Biophysical analysis of heat-induced structural maturation of glutamate dehydrogenase from a hyperthermophilic archaeon

Tertiary structure of the recombinant glutamate dehydrogenase from Thermococcus kodakaraensis KOD1 (Tk-rGDH) converts into an intact form induced by the heat treatment. This phenomenon, heat-induced structural maturation, means that high temperature plays an important role in the proper folding and oligomerization of Tk-rGDH. In this work, we analyzed the heat-induced structural maturation of Tk-rGDH by differential scanning microcalorimetry (DSC), circular dichroism (CD), and activity measurements. In DSC measurements, the peak of adsorption of non-heated Tk-rGDH (nh-Tk-rGDH) was two times smaller than that of Tk-rGDH heated at 70 degrees C for 30 min (h-Tk-rGDH). The transition temperature (T(m)) of h-Tk-rGDH was 115 degrees C, which was about 3 degrees C higher than that of nh-Tk-rGDH. In the presence of 0.5 M NaCl, the nh-Tk-rGDH showed two peaks at 107 degrees C and 114 degrees C, while the h-Tk-rGDH showed a single peak at 115.7 degrees C. The heat-induced conformational change process was monitored by changes in CD intensity at 222 nm, and the result showed that heat-induced structural maturation is irreversible. The heat treatment at 70 degrees C showed the highest enhancement in activity, which was 15% larger than that of heat-treated Tk-rGDH at 40 degrees C. The results indicate that heat-induced structural maturation involves an irreversible process, transforming the non-heated form to the stable and active form.     

Synthesis of Oriented Ba2NaNb5O15 (BNN) Thin Films from an Alkoxy-derived Precursor

Ba₂NaNb₅O₁₅ (BNN) thin films with a tungsten bronze structure were fabricated using a precursor solution that was synthesized from barium, sodium, and niobium alkoxides. Highly (002)-oriented BNN films were prepared successfully on Pt(100)/MgO(100) substrates at 700°C, using a BNN underlayer. X-ray pole-figure measurement showed that the BNN films that crystallized on Pt(100)/MgO(100) substrates had two in-plane orientations. The remanent polarization and coercive field of the BNN film (thickness of 1.0 µm) that was crystallized at 700°C were 12.3 µC/cm² and 101 kV/cm, respectively, at –150°C (123 K). BNN films on fused silica substrates exhibited second harmonic generation upon irradiation with 1064 nm light.     

Fast response varifocal lenses using KTa(1-x)Nb(x)O3 crystals and a simulation method with electrostrictive calculations

We fabricated cylindrical varifocal lenses with fast responses by using the strong Kerr effect of KTa(1-x)Nb(x)O(3) (KTN) single crystals. We observed focus shifts of up to 87 mm with the assistance of a 250 mm focal length lens, which corresponds to a focus shift from infinity to 720 mm by the KTN lens itself. The response time was as fast as 1 μs. We also present a simulation method for calculating refractive index distributions in KTN single crystals, which is essential when designing the lens. The method is characterized by the strain contribution, which has not conventionally been typical of electro-optic simulations. We used this method to explain the refractive index modulations that are characteristic of the varifocal lenses.     

A verification and validation method and its application to digital safety systems in ABWR nuclear power plants

A verification and validation (V&V) method has been developed and applied to newly developed digital safety systems for the first ABWR plant, Kashiwazaki–Kariwa unit No. 6 of the Tokyo Electric Power Company. This paper describes the method and experience gained from its application. The method was developed on the basis of domestic and foreign standards and guidelines, and covers the more concrete procedures required for actual V&V. The application of problem oriented language (POL) helps make the V&V feasible and reliable. A personal-computer-based automatic test tool for the validation test has been developed and utilized. This tool is used to carry out the pre-defined validation test procedure automatically and produce a test report, and it was found to be effective in reducing the time and manpower required for the validation test. The validation test covers dynamic transient tests in which the response of the digital safety system against the simulated design based transients are tested. The tool enables automatic execution of the dynamic test.     

Three-dimensional multiinkdot automata

Recently, related to the open problem whether deterministic and nondeterministic space (especially lower-level) complexity classes are separated, inkdot Turing machines were introduced. On the other hand, due to the advances in many application areas about three-dimensional information processing, the research on three-dimensional automata as the computation of three-dimensional pattern processing has been meaningful. From this viewpoint, we propose a three-dimensional multiinkdot automata, and investigate some properties of three-dimensional multiinkdot automata.This work was presented, in part, at the 9th International Symposium on Artificial Life and Robotics, Oita, Japan, January 28–30, 2004     

Thermal-hydraulic behavior of a marine reactor during oscillations

The effect of ship motion, such as heaving and rolling, on the thermal-hydraulic behavior of marine reactors was investigated. The COBRA-IV-I CODE was modified to analyse the thermal-hydraulic performance on the critical heat flux under oscillating acceleration conditions. The critical heat flux in the code was verified experimentally using freon as a comparison. The Critical Heat Flux Ratio (CHFR) at the hottest channel of the PWR subchannel was analysed using the same code. A system code RETRAN-02/MOD2-GRAV was developed by improving RETRAN-02/MOD2 to simulate the thermal hydraulic transient under ship motion. It was verified by comparison using the experimental results of both two-phase natural circulation flow under heaving motion and single-phase natural circulation flow at an inclined attitude. The code was used to analyse reactor plant behavior in the nuclear ship Mutsu. Natural circulation flow during rolling motion was investigated experimentally. The characteristics of loop flow and core flow rates were clarified. The core flow rate correlated well with the Reynolds number of rolling motion.     

A microscopic study on the oxidative stabilization of a coal-tar-based mesophase pitch and its blends with PVC pitch

Coal-tar-based mesophase pitch and its blends with PVC pitch at 5 or 10 wt% were oxidatively stabilized at 230, 270 and 300°C for variable periods to clarify the progress of stabilization and the effects of the blending with PVC pitch on the stabilization reactivity. PVC pitch which was prepared from PVC by heat-treatment at 420°C for 2h enhanced the stabilization reactivity of whole pitch fibres to shorten the stabilization time to a half of that for mesophase pitch alone. PVC pitch carrying considerable amounts of aliphatic components and large molecular weight may initiate, as a trigger, the stabilization reactions of mesophase constituent molecules. Carbonized fibres of 30μm diameter after stabilization at 270 and 300°C exhibited a skin-core structure, while fibres of 10μm diameter showed no skin-core structure, indicating a homogeneous progress of stabilization in the radial direction of the latter fibres. Lower stabilization temperatures provoked no skin-core structure even in the thick fibres. The rate of core diminishing became relatively slower in the later stage of the stabilization, even when the reactivity of the pitch fibres was enhanced by blending with PVC pitch and using a higher stabilization temperature. The diffusion of the oxidant and stabilization reactivity of the pitch fibres are discussed comparatively.     

Gene cloning and characterization of fructose-1,6-bisphosphate aldolase from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1

The fructose-1,6-bisphosphate (FBP) aldolase gene from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 was cloned. The gene encoding FBP aldolase (Tk-Fba) was expressed in Escherichia coli and the purified recombinant protein was characterized at high temperature. Tk-Fba is a homodecamer with a subunit molecular mass of 31,283 Da. The amino acid sequence, decameric conformation, formation of a Schiff-base intermediate, and stimulation (286%) of FBP cleavage activity by citrate suggested that Tk-Fba belonged to Class IA, a subtype of the classical Class I aldolases. The specific activity for the FBP cleavage reaction was 18.9 U/mg, which was much higher than those of other Class IA type FBP aldolases. Tk-Fba was extremely thermostable since the optimum temperature seemed to be above 100 degrees C. The optimum pH for Tk-Fba was determined to be 5.0 in the absence of citrate, while it shifted to around 7.0 in the presence of citrate. Tk-Fba accepted FBP and fructose-1-phosphate as substrates and K(m) values were determined to be 0.063 mM and 4.37 mM, respectively. In addition to citrate, phosphoenolpyruvate and pyrophosphate were also found to be potent activators of Tk-Fba, enhancing activities up to 346% and 201%, respectively. Erythrose-4-phosphate acted as an inhibitor and caused a decrease in the activity to 49%. Tk-Fba also catalyzed the condensation reaction with a similar activity level (14.9 U/mg) to that for FBP cleavage. However, none of the above compounds seemed to have a significant effect on the condensation reaction by Tk-Fba. These results suggest a regulatory function of Tk-Fba toward the catabolic direction of sugar metabolism in T. kodakaraensis KOD1.     

An Integrated Field-Effect Electrode for Biopotential Recording

An integrated liquid-oxide-semiconductor FET structure is described. The structure is similar to a conventional n-channel depletion mode MOSFET except the gate metal is omitted. Immersing the device in a saline solution, the drain current is controlled by the potential between the source and the solution. Operated as an active recording electrode, the structure has several properties useful in neurological research.