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Intracellular protein motors have evolved to perform specific tasks critical to the function of cells such as intracellular trafficking and cell division. Kinesin and dynein motors, for example, transport cargoes in living cells by walking along microtubules powered by adenosine triphosphate hydrolysis. These motors can make discrete 8 nm centre-of-mass steps and can travel over 1 µm by changing their conformations during the course of adenosine triphosphate binding, hydrolysis and product release. Inspired by such biological machines, synthetic analogues have been developed including self-assembled DNA walkers that can make stepwise movements on RNA/DNA substrates or can function as programmable assembly lines. Here, we show that motors based on RNA-cleaving DNA enzymes can transport nanoparticle cargoes-CdS nanocrystals in this case-along single-walled carbon nanotubes. Our motors extract chemical energy from RNA molecules decorated on the nanotubes and use that energy to fuel autonomous, processive walking through a series of conformational changes along the one-dimensional track. The walking is controllable and adapts to changes in the local environment, which allows us to remotely direct 'go' and 'stop' actions. The translocation of individual motors can be visualized in real time using the visible fluorescence of the cargo nanoparticle and the near-infared emission of the carbon-nanotube track. We observed unidirectional movements of the molecular motors over 3 µm with a translocation velocity on the order of 1 nm min−1 under our experimental conditions. [ABSTRACT FROM AUTHOR]

Highlights: [•] A high-throughput screening system was established for exocellulase screening. [•] CelEx-BR12 was first isolated from metagenomic resources for exocellulase activity. [•] CelEx-BR12 is a multifunctional enzyme having endo/exocellulase, xylanase activities. [Copyright &y& Elsevier]

The dynamic reconfiguration of a hierarchically assembled tubular structure is demonstrated using the DNA origami technique. Short cylindrical DNA origami monomers are synthesized and linked into elongated tubules, which can then be disassembled via toehold-mediated strand displacement. The disassembled subunits are subsequently linked into tubules of a different chirality. The reconfiguration is performed with the subunits carrying dumbbell hairpin DNA oligonucleotides or gold nanoparticles (AuNPs). The reconfiguration of higher order origami structures presented here is useful for constructing dynamic nanostructures that exceed the size limit of single DNA origami and may facilitate the study of molecular or particle interactions by tuning their relative distance and organization. [ABSTRACT FROM AUTHOR]

Abstract: Exocellulases play a key role in cleaving the accessible ends of cellulose molecules to release soluble glucose and cellobiose. To date, there have been no screens for exocellulase owing to assay protocol limitations, the high cost of substrates, and low activity of exocellulases compared with endocellulases. This study is the first to demonstrate direct screening for exocellulase activity using a robotic, high-throughput screening (HTS) system. Cell growth in 96-well plates was measured by monitoring optical density over 11–14h at 37°C with agitation. Fluorescence of methylumbelliferyl groups released from 4-methylumbelliferyl-β-D-cellobioside was determined using a VICTOR3 microplate reader. This new HTS system enabled activity verification of more than 104 clones per day. As a result, we obtained four exocellulases clones (CelEx-SF301, CelEx-SF309, CelEx-BR12 and CelEx-BR15) from 29,006 metagenomic fosmid clones that had previously been prepared from sweet potato field soil microbes and rumen fluid. This powerful approach could be effectively applied to screen various metagenomic resources for new enzymes. [Copyright &y& Elsevier]

Abstract: In previous studies, we identified sedative effects of Scutellaria baicalensis extracts and found that these extracts or their constituents may also have anticonvulsive effects. Wogonin is a natural product isolated from S. baicalensis, which possesses central nervous system effects such as anxiolytic and neuroprotective activities. In this study, we investigated the effects of wogonin on convulsion related behaviors, such as myorelaxation, motor coordination, and anticonvulsant effects of wogonin on chemical induced seizure and electroshock seizure in mice or rats. The effect of wogonin on membrane potential was also observed. Wogonin was intraperitoneally injected into mice or rats 30 min prior to testing. Animals treated with wogonin did not change locomotor activities as well as endurance times on the rota-rod, which indicates that wogonin did not cause a sedative and myorelaxation effect. Wogonin significantly blocked convulsion induced by pentylenetetrazole and electroshock but not convulsion induced by strychnine. Wogonin also significantly reduced the electrogenic response score, but flumazenil treatment reversed this decrease to the level of the control group. The wogonin treatment increased Cl-influx into the intracellular area as dose increased. Flumazenil and bicuculline treatment, however, inhibited the Cl− influx induced by wogonin. These results indicate that the anticonvulsive effects produced by wogonin were mediated by the GABAergic neuron. [Copyright &y& Elsevier]

Based on the previous study for the water-air mixture, the characteristics of AC electromagnetic flowmeter were investigated in the liquid metal two-phase flow. Two-phase flow experiments, encompassing bubbly to slug flow regimes, were conducted with the liquid sodium–nitrogen mixture. The simple relation ΔUTP=ΔUSP(1−α), relating flowmeter signal between single-phase flow and two-phase flow, was verified with measurements of the potential differences and the void fraction α for a bubbly flow regime in water-air loop. Whereas there was no difference in the shape of the raw signal between single-phase flow and bubbly flow, the signal amplitude of bubbly flow was greater than that of single-phase flow under the same liquid flow (sodium or water) rates, since the passage area of the liquid flow was reduced. In the case of slug flow, the phase and the amplitude of the flowmeter output showed a change in the flow characteristics around each slug bubble and the position of a slug bubble itself. Therefore, the electromagnetic flowmeter shows a good potential as a useful device for identifying the flow regimes. The void fraction can be simply measured with two electromagnetic flowmeters for a low-void fraction flow such as bubbly flow. Thus, the flowmeters can be used to measure the void fraction for a liquid sodium. [Copyright &y& Elsevier]

Corynebacterium glutamicum is an industrial strain used for the production of valuable chemicals such as L-lysine and L-glutamate. Although C. glutamicum has various industrial applications, a limited number of tunable systems are available to engineer it for efficient production of platform chemicals. Therefore, in this study, we developed a novel tunable promoter system based on repeats of the Vitreoscilla hemoglobin promoter (Pvgb). Tunable expression of green fluorescent protein (GFP) was investigated under one, four, and eight repeats of Pvgb (Pvgb, Pvgb4, and Pvgb8). The intensity of fluorescence in recombinant C. glutamicum strains increased as the number of Pvgb increased from single to eight (Pvgb8) repeats. Furthermore, we demonstrated the application of the new Pvgb promoter-based vector system as a platform for metabolic engineering of C. glutamicum by investigating 5-aminovaleric acid (5-AVA) and gamma-aminobutyric acid (GABA) production in several C. glutamicum strains. The profile of 5-AVA and GABA production by the recombinant strains were evaluated to investigate the tunable expression of key enzymes such as DavBA and GadBmut. We observed that 5-AVA and GABA production by the recombinant strains increased as the number of Pvgb used for the expression of key proteins increased. The recombinant C. glutamicum strain expressing DavBA could produce higher amounts of 5-AVA under the control of Pvgb8 (3.69 ± 0.07 g/L) than the one under the control of Pvgb (3.43 ± 0.10 g/L). The average gamma-aminobutyric acid production also increased in all the tested strains as the number of Pvgb used for GadBmut expression increased from single (4.81–5.31 g/L) to eight repeats (4.94–5.58 g/L). [ABSTRACT FROM AUTHOR]

Background: Recent interest has been focused on the production of platform chemicals from renewable biomass due to increasing concerns on global warming and depletion of fossil fuel reserves. Microbial production of platform chemicals in biorefineries has been suggested to be a promising solution for these problems. Gamma-aminobutyrate (GABA), a versatile bulk chemical used in food and pharmaceutical industry, is also used as a key monomer for nylon 4. GABA can be biologically produced by decarboxylation of glutamate. Results: In this study, we examined high glutamate-producing Corynebacterium glutamicum strains as hosts for enhanced production of GABA from glucose and xylose as carbon sources. An Escherichia coli gadB mutant with a broad pH range of activity and E. coli xylAB genes were expressed under the control of a synthetic H36 promoter. When empty fruit bunch (EFB) solution was used as carbon source (45 g/L glucose and 5 g/L xylose), 12.54 ± 0.07 g/L GABA was produced by recombinant C. glutamicum H36GD1852 expressing E. coli gadB mutant gene and xylAB genes. Batch fermentation of the same strain resulted in the production of 35.47 g/L of GABA when EFB solution was added to support 90 g/L glucose and 10 g/L xylose. Conclusions: This is the first report of GABA production by recombinant C. glutamicum strains from co-utilization of glucose and xylose from EFB solution. Recombinant C. glutamicum strains developed in this study should be useful for an efficient and sustainable production of GABA from lignocellulosic biomasses. [ABSTRACT FROM AUTHOR]

Abstract: We have developed a system to detect three hydrolytic enzymes (cellulase, lipase, and protease) using a single sodium dodecyl sulfate (SDS) gel and an electrotransfer system. After electrophoresis, proteins in the gel were transferred to three sandwiched substrate gels containing glycerol tributyrate, azo-carboxymethyl cellulose (Azo-CMC), and fibrin for detection of cellulase, lipase, and protease, respectively. We show that three cellulases (from a Paenibacillus sp. and two Bacillus sp. strains), one lipase (from a Staphylococcus sp.), and two proteases (from two Bacillus sp. strains) can be detected simultaneously with our zymogram system. [Copyright &y& Elsevier]