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11.
Production,purification and characterisation of a novel halostable xylanase from Bacillus sp. NTU-06
Bacillus sp. NTU-06 was used to produce xylanase, which is an important industrial enzyme used in the pulp and paper industry. The enzyme was purified by fast protein liquid chromatography (FPLC) and had a molecular mass of 24 kDa. The enzyme was active over a concentration range of 0–20% sodium chloride in culture broth, although its activity was optimal in 5% sodium chloride. A salinity stability test showed that 43% of the enzyme activity was retained after 4 h in 20% sodium chloride. Xylanase activity was maximal at pH 8.0 and 40°C. The enzyme was somewhat thermostable, retaining 20% of the original activity after incubation at 70°C for 4 h. The xylanase had Km and Vmax values of 3.45 mg mL−1 and 387.3 µmol min−1mg−1, respectively. The deduced internal amino acid sequence of Bacillus sp. NTU-06 xylanase resembled the sequence of beta-1,4-endoxylanase, which is a member of glycoside hydrolase family 11. Some of the novel characteristics that make this enzyme potentially effective in xylan biodegradation are discussed. 相似文献
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Colloidal gold-based immunochromatographic assay for detection of botulinum neurotoxin type B 总被引:6,自引:0,他引:6
Chiao DJ Shyu RH Hu CS Chiang HY Tang SS 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2004,809(1):37-41
A rapid immunochromatographic assay was developed to detect botulinum neurotoxin type B (BoNT/B). The assay was based on the sandwich format using polyclonal antibody (Pab). The thiophilic gel purified anti-BoNT/B Pab was immobilized to a defined detection zone on a porous nitrocellulose membrane and conjugated to colloidal gold particles that served as a detection reagent. The BoNT/B-containing sample was added to the membrane and allowed to react with Pab-coated particles. The mixture was then passed along the porous membrane by capillary action past the Pab in the detection zone, which will bind the particles that had BoNT/B bound to their surface, giving a red colour within this detection zone with an intensity proportional to BoNT/B concentration. In the absence of BoNT/B, no immunogold was bound to the solid-phase antibody. With this method, 50 ng/ml of BoNT/B was detected in less than 10 min. The assay sensitivity can be increased by silver enhancement to 50 pg/ml. The developed BoNT/B assay also showed no cross reaction to type A neurotoxin (BoNT/A) and type E neurotoxin (BoNT/E). 相似文献
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Human TOB, an antiproliferative transcription factor, is a poly(A)-binding protein-dependent positive regulator of cytoplasmic mRNA deadenylation 总被引:1,自引:0,他引:1 下载免费PDF全文
Ezzeddine N Chang TC Zhu W Yamashita A Chen CY Zhong Z Yamashita Y Zheng D Shyu AB 《Molecular and cellular biology》2007,27(22):7791-7801
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This study was to examine the immune response of white shrimp Litopenaeus vannamei and its resistance against Vibrio alginolyticus and WSSV when shrimp received the Sargassum hemiphyllum var. chinense powder and its hot-water extract. Both powder and extract showed activation of prophenoloxidase and generation of superoxide anion in the shrimp in vitro. The haemocyte count, phenoloxidase (PO) activity, respiratory burst, and lysozyme activity were examined after the shrimp were immersed in seawater containing S. hemiphyllum var. chinense powder or its extract at 0, 100, 300, and 500 mg L?1 for 1, 3, and 5 h. These immune parameters of shrimp immersed in 300 and 500 mg L?1 powder, and 100 and 300 mg L?1 extract were significantly higher than those of control shrimp after 3 h, but slightly decreased after 5 h. In another experiment, shrimp immersed in seawater containing the powder or the extract at 0, 100, 300, and 500 mg L?1 after 3 h were challenged with V. alginolyticus at 8 × 10? colony-forming unit (cfu) shrimp?1, or challenged with WSSV at 1 × 10? copies shrimp?1, and then placed in seawater. Survival rate of shrimp immersed in 500 mg L?1 powder was significantly higher than that of control shrimp after 24-120 h in the V. alginolyticus-challenge test, and after 72 h in the WSSV-challenge test, respectively. Survival rate of shrimp immersed in 300 mg L?1 extract was significantly higher than that of control shrimp after 72-120 h in both V. alginolyticus-challenge and WSSV-challenge tests. It was concluded that the shrimp immersed in seawater containing the powder at 500 mg L?1, and the extract at 300 mg L?1 had increased immunity and resistance against V. alginolyticus infection, and the shrimp that received extract at 300 mg L?1 showed resistance against WSSV infection. 相似文献
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Yamashita A Chang TC Yamashita Y Zhu W Zhong Z Chen CY Shyu AB 《Nature structural & molecular biology》2005,12(12):1054-1063
In mammalian cells, the enzymatic pathways involved in cytoplasmic mRNA decay are incompletely defined. In this study, we have used two approaches to disrupt activities of deadenylating and/or decapping enzymes to monitor effects on mRNA decay kinetics and trap decay intermediates. Our results show that deadenylation is the key first step that triggers decay of both wild-type stable and nonsense codon-containing unstable beta-globin mRNAs in mouse NIH3T3 fibroblasts. PAN2 and CCR4 are the major poly(A) nucleases active in cytoplasmic deadenylation that have biphasic kinetics, with PAN2 initiating deadenylation followed by CCR4-mediated poly(A) shortening. DCP2-mediated decapping takes place after deadenylation and may serve as a backup mechanism for triggering mRNA decay when initial deadenylation by PAN2 is compromised. Our findings reveal a functional link between deadenylation and decapping and help to define in vivo pathways for mammalian cytoplasmic mRNA decay. 相似文献
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The retinoid-inducible gene 1 (RIG1) protein is a retinoid-inducible growth regulator. Previous studies have shown that the RIG1 protein inhibits the signaling pathways of Ras/mitogen-activated protein kinases. However, neither the mode of action nor the site of inhibition of RIG1 is known. This study investigated the effects of RIG1, and the mechanisms responsible for these effects, on the activation of Ras proteins in HtTA cervical cancer cells. RIG1 reduced the levels of activated Ras (Ras-GTP) and total Ras protein in cells transfected with mutated H-, N-, or K-Ras(G12V), or in cells transfected with the wild type H- or N-Ras followed by stimulation with epidermal growth factor. The half-life of Ras protein decreased from more than 36 h in control cells to 18 h in RIG1-transfected cells. RIG1 immunoprecipitated with the Ras protein in co-transfected cellular lysates. In contrast to the predominant plasma membrane localization in control cells, the H-Ras fusion protein EGFP-H-Ras was localized within a discrete cytoplasmic compartment where it co-localized with RIG1. RIG1 inhibited more than 93% of the Elk- and CHOP-mediated transactivation induced by H- or K-Ras(G12V). However, RIG1 did not inhibit the transactivation induced by MEK1 or MEK3, and failed to suppress the phosphorylation of extracellular signal-regulated kinases 1 and 2 induced by the constitutively activated B-Raf(V599E). The RIG1 with carboxyl terminal truncation (RIG1DeltaC) did not immunoprecipitate with Ras and had no effect on Ras activation or transactivation of the downstream signal pathways. These data indicate that RIG1 exerts its inhibitory effect at the level of Ras activation, which is independent of Ras subtype but dependent on the membrane localization of the RIG1 protein. This inhibition of Ras activation may be mediated through downregulation of Ras levels and alteration of Ras subcellular distribution. 相似文献
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