Full-Spectrum Targeted Mutagenesis in Plant and Animal Cells

Directed evolution is a powerful approach for protein engineering and functional studies. However, directed evolution outputs from bacterial and yeast systems do not always translate to higher organisms. In situ directed evolution in plant and animal cells has previously been limited by an inability to introduce targeted DNA sequence diversity. New hypermutation tools have emerged that can generate targeted mutations in plant and animal cells, by recruiting mutagenic proteins to defined DNA loci. Progress in this field, such as the development of CRISPR-derived hypermutators, now allows for all DNA nucleotides within user-defined regions to be altered through the recruitment of error-prone DNA polymerases or highly active DNA deaminases. The further engineering of these mutagenesis systems will potentially allow for all transition and transversion substitutions to be generated within user-defined genomic windows. Such targeted full-spectrum mutagenesis tools would provide a powerful platform for evolving antibodies, enzymes, structural proteins and RNAs with specific desired properties in relevant cellular contexts. These tools are expected to benefit many aspects of biological research and, ultimately, clinical applications.     

Automated Flowsheet Synthesis Using Hierarchical Reinforcement Learning: Proof of Concept

Recently we showed that reinforcement learning can be used to automatically generate process flowsheets without heuristics or prior knowledge. For this purpose, SynGameZero, a novel two-player game has been developed. In this work we extend SynGameZero by structuring the agent's actions in several hierarchy levels, which improves the approach in terms of scalability and allows the consideration of more sophisticated flowsheet problems. We successfully demonstrate the usability of our novel framework for the fully automated synthesis of an ethyl tert-butyl ether process.     

Cortactin Is Required for Efficient FAK,Src and Abl Tyrosine Kinase Activation and Phosphorylation of Helicobacter pylori CagA

Cortactin is a well-known regulatory protein of the host actin cytoskeleton and represents an attractive target of microbial pathogens like Helicobacter pylori. H. pylori manipulates cortactin’s phosphorylation status by type-IV secretion-dependent injection of its virulence protein CagA. Multiple host tyrosine kinases, like FAK, Src, and Abl, are activated during infection, but the pathway(s) involved is (are) not yet fully established. Among them, Src and Abl target CagA and stimulate tyrosine phosphorylation of the latter at its EPIYA-motifs. To investigate the role of cortactin in more detail, we generated a CRISPR/Cas9 knockout of cortactin in AGS gastric epithelial cells. Surprisingly, we found that FAK, Src, and Abl kinase activities were dramatically downregulated associated with widely diminished CagA phosphorylation in cortactin knockout cells compared to the parental control. Together, we report here a yet unrecognized cortactin-dependent signaling pathway involving FAK, Src, and Abl activation, and controlling efficient phosphorylation of injected CagA during infection. Thus, the cortactin status could serve as a potential new biomarker of gastric cancer development.     

Evaluation of wind power forecasts—An up‐to‐date view

Wind power forecast evaluation is of key importance for forecast provider selection, forecast quality control, and model development. While forecasts are most often evaluated based on squared or absolute errors, these error measures do not always adequately reflect the loss functions and true expectations of the forecast user, neither do they provide enough information for the desired evaluation task. Over the last decade, research in forecast verification has intensified, and a number of verification frameworks and diagnostic tools have been proposed. However, the corresponding literature is generally very technical and most often dedicated to forecast model developers. This can make forecast users struggle to select the most appropriate verification tools for their application while not fully appraising subtleties related to their application and interpretation. This paper revisits the most common verification tools from a forecast user perspective and discusses their suitability for different application examples as well as evaluation setup design and significance of evaluation results.     

Lipid modification of amino acids,carbohydrates and polyols

Agricultural crops provide a considerable reservoir of useful and low cost raw materials like fats and oils, plant proteins and carbohydrates. By selective combination of their molecular constituents (e. g. fatty acids, glycerol, amino acids, saccharides), a wide variety of surface active materials can be prepared. Due to their molecular constitution, all of them are potentially biodegradable. In this article we describe two generally applicable methods for the synthesis of such combination products using fatty acids from plant oils – one of the most important renewable materials as feedstock for the chemical industry – in combination with amino acids and monosaccharides including sugar alcohols. Using the high selectivities displayed by lipases, which catalyze a wide variety of transformations under very mild conditions (pH, temperature) with high atom efficiency in various non‐toxic solvents and without the production of by‐products or waste, numerous useful products can be obtained ranging all the way from surface active compounds (detergents and emulsifiers for food and personal care products) such as mono‐ and diglycerides, sugar esters, synthetic building blocks, products for food and dietary applications, all the way to novel methods for the production of biofuels (biodiesel). In our continuing effort to devise novel and generally applicable methods for the syntheses of such combination products we recently discovered a novel class of building blocks, fatty acid modified anhydrides of hydroxy carboxylic acids such as citric acid, tartaric acid and malic acid which led to alternative synthetic approaches towards new classes of the title compounds.     

Parallel integration of aligned carbon strings in polymer composite: Dielectrophoretic preparation,finite element simulation,and electrical characterization

We report on dielectrophoretic alignment of carbon black particles into radially arranged string‐like assemblies in oligourethane mixtures followed by photopolymerization. Using finite element modelling and optical microscopy we find significant difference in field distributions depending on the substrate beneath aligned films. On glass, the field is concentrated between the electrode tips leading to string‐like assemblies between the tips. On oxide covered silicon, the field is distributed along the electrode circumference leading to more distributed fractal assemblies. Using comprehensive dc resistance measurements and ac‐impedance spectroscopy we show that the resistance of the strings varies from 120 kΩ to 5 MΩ with a mean of 1.03 MΩ. Strings on oxide covered silicon show significantly higher resistances than the strings on glass. We also demonstrate the effect of aging and an increase in resistance through elongating aligned strings. POLYM. COMPOS., 36:1866–1874, 2015. © 2014 Society of Plastics Engineers     

Brominative Deoxygenation of some aldehydes and ethers

Three aldehydes ( 4a–c ) are transformed into 1,1-dibromides ( 6a–c ) by 2,2,2-tribromo-2,2-dihydro-1,3,2-benzodioxaphosphole ( 2 ). This reagent ( 2 ) is also very active in the cleavage of ethers; its reactions may show some features of carbonium as well as of S_N2 character.     

Mechanical characterization and fractography of glass fiber/polyamide (PA6) composites

The mechanical properties of the glass fiber reinforced Polyamide (PA6) composites made by prepreg tapes and commingled yarns were studied by in‐plane compression, short‐beam shear, and flexural tests. The composites were fabricated with different fiber volume contents (prepregs—47%, 55%, 60%, and commingled—48%, 48%, 49%, respectively) by using vacuum consolidation technique. To evaluate laminate quality in terms of fiber wet‐out at filament level, homogeneity of fiber/matrix distribution, and matrix/fiber bonding standard microscopic methods like optical microscopy and scanning electron microscopy (SEM) were used. Both commingled and prepreg glass fiber/PA6 composites (with V_f ∼ 48%) give mechanical properties such as compression strength (530–570 MPa), inter‐laminar shear strength (70–80 MPa), and transverse strength (80–90 MPa). By increasing small percentage in the fiber content show significant rise in compression strength, slight decrease in the ILSS and transverse strengths, whereas semipreg give very poor properties with the slight increase in fiber content. Overall comparison of mechanical properties indicates commingled glass fiber/PA6 composite shows much better performance compared with prepregs due to uniform distribution of fiber and matrix, better melt‐impregnation while processing, perfect alignment of glass fibers in the composite. This study proves again that the presence of voids and poor interface bonding between matrix/fiber leads to decrease in the mechanical properties. Fractographic characterization of post‐failure surfaces reveals information about the cause and sequence of failure. POLYM. COMPOS., 36:834–853, 2015. © 2014 Society of Plastics Engineers