstrated by lipo-poly-saccharide stimulation as well as pharmaceutical treatment of the cells. Overall, with this thesis, a new quantitative proteomics strategy for the enzymes of the ARA cascade is established which can be easily extended to further proteins. The use of this technique in combination with targeted oxylipin metabolomics has provided new insights into the mechanisms involved in the modulation of the ARA cascade in human immune cells.     

An evaluation of approaches to model checking real-time task schedulability analysis

International Journal on Software Tools for Technology Transfer - This article is a follow-up contribution that extends the conference paper (Nxumalo, in: Laarman, Sokolova (eds)Model Checking...     

Adsorption auf der ACHEMA 2022

After a longer break due to the pandemic, ACHEMA was held again in Frankfurt in 2022, where, among others, several companies from the adsorption sector presented themselves and their products. The broad exhibitor field ranged from adsorbents to measuring devices and plant engineering. The report covers trends and innovations in adsorption technology, which were presented at the booths and discussed in talks with the exhibitors.