Expression of a gene encoding chitinase (pCA 8 ORF) from Aeromonas sp. no. 10S-24 in Escherichia coli and enzyme characterization

A gene encoding chitinase from Aeromonas sp. no. 10S-24 was expressed using pTrc99A in Escherichia coli JM 105 which yielded a 5-fold higher activity than when pUC19 was used. Three different truncated enzymes (SA-1, SA-2 and SA-3) were obtained after purification. Their isoelectric points were 7.0, 6.9, and 6.7, respectively. The enzymes showed two optimum pHs, 4.0 and 7.0, when incubated with ethylene glycol chitin as the substrate, and were stable over a wide pH range (3.0-9.0). The optimum temperature was 60 degrees C and the enzymes were stable up to 50 degrees C. The chitinases exhibited wide substrate specificities for chitin-related compounds.     

Establishment of a simple system to analyse the molecular interaction in the agglutination of Saccharomyces cerevisiae

Saccharomyces cerevisiae a-agglutinin, which is involved in mating and covalently anchoring to the cell wall, consists of two components, Aga1p and Aga2p, whose syntheses are individually regulated. To facilitate the analysis of the protein-protein interaction on agglutination between a- and alpha-agglutinins, the construction of a yeast strain (MATa) with the functional protein prepared by genetic fusion of Aga1p- and Aga2p-encoding genes and by the expression system using the UPR-ICL promoter derived from the n-alkane-assimilating yeast, Candida tropicalis, which is functional under the condition of lower glucose concentration was tried and the agglutination ability of the constructed strain was evaluated with a yeast strain (MATa) which expressed AGalpha1 encoding alpha-agglutinin under the control of the same promoter. The genes were integrated into the yeast chromosomes. Cell agglutination between both (MATa) strains was observed microscopically when these two strains were mix-cultured to a glucose-decreased concentration. The agglutination was further confirmed by the sedimentation test and by the quantification using a filter. These results proved that the constructed Aga1p-Aga2p fusion protein was enoughly functional for the interaction with the Agalpha1 protein, and that this phenomenon occurred dependent on glucose concentration, but independent of the peptide pheromones secreted by the cells of the opposite mating types. Using this system, the role of two disulphide linkages between Aga1p and Aga2p on the binding activity between Aga2p and Aga1p was first evaluated. Under the treatment by the SH-compound (dithiothreitol), in which Agalpha2p is easily released into the medium from the intact cell surface, the Aga1p and Aga2p fusion protein was a good tool to make clear the role of the disulphide linkages. As a result, the linkages had a significant effect on not only the assembly but also the binding activity. The novel and simple system described here may further facilitate the study of molecular interaction in agglutination.     

TRANSIENT THERMAL SINGULAR STRESSES OF MULTIPLE CRACKING IN A W-Cu FUNCTIONALLY GRADED DIVERTOR PLATE

We consider the transient thermal singular stresses of multiple cracking in a functionally graded divertor plate due to a thermal shock. The plate is made of a graded layer bonded between a homogeneous substrate and a homogeneous coating, and it is subjected to a cycle of heating and cooling on the coating surface of the plate. The surface layer contains a parallel array of embedded or edge cracks perpendicular to the boundaries. The thermal and elastic properties of the material are dependent on the temperature and the position. Finite element calculations are carried out, and the transient thermal stress intensity factors are shown graphically.     

Ultrashort X-ray pulse generation using subpicosecond electron linac

As a promising tool for ultrafast material analyses, we propose to utilize the X-ray pulse which may be generated in a quite simple manner using subpicosecond electron linacs. The properties of the X-ray were numerically studied with the EGS4 code. Verification of the X-ray generation was also conducted at the Nuclear Engineering Research Laboratory (NERL) linac and clear diffraction patterns of characteristic X-ray were obtained for typical single crystals.     

A geometric method of sector decomposition

We propose a new geometric method of IR factorization in sector decomposition. The problem is converted into a set of problems in convex geometry. The latter problems are solved using algorithms in combinatorial geometry. This method provides a deterministic algorithm and never falls into an infinite loop. The number of resulting sectors depends on the algorithm of triangulation. Our test implementation shows smaller number of sectors comparing with other existing methods with iterations.     

Direct electron microscopic observation of transcrystalline layers in microfibrillar reinforced polymer-polymer composites

Microfibrillar reinforced composites (MFC) comprising an isotropic matrix from a lower melting polymer, i.e., low density polyethylene (LDPE), reinforced by microfibrils of a higher melting polymer, recycled from bottles, i.e., poly(ethylene terephthalate) (PET), were processed under industrially relevant conditions via injection molding in a weight ratio of PET/LDPE = 50/50. Dog bone samples with MFC structure were characterized by means of scanning (SEM) and transmission (TEM) electron microscopy. SEM observations on cryogenic fracture surfaces show an isotropic LDPE matrix reinforced by more or less randomly distributed PET microfibrils. By means of TEM on stained ultrathin slices one observes the formation of transcrystalline layers of LDPE matrix on the surface of the PET microfibrils. In these layers the crystalline lamellae are aligned parallel to each other and are placed perpendicularly to the fibril surfaces. This is in contrast to the bulk matrix where the lamellae are quasi-randomly arranged.     

Development and evaluation of a diamond electron source for electron beam instruments

Since diamonds indicated low or negative electron affinity, they have been expected to be high performance electron emitters. New diamond electron sources have been developed, which are configured for electron beam instruments such as electron beam lithography systems and electron microscopes. As a result of thermionic emission evaluations, an electron emission current of practical level (116 µA at 600 °C) was obtained. The energy spread of an electron beam generated by thermionic emission was measured as 0.23 eV (FWHM), the value of which was lower than those of a lanthanum hexaborate (LaB₆) cathode and a zirconium oxide/tungsten (ZrO/W) cathode measured together. The result of field emission evaluations, a practical beam current (459 pA) and stability (6% rms for 10 h) for scanning electron microscope (SEM) was achieved. The energy spread of a high-current electron beam of over 200 pA generated by field emission from a diamond emitter reached about the same as that of a cold field emission cathode made of tungsten. A scanning electron micrograph (SEM) with a diamond electron source having a tip of submicron order allowed measurement a resolution of 5 nm. It was found that unprecedentedly high-current and high-convergence electron sources could be generated by using diamonds.     

Mixed powder coating film using thermoplastic polyester and its alkaline resistance

To improve alkaline resistance, a newly mixed powder coating film using thermoplastic polyethylene terephthalate (PET) was investigated. Two kinds of polyvinyl butyral (PVB) and two kinds of polyamide (PA) were chosen as the secondary polymer. The melting temperatures (T _ms) of these materials were lower than that of primary PET polymer so the mixed powder coating was able to form a dual phase film through a fluidized bed coating process. Microscope and Fourier transform infrared spectroscopy (FTIR) observations revealed that the dual phase structure was indeed successfully formed and there was a secondary layer over the entire surface area of the film. This mixed powder coating film significantly enhanced the alkaline resistance to an environment filled with NaOH solution at 40°C in which a secondary material would be effective in protecting the film. With regard to both alkaline resistance and film formability, PET/PVB was superior to the PET/PA mixture.     

Phase-diagram study for the Al2O3-CaF2-SiO2 system

Phase relations for the Al₂O₃-CaF₂-SiO₂ system were investigated from 1673 to 1723 K. The hotfilament technique was applied to observe a two-liquid region and liquidus for the ternary system. The liquidus saturated with SiO₂ was investigated at 1673 K by using the hot-filament technique and the chemical equilibrium technique. In this system, the addition of Al₂O₃ to the CaF₂-SiO₂ system reduces the congruent temperature. A small substitution of Al₂O₃ for CaF₂ increases the solubility of SiO₂, namely, if the region of liquid phase could be enlarged. These results suggest that Al₂O₃ would be an effective substitute of CaF₂ in slag for steelmaking.