Controlling Enzymatic Activity by Modulating the Oligomerization State via Chemical Rescue and Optical Control

Selective switching of enzymatic activity has been a longstanding goal in synthetic biology. Drastic changes in activity upon mutational manipulation of the oligomerization state of enzymes have frequently been reported in the literature, but scarcely exploited for switching. Using geranylgeranylglyceryl phosphate synthase as a model, we demonstrate that catalytic activity can be efficiently controlled by exogenous modulation of the association state. We introduced a lysine-to-cysteine mutation, leading to the breakdown of the active hexamer into dimers with impaired catalytic efficiency. Addition of bromoethylamine chemically rescued the enzyme by restoring hexamerization and activity. As an alternative method, we incorporated the photosensitive unnatural amino acid o-nitrobenzyl-O-tyrosine (ONBY) into the hexamerization interface. This again led to inactive dimers, but the hexameric state and activity could be recovered by UV-light induced cleavage of ONBY. For both approaches, we obtained switching factors greater than 350-fold, which compares favorably with previously reported activity changes that were caused by site-directed mutagenesis.     

Development of a PM 10/PM 2.5 Cascade Impactor and In-Stack Measurements

Combustion and industrial processes are an important source of particles. Due to the new PM 10 and PM 2.5 standards for ambient air quality, a sampling system for PM 10/PM 2.5 in-stack measurements was designed and calibrated. In this new system, the exhaust gas is isokinetically sucked into a two stage impactor through the inlet of a plane filter device and the aerosol is fractionated in the particle size classes >10 w m, 10-2.5 w m, and <2.5 w m. Due to a relatively high volume flow (ca. 3.2 m 3 /h, depending on exhaust gas conditions), sampling times are kept short, e.g., 30 min for dust concentrations of 10 mg/m 3 . The impactor was calibrated in the laboratory and then operated at various industrial plants. Parallel measurements with identical devices showed average standard deviations of 3.1% (PM 10) and 3.4% (PM 2.5). Measurements of the cascade impactor together with the plane filter device gave plausible results and average PMx/TSP ratios of 0.49 (PM 2.5/TSP) and 0.78 (PM 10/TSP), showing a large variability for different processes. Elemental analysis using total-reflection X-ray fluorescence spectrometry, together with the size-fractionated sampling, proved to give characteristic patterns of the emitted aerosols, which can be used for a subsequent fingerprint modelling for source apportionment of ambient air pollution.     

Nanogel-modified polycaprolactone microfibres with controlled water uptake and degradability

In this work we prepared composite poly(caprolactone) (PCL) microfibres decorated with temperature-sensitive poly(N-vinylcaprolactam) nanogels by an one-step electropsinning process. Microfibres with variable internal structure were prepared by using two different solvent systems: methanol/toluene (Me/Tol) and chloroform/dimethylformamide (Ch/DMF). Our experimental data shows that the nature of the solvent mixtures allows obtaining microfibres with different morphologies: Microfibres with nanogels on the fibre surface (Me/Tol) and microfibres with nanogels in the fibre interior (Ch/DMF). The morphology of composite fibres was visualized by scanning electron microscopy (SEM) and their properties investigated by differential scanning calorimetry (DSC), thermo-gravimetric analysis (TGA) and contact angle measurements. The results show that combining hydrophobic poly(caprolactone) with hydrophilic nanogels leads to microfibres exhibiting controlled swelling in water. Additionally, the thermo-sensitive properties of the nanogels are maintained whether they are on the surface or inside of the fibres. The presence of nanogels in the fibre structure also allows regulating their degradability.     

A review of occupant control on natural ventilation

According to IPCC (Intergovernmental Panel on Climate Change), the largest use of energy in commercial buildings is space heating in colder climates and air conditioning in hot climates. In Europe, the Directive on the energy performance of buildings EPBD (European Energy Performance of Buildings Directive) [1] provides a framework for national building performance regulations and calculation procedures. However, there are often large discrepancies between calculated and measured energy performance of buildings. One main reason is the behaviour of occupants, which is often not reflected in calculation models.This paper presents a literature review of various parameters influencing the effectiveness of occupant controlled natural ventilation. Additionally possibilities to implement these influences into calculation procedures/building simulation and adaptive thermal comfort evaluation are discussed.     

Hydrogen production from carrot pulp by the extreme thermophiles Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana

Hydrogen was produced from carrot pulp hydrolysate, untreated carrot pulp and (mixtures of) glucose and fructose by the extreme thermophiles Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana in pH-controlled bioreactors. Carrot pulp hydrolysate was obtained after enzymatic hydrolysis of the polysaccharide fraction in carrot pulp. The main sugars in the hydrolysate were glucose, fructose, and sucrose.     

Computer-Assisted Selective Optimization of Side-Activities—from Cinalukast to a PPARα Modulator

Automated computational analogue design and scoring can speed up hit-to-lead optimization and appears particularly promising in selective optimization of side-activities (SOSA) where possible analogue diversity is confined. Probing this concept, we employed the cysteinyl leukotriene receptor 1 (CysLT₁R) antagonist cinalukast as lead for which we discovered peroxisome proliferator-activated receptor α (PPARα) modulatory activity. We automatically generated a virtual library of close analogues and classified these roughly 8000 compounds for PPARα agonism and CysLT₁R antagonism using automated affinity scoring and machine learning. A computationally preferred analogue for SOSA was synthesized, and in vitro characterization indeed revealed a marked activity shift toward enhanced PPARα activation and diminished CysLT₁R antagonism. Thereby, this prospective application study highlights the potential of automating SOSA.     

Biaxial strength and slow crack growth in porous alumina with silica sintering aid

Biaxial strength, fracture toughness and subcritical crack growth are reported for coarse grained porous alumina ceramics. The materials were prepared with a varying amount of a silica sintering aid, which resulted in the formation of a glassy secondary phase at the grain boundaries. Crystalline mullite was additionally found in the material with the highest silica content. The biaxial strength, measured by Ball-on-Ring and Ball-on-3-Balls, was highest for the material without mullite at the grain boundaries, and the biaxial strength decreased with increasing porosity. The fracture toughness of the materials was in the range of 1.7–1.9 MPa m^0.5. Measurements of subcritical crack growth by a modified lifetime method in air and aqueous environments demonstrated a higher crack growth rate in water and acid relative to in air. The effect of porosity and grain boundary phase were discussed in relation to subcritical crack growth and fracture mode in the coarse grained alumina ceramics.     

Effects of Particle Size and Molecular Weight of Polyethylenimine on Properties of Nanoparticulate Silicon Dispersions

The effect of particle size on the zeta potential and rheology of nanoparticulate SiO₂ dispersions stabilized with the polymer polyethylenimine (PEI) was investigated experimentally. The particle size and molecular weight of the polymer had only a small influence on the amount of PEI needed and the achieved zeta potential of the suspensions. The polymer concentration range within which a stable dispersion could be produced was very narrow for the smallest particle size. Higher-molecular-weight PEI was more suitable to stabilize small particle dispersions. Under optimum conditions, Newtonian flow behavior was achieved for dispersions of 20 nm particles with a solids content as high as 25 vol%.     

Hope for GWAS: Relevant Risk Genes Uncovered from GWAS Statistical Noise

Hundreds of genetic variants have been associated to common diseases through genome-wide association studies (GWAS), yet there are limits to current approaches in detecting true small effect risk variants against a background of false positive findings. Here we addressed the missing heritability problem, aiming to test whether there are indeed risk variants within GWAS statistical noise and to develop a systematic strategy to retrieve these hidden variants. Employing an integrative approach, which combines protein-protein interactions with association data from GWAS for 6 common diseases, we found that associated-genes at less stringent significance levels (p < 0.1) with any of these diseases are functionally connected beyond noise expectation. This functional coherence was used to identify disease-relevant subnetworks, which were shown to be enriched in known genes, outperforming the selection of top GWAS genes. As a proof of principle, we applied this approach to breast cancer, supporting well-known breast cancer genes, while pinpointing novel susceptibility genes for experimental validation. This study reinforces the idea that GWAS are under-analyzed and that missing heritability is rather hidden. It extends the use of protein networks to reveal this missing heritability, thus leveraging the large investment in GWAS that produced so far little tangible gain.