Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans

β-Glucan is widely distributed in various plants and microorganisms and is composed of β-1,3-linked d-glucose units. It may have a branched short or long side chain of glucose units with β-1,6- or β-1,4-linkage. Numerous studies have investigated different β-glucans and revealed their bioactivities. To understand the structure-function relationship of β-glucan, we constructed a split-luciferase complementation assay for the structural analysis of long-chain β-1,6-branched β-1,3-glucan. The N- and C-terminal fragments of luciferase from deep-sea shrimp were fused to insect-derived β-1,3-glucan recognition protein and fungal endo-β-1,6-glucanase (Neg1)-derived β-1,6-glucan recognition protein, respectively. In this approach, two β-glucan recognition proteins bound to β-glucan molecules come into close proximity, resulting in the assembly of the full-length reporter enzyme and induction of transient luciferase activity, indicative of the structure of β-glucan. To test the applicability of this assay, β-glucan and two β-glucan recognition proteins were mixed, resulting in an increase in the luminescence intensity in a β-1,3-glucan with a long polymer of β-1,6-glucan in a dose-dependent manner. This simple test also allows the monitoring of real-time changes in the side chain structure and serves as a convenient method to distinguish between β-1,3-glucan and long-chain β-1,6-branched β-1,3-glucan in various soluble and insoluble β-glucans.     

Characterisation of the Dynamic Interactions between Complex N-Glycans and Human CD22

CD22 (Siglec-2) is a B-cell surface inhibitory protein capable of selectively recognising sialylated glycans, thus dampening autoimmune responses against self-antigens. Here we have characterised the dynamic recognition of complex-type N-glycans by human CD22 by means of orthogonal approaches including NMR spectroscopy, computational methods and biophysical assays. We provide new molecular insights into the binding mode of sialoglycans in complex with h-CD22, highlighting the role of the sialic acid galactose moieties in the recognition process, elucidating the conformational behaviour of complex-type N-glycans bound to Siglec-2 and dissecting the formation of CD22 homo-oligomers on the B-cell surface. Our results could enable the development of additional therapeutics capable of modulating the activity of h-CD22 in autoimmune diseases and malignancies derived from B-cells.     

Possibility of Japanese Cedar Pollen Causing False Positives in the Deep Mycosis Test

Because Japanese cedar pollen (JCP) contains beta-1,3-d-glucan (BG), there is concern that its lingering presence in the atmosphere, especially during its scattering period, may cause false positives in the factor-G-based Limulus amebocyte lysate (LAL) assay used to test for deep mycosis (i.e., G-test). Hence, we examined whether the LAL assay would react positively with substances contained in JCP by using the G-test to measure JCP particles and extracts. BG was purified from the JCP extract on a BG-specific affinity column, and the percentage extractability was measured using three different BG-specific quantitative methods. The G-test detected 0.4 pg BG in a single JCP particle and 10 fg from a single particle in the extract. The percentage extractability of JCP-derived BG was not significantly different among the three quantitative methods. As the JCP particles should technically have been removed during serum separation, they should be less likely to be a direct false-positive factor. However, given that the LAL-assay-positive substances in the JCP extract were not distinguishable by the three BG-specific quantitative methods, we conclude that they may cause the background to rise. Therefore, in Japan false positives arising from JCP contamination should be considered when testing patients for deep mycosis.     

Multimodal sensing and active continuous closed‐loop feedback for achieving reliable manipulation in the outdoor physical world

The use of field robots can greatly decrease the amount of time, effort, and associated risk compared to if human workers were to carryout certain tasks such as disaster response. However, transportability and reliability remain two main issues for most current robot systems. To address the issue of transportability, we have developed a lightweight modularizable platform named AeroArm. To address the issue of reliability, we utilize a multimodal sensing approach, combining the use of multiple sensors and sensor types, and the use of different detection algorithms, as well as active continuous closed‐loop feedback to accurately estimate the state of the robot with respect to the environment. We used Challenge 2 of the 2017 Mohammed Bin Zayed International Robotics Competition as an example outdoor manipulation task, demonstrating the capabilities of our robot system and approach in achieving reliable performance in the fields, and ranked fifth place internationally in the competition.     

Evaluating building fire safety through egress prediction: A standard application in Japan

Egress calculation, or predicting human flow during egress, is widely applied in Japan using a standardized method for evaluating the fire safety of building plans. This paper illustrates the methods involved in egress calculation using concrete examples and a sample plan. The egress calculation plays an important role in evaluating building plans and in complementing existing fire regulations. However, further investigation needs to be done into human behavior and how it factors into egress.     

High performance films of cellulose butyral derivative having a necklace-like annular structure in the side chains

We fabricated high performance films using cellulose butyral (CB) synthesized from native cellulose. Two-step reactions were adopted to produce the derivative CB, including etherification of cellulose with glycidol in NaOH/urea aqueous solution to yield O-(2,3-dihydroxypropyl) cellulose (DHPC), and butyralization of DHPC. Both DHPC and CB products were easily processed into a thin film by hot-press molding. The butyral modifier significantly improved the tenacity of highly ductile DHPC, by virtue of the possible chain-entangling action of the ring structures in the stretching process. Thereby the film toughness was markedly enhanced. The CB films exhibited excellent optical transparency and a good adhesive property to glass plates. Thus the films may be comparable to commercial poly(vinyl butyral) (PVB) films in optical and mechanical performances and therefore possess a potential applicability as interlayer for laminated glasses.     

Characteristic scales of two‐dimensional turbulence in polymer solutions

An experimental study has been performed to investigate the relationship between the extensional viscosity of polymers and the turbulent drag reduction. Self‐standing flowing soap film was used to generate two‐dimensional (2‐D) turbulent flow to eliminate shear stress. Two types of polymers having different flexibilities were added to the 2‐D turbulence. The effects of these polymers were visualized by the interference pattern of flowing soap films. The vortex deformation by adding polymers was analyzed by Fourier transformation and wavelet transformation. The scaling exponents of the power spectrum of interference patterns indicate that the mechanism of turbulence laminarization due to the extensional viscosity is anisotropic. A wavelet analysis reveals the high and low fluctuations of the polymer‐added flow. Results from wavelet analysis indicate disappearing of original vortices, and appearing of new structures in low frequency in 2‐D flow. © 2014 American Institute of Chemical Engineers AIChE J, 60: 1854–1862, 2014     

Dynamic mechanical properties of three‐component composites (acrylic polymer/epoxy/SiO2) in the glass‐transition region

In previous studies, we reported the linear and nonlinear rheological properties of three‐component composites consisting of acrylic polymer (AP), epoxy resin (EP), and various SiO₂ contents (AP/EP/SiO₂) in the molten state. In this study, the dynamic mechanical properties of AP/EP/SiO₂ composites with different particle sizes (0.5 and 8 μm) were investigated in the glass‐transition region. The EP consisted of three kinds of EP components. The α relaxation due to the glass transition shifted to a higher temperature with an increase in the volume fraction (?) for the AP/EP/SiO₂ composites having a particle size of 0.5 μm, but the α relaxation scarcely shifted for the composite having a particle size of 8 μm as a general result. This result suggested that the SiO₂ nanoparticles that were 0.5 μm in size adsorbed a lot of the low‐glass‐transition‐temperature (T_g) component because of their large surface area. The AP/SiO₂ composites did not exhibit a shift in T_g; this indicated that the composite did not adsorb any component. The modulus in the glassy state (E′_g) exhibited a very weak &phis; dependence for the AP/EP/SiO₂ composites having particle sizes of 0.5 and 8 μm, although E′_g of the AP/SiO₂ composites increased with &phis;. The AP/EP/SiO₂ composites exhibited a peculiar dynamic mechanical behavior, although the AP/SiO₂ composites showed the behavior of general two‐component composites. Scanning electron microscopic observations indicated that some components in the EP were adsorbed on the surface of the SiO₂ particles. We concluded that the peculiar behavior of the AP/EP/SiO₂ composites was due to the selective adsorption of the EP component. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40409.     

Electromagnetic silicon MEMS resonator

We have fabricated a silicon MEMS resonator aiming at multimodal sensors, and evaluated vibration characteristics by electromagnetic drive and induced electromotive force detection. The resonance frequency of the driving voltage of 0.6 Vpp shows torsional vibration of approximately 87 kHz, and the resonance frequency is shifted toward the low‐frequency side as the driving voltage increases. Resonance characteristics due to temperature change and film stress were evaluated.