Identification of an insertion sequence, IS1203 variant, in a Shiga toxin 2 gene of Escherichia coli O157:H7

An insertion sequence composed of 1310 bp was found in Shiga toxin 2 genes of some isolates of Escherichia coli O157:H7. This insertion sequence showed extremely high homology with IS1203 of E. coli O111:H(-). This IS1203 variant was inserted in the region encoding the amino-terminus of the B subunit with a duplication of 3 bp at the target site, resulting in inactivation of the Shiga toxin 2 gene.     

Host-dependent activation of IS1203v excision in Shiga toxin-producing Escherichia coli

IS1203v is an insertion sequence (IS) which is identical to the most abundant IS elements in the genome of Escherichia coli O157:H7. However, there is no sequence homologous to IS1203v in the genome of E. coli K-12. We constructed a system to analyze the excision frequency of IS1203v, and demonstrated that the frequency in E. coli O157:H7 was approximately 10(5) times higher than that in E. coli K-12. We also investigated the excision frequencies of IS1203v in various E. coli isolates, and showed that the excision frequencies of IS1203v-possessing strains were approximately 10(3) times higher than those of IS1203v-nonpossessing strains. The results suggest that the IS1203v-possessing strains use a common system to enhance IS1203v excision.     

Identification and Characterization of D‐Succinylase,and a Proposed Enzymatic Method for D‐Amino Acid Synthesis

Chiral amino acids are important intermediates for the pharmaceutical industry. We have developed a novel one‐pot enzymatic method for D ‐amino acid synthesis by the dynamic kinetic resolution of N‐succinyl‐dl ‐amino acids using D ‐succinylase (DSA) and N‐succinylamino acid racemase (NSAR, EC 4.2.1.113). The DSA from Cupriavidus sp. P4‐10‐C, which hydrolyzes N‐succinyl‐D ‐amino acids enantioselectively to their corresponding D ‐amino acids, was identified for the first time by screening soil microorganisms. Subsequently, the DSA gene was cloned and overexpressed in Escherichia coli. DSA was shown to comprise two subunits with molecular masses of 26 kDa and 60 kDa. Additionally, the NSAR gene from Geobacillus stearothermphilus NCA1503, which racemizes N‐succinylamino acids, was also cloned and overexpressed in E. coli. The highly purified DSA and NSAR prepared from each recombinant E. coli were characterized and used for D ‐amino acid synthesis. A one‐pot enzymatic method converted 100 mM N‐succinyl‐dl ‐phenylalanine to D ‐phenylalanine in 91.1% conversion with 86.7% ee. This novel enzymatic method may be useful for the industrial production of many D ‐amino acids.

     

Preventive control based on complex-valued load-flow technique to avoid voltage instability in power systems

This paper presents a method for obtaining preventive control strategies based on nonlinear optimization for power systems which will incur voltage instability when load demand increases. An algorithm is proposed to determine an appropriate and effective control action taking into account the operating constraints to widen the margin between the present operating point and a voltage collapse point, thereby improving the system state. The method proposed here is based on a complex-valued load-flow technique using the Newton-Raphson method which has been developed already by the authors. Consequently, the preventive control algorithm also can evaluate the voltage instability in the event the present operating point becomes closer to the critical point and the system state becomes unfeasible at increased loading point. Further, in optimization, two-types of objective functions are introduced so that voltage stabilization and dissolution of constraint violation can be attained simultaneously. In addition, this paper discusses the extension of the proposed preventive control to the successive control method which carries out the control action to avoid voltage collapse while the load demand is increasing. Numerical examples for various model systems demonstrate the effectiveness of the proposed method.     

Optimal planning for introducing dispersed generating sources in power systems based on an economical viewpoint

This paper discusses optimal planning problems for dispersed generating sources so as to provide a guideline for their introduction to power systems. First, a recursive method is developed from the viewpoint of supplying electric power only where solutions for one unit are superposed. The computational loads are decreased by selecting alternatives of buses based on the so-called penalty factor. However, for effective utilization of dispersed generating sources, it is indispensable to consider their thermal merits in addition to electric power. After applying the Kuhn-Tucker theory to the optimal dispatching problem including boiler fuel costs, thermal values of the dispersed generating units are evaluated quantitatively. Finally, the above-mentioned technique for the optimal planning basically is also effective and is extended into this case. The validity of the proposed algorithms are demonstrated using a real-scale model system.     

Improved coupling efficiency of a strained InGaAs-AlGaAs quantum-well laser into a fiber Bragg grating

We describe a highly efficient coupling of up to 78% into a single mode fiber (SMF) by utilizing a low-divergence-angle InGaAs-AlGaAs laser and a new high-NA aspheric lens, resulting in the coupled optical output of over 300 mW with a kink-free performance. The kink-free 300 mW of fiber-coupled power is the largest power reported from a diode laser wavelength stabilized using a fiber Bragg grating. A 3% fiber Bragg grating makes the optical output stabilize at a wavelength of 1.016 /spl mu/m for an optical output power up to 310 mW and without any distinct decrease of the optical output. The coupling tolerance for lateral offsets is improved by utilizing a thermal-diffused expanded-core technique even in the case of using a high-NA coupling aspheric lens. We have also achieved a 300-mW fiber-coupled wavelength-stabilized InGaAs-AlGaAs semiconductor laser butterfly module using a fiber Bragg grating.     

Reconstructed three-dimensional images of flow-sorted nuclei hybridized with chromosome-specific probes

We demonstrated that the three-dimensional (3-D) locational and morphological differences of chromosome 17 are dependent on each cell cycle phase in the clinical materials. Cell suspensions prepared from hypertrophied tonsil were hybridized with chromosome 17 whole painting probe or its centromeric probe and the probes were detected with fluorescein isothiocyanate. Then the cells were sorted from G(0+1), S-, and G(2+M)-phase fractions by flow cytometry and observed by confocal laser scanning microscopy to obtain the serial optical sections. The 3-D images were obtained by assembling these sections using a computerized image analysis device. The distribution of centromeric copies was analyzed statistically, and the data values were not a population of random distribution within a sphere. The copies were observed in the periphery of the nuclei in G(0+1)- and S-phase. The 3-D images revealed that chromosome 17 was oval in shape in the G(0+1)-phase nucleus, and was changing into a flame shape in the S-phase, with arms stretching out along the nuclear membrane, and looked bush shaped in G2-phase. The eccentric distribution of chromosome 17 in G(0+1)- and S-phase nuclei may reflect the optimal efficiency of incorporating and/or releasing essential materials and products.     

Effect of non-ionic detergents on apparent enzyme mechanism: V121A mutant of Streptomyces cholesterol oxidase endowed with enhanced sensitivity towards detergents

One of the mutants of Streptomyces cholesterol oxidase with the Val121Ala mutation (V121A) was kinetically analysed. Although the reaction rate-substrate concentration curve of wild type follows a simple Michaelis-Menten equation, that of V121A is sigmoidal. The cooperativity was apparent and caused by non-ionic detergents that were used as a solvent of cholesterol. The concentration dependence of V121A on detergents was more significant than that of wild type, although the reaction rates of both enzymes decrease as the concentrations of detergents increase. Further experiments suggested that less hydrophobic interactions between V121A and detergents should be responsible for the apparent cooperativity. Since Val121 is in a hydrophobic loop located near the active site, the mutational effect is structurally discussed.