Isolation and characterization of a gene coding for chitin deacetylase specifically expressed during fruiting body development in the basidiomycete Flammulina velutipes and its expression in the yeast Pichia pastoris

ArticleFEMS MICROBIOLOGY LETTERS. 289(2):130-137 (2008)journal articl

Purification and characterization of UDP-glucose: hydroxycoumarin 7-O-glucosyltransferase, with broad substrate specificity from tobacco cultured cells

The enzyme UDP-glucose: hydroxycoumarin 7-O-glucosyltransferase (CGTase), which catalyzes the formation of scopolin from scopoletin, was purified approximately 1200-fold from a culture of 2,4-D-treated tobacco cells (Nicotiana tabacum L. cv. Bright Yellow T-13) with a yield of 7%. Purification to apparent homogeneity, as judged by SDS-PAGE, was achieved by sequential anion-exchange chromatography, hydroxyapatite chromatography, gel filtration, a second round of anion-exchange chromatography, and affinity chromatography on UDP-glucuronic acid agarose. The purified enzyme had a pH optimum of 7.5, an isoelectric point (pI) of 5.0, and a molecular mass of 49 kDa. The enzyme did not require metal cofactors for activity. Its activity was inhibited by Zn2+, Co2+ and Cu2+ ions, as well as by SH-blocking reagents. The K-m values for UDP-glucose, scopoletin and esculetin were 43, 150 and 25 mu M. respectively. A study of the initial rate of the reaction suggested that the reaction proceeded via a sequential mechanism. The purified enzyme preferred hydroxycoumarins as substrates but also exhibited significant activity with flavonoids. A database search using the amino terminus amino acid sequence of CGTase revealed strong homology to the amino acid sequences of other glucosyltransferases in plants.ArticlePlant Science. 157(1):105-112 (2000)journal articl

Trends in life science grid: from computing grid to knowledge grid

BACKGROUND: Grid computing has great potential to become a standard cyberinfrastructure for life sciences which often require high-performance computing and large data handling which exceeds the computing capacity of a single institution. RESULTS: This survey reviews the latest grid technologies from the viewpoints of computing grid, data grid and knowledge grid. Computing grid technologies have been matured enough to solve high-throughput real-world life scientific problems. Data grid technologies are strong candidates for realizing "resourceome" for bioinformatics. Knowledge grids should be designed not only from sharing explicit knowledge on computers but also from community formulation for sharing tacit knowledge among a community. CONCLUSION: Extending the concept of grid from computing grid to knowledge grid, it is possible to make use of a grid as not only sharable computing resources, but also as time and place in which people work together, create knowledge, and share knowledge and experiences in a community

NK Cell–Like Behavior of Vα14i NK T Cells during MCMV Infection

Immunity to the murine cytomegalovirus (MCMV) is critically dependent on the innate response for initial containment of viral replication, resolution of active infection, and proper induction of the adaptive phase of the anti-viral response. In contrast to NK cells, the Vα14 invariant natural killer T cell response to MCMV has not been examined. We found that Vα14i NK T cells become activated and produce significant levels of IFN-γ, but do not proliferate or produce IL-4 following MCMV infection. In vivo treatment with an anti-CD1d mAb and adoptive transfer of Vα14i NK T cells into MCMV-infected CD1d−/− mice demonstrate that CD1d is dispensable for Vα14i NK T cell activation. In contrast, both IFN-α/β and IL-12 are required for optimal activation. Vα14i NK T cell–derived IFN-γ is partially dependent on IFN-α/β but highly dependent on IL-12. Vα14i NK T cells contribute to the immune response to MCMV and amplify NK cell–derived IFN-γ. Importantly, mortality is increased in CD1d−/− mice in response to high dose MCMV infection when compared to heterozygote littermate controls. Collectively, these findings illustrate the plasticity of Vα14i NK T cells that act as effector T cells during bacterial infection, but have NK cell–like behavior during the innate immune response to MCMV infection

Purification and Characterization of a Novel Bacteriocin Produced by Enterococcus faecalis Strain RJ-11

APPEAL FROM THE THIRD JUDICIAL DISTRICT COURT, SALT LAKE COUNTY, JUDGE LESLIE A. LEWIS PRESIDING, DENYING DEFENDANT\u27S MOTION TO SUPPRESS EVIDENC

Transformation of Tobacco Cultured Cell by Particle Bombardment : Expression of Phenylalanine Ammonia-lyase Gene in Transgenic Tobacco Cells.

The plasmid DNA containing a NPTⅡ as a selectable marker gene and a tobacco PAL cDNA combined with the CaMV 35S promoter and NOS terminator was transformed into tobacco cells by a particle bombardment device based on flowing helium. The most effcient transformation was achieved when the amount of DNA-coated tungsten particle was 0.2㎎ per projectile, the amount of plasmid DNA was 4μg/㎎ of tungsten particles, the helium pressure accelerated was 3㎏/c㎡, and the distance between the sample and the projectile was 15cm. Genetic analysis of kanamycin-resistant transformants obtained　showed that the PAL cDNA construct integrated into the genome of tobacco cells. PAL activity of the transformant increased almost 4-fold and scopoletin content increased more than 2-fold as compared to nontrasformed cells.ヘリウムガス圧式パーティクルガンにより，タバコ培養細胞のPALcDNAを同細胞へ再導入し，形質転換体を得た．タバコ培養細胞のPALcDNAをカリフラワーモザイクウイルス(CaMV)の35Sプロモーターとノパリン合成酵素(NOS)遺伝子のターミネーターに接続し，ネオマイシンホスホトランスフェラーゼⅡ(NPTⅡ)遺伝子とともにタバコ培養細胞へ導入し，形質転換体をカナマイシン培地で選抜した．また，パーティクルガンによる遺伝子導入の最適条件としては，加速圧力3㎏/c㎡，0.2㎎タングステン粒子/プロジェクタイル，4μg/㎎タングステン粒子，プロジェクタイルからサンプルまでの距離は15㎝であった．そして得られた形質転換体の中には，非形質転換体と比較して，PAL酵素活性で4倍，スコポレチン生成量で2倍以上のものが確認された．ArticlePlant tissue culture letters. 12(2):165-171 (1995)journal articl