Dibenzothiophene desulfurizing enzymes from moderately thermophilic bacterium Bacillus subtilis WU-S2B: purification, characterization and overexpression

The moderately thermophilic bacterium Bacillus subtilis WU-S2B desulfurized dibenzothiophene (DBT) at 50 degrees C through the selective cleavage of carbon-sulfur bonds. In this study, three enzymes involved in the microbial DBT desulfurization were purified and characterized. The first two enzymes, DBT monooxygenase (BdsC) and DBT sulfone monooxygenase (BdsA), were purified from the wild-type strain, and the last one, 2'-hydroxybiphenyl 2-sulfinic acid desulfinase (BdsB), was purified from the recombinant Escherichia coli overexpressing the gene, bdsB, with chaperonin genes, groEL/ES. The genes of BdsC and BdsA were also overexpressed. The molecular weights of BdsC and BdsA were determined to be 200 and 174 kDa, respectively, by gel filtration chromatography, suggesting that both enzymes had four identical subunits. BdsB had a monomeric structure of 40 kDa. The three enzymes were characterized and compared with the corresponding enzymes (DszC, DszA, and DszB) of mesophilic desulfurization bacteria. The specific activities of BdsC, BdsA, and BdsB were 84.2, 855, and 280 units/mg, respectively, and the latter two activities were higher than those of DszA and DszB. The heat stability and optimum temperature of BdsC, BdsA, and BdsB were higher than those of DszC, DszA, and DszB. Other enzymatic properties were investigated in detail.     

The oxidation behavior of an iron-nickel alloy containing yttrium or rare earth elements between 900°C and 1200°C

A study of oxidation of an iron-nickel alloy with and without yttrium or rare earth additions is made to provide information over a range of temperature. The additions improve the oxidation resistance in air. The microfractograph features of oxide scale are discussed. It is suggested that pores existing in the interface between oxide scale and substrate may be an important factor for the good adherence which is obtained. In addition the pegging mechanism of rare earth element oxides may improve the adherence of oxide scale to substrate. An incubation period of oxidation in the alloys containing yttrium or rare earth elements is described and discussed in terms of net weight gain and weight loss due to gas evaporation.     

Development of an Artificial Lipid-Based Membrane Sensor with High Selectivity and Sensitivity to the Bitterness of Drugs and with High Correlation with Sensory Score

This paper reports the development of membrane sensors based on an artificial lipid and plasticizers with high selectivity and sensitivity to drug bitterness by using bis(1-butylpentyl) adipate (BBPA), bis(2-ethylhexyl) sebacate (BEHS), phosphoric acid tris(2-ethylhexyl) ester (PTEH), and tributyl o-acetylcitrate (TBAC) as a plasticizer and phosphoric acid di-n-decyl ester (PADE) as an artificial lipid to optimize surface hydrophobicity of the sensors. In addition, a sensor with highly correlated bitterness sensory score was developed by blending BBPA and TBAC to detect the bitterness suppression effect of sucrose, and other bitter-masking materials. Therefore, this sensor can be used to evaluate the bitterness of various drug formulations with high accuracy. Copyright © 2009 Institute of Electrical Engineers of Japan. Published by John Wiley & Sons, Inc.     

Improvement and induction property of radiation-responsive promoter through DNA shuffling of 5′-flanking regions of the human p21 gene

A promoter that augments gene expression in response to stimulation of ionizing radiation would be a desired tool for radiogenetic therapy, a combination of radiotherapy and gene therapy. Although various promoters occurring naturally or artificially have been used for researches, one showing higher reactivity to ionizing radiation is desirable. In the present study, we attempted to improve a radiation-responsive promoter of the p21 through a technique called DNA shuffling. A library of DNA fragments was constructed by re-ligation of randomly digested promoter fragments and improved promoters were chosen out of the library. We repeated this process twice to obtain a promoter showing 2.6-fold better reactivity to ionizing radiation compared with its parent, p21 promoter after 10 Gy γ-ray irradiation. Nucleotide sequence analyses revealed that the obtained promoter was densely packed with some of the cis-acting elements including binding sites for p53, NF-κB, NRF-2, AP-1 and NF-Y more than p21 promoter. In addition, it was shown that its induction by ionizing radiation was dependent upon p53 status of a cell line, suggesting that the promoter retained properties of the p21 promoter. This technique is simple and efficient to improve a promoter responsive to other stimulus of interest besides IR.