• Journal of fish pathology
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• v.21 no.1
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• pp.1-11
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• 2008

We report the existence of new type of phosphatidylcholine-hydrolyzing phospholipase D (PLD), which has been characterized and partially purified in the scuticociliate, Uronema marinum. The enzyme from partial purification showed that it was existed in membrane fraction and was a neutral PLD, which catalyzed both transphosphatidylation and hydrolysis reaction. The activity of partially purified membrane-bound PLD was also found to be optimal at pH 7.0-7.5 for 2 hours at 37℃ and depended strictly on the presence of Ca2+ (2.5 mM) and Mg2+ (1.6 mM). Immunoblot analysis indicated that the enzyme was distinct from hPLD1 (human PLD1) and hPLD2 (human PLD2) because it was not recognized by a polyclonal antibody raised to the 12 terminal amino acid of these enzymes. We also found that the membrane-bound PLD is a PIP2-dependent PLD and that GTP-binding proteins are not implicated in the regulation of this enzyme: This enzyme activity is markedly stimulated by phosphatidylinositol 4,5-bisphosphate (PIP2) but not by the small G-protein Arf and GTPrS. In addition, this enzyme was capable of hydrolyzing phosphatidylcholine (PC) but not phosphatidylethanolamine (PE), implying that PC was a preferred substrate.

• Journal of Soil and Groundwater Environment
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• v.20 no.1
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• pp.56-64
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• 2015

Acid mine drainage occurrence is a serious environmental problem by mining industry, it usually contains high levels of metal ions, such as iron, copper, zinc, aluminum, and manganese, as well as metalloids of which arsenic is generally of the greatest concern. An indigenous plant extract was used to produce calcium carbonate from Canavalia ensiformis as effective biomaterial, and its ability to form the calcium carbonate under stable conditions was compared to that of purified urease. X-ray diffraction and scanning electron microscopy were employed to elucidate the mechanism of calcium carbonate formation from the crude plant extracts. The results revealed that urease in the plant extracts catalyzed the hydrolysis of urea in liquid state cultures and decreased heavy metal amounts in the contaminated soil. The heavy metal amounts were decreased in the leachate from the treated mine soil; 31.7% of As, 65.8% of Mn, 50.6% of Zn, 51.6% of Pb, 45.1% of Cr, and 49.7% of Cu, respectively. The procedure described herein is a simple and beneficial method of calcium carbonate biomineralization without cultivation of microorganisms or further purification of crude extracts. This study suggests that crude plant extracts of Canavalia ensiformis have the potential to be used in place of purified forms of the enzyme during remediation of heavy metal contaminated soil.

• Journal of Microbiology and Biotechnology
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• v.20 no.10
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• pp.1378-1385
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• 2010

A novel agarolytic bacterium, KY-YJ-3, producing extracellular agarase, was isolated from the freshwater sediment of the Sincheon River in Daegu, Korea. On the basis of Gram-staining data, morphology, and phylogenetic analysis of the 16S rDNA sequence, the isolate was identified as Cellvibrio sp. By ammonium sulfate precipitation followed by Toyopearl QAE-550C, Toyopearl HW-55F, and MonoQ column chromatographies, the extracellular agarase in the culture fluid could be purified 120.2-fold with a yield of 8.1%. The specific activity of the purified agarase was 84.2 U/mg. The molecular mass of the purified agarase was 70 kDa as determined by dodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The optimal temperature and pH of the purified agarase were $35^{\circ}C$ and pH 7.0, respectively. The purified agarase failed to hydrolyze the other polysaccharide substrates, including carboxymethyl-cellulose, dextran, soluble starch, pectin, and polygalacturonic acid. Kinetic analysis of the agarose hydrolysis catalyzed by the purified agarase using thin-layer chromatography showed that the main products were neoagarobiose, neoagarotetraose, and neoagarohexaose. These results demonstrated that the newly isolated freshwater agarolytic bacterium KY-YJ-3 was a Cellvibrio sp., and could produce an extracellular ${\beta}$-agarase, which hydrolyzed agarose to yield neoagarobiose, neoagarotetraose, and neoagarohexaose as the main products.

• Bulletin of the Korean Chemical Society
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• v.27 no.1
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• pp.65-70
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• 2006

An amperometric glucose biosensor has been developed based on the use of the nanoporous composite film of sol-gel-derived zirconia and perfluorosulfonated ionomer, Nafion, for the encapsulation of glucose oxidase (GOx) on a platinized glassy carbon electrode. Zirconium isopropoxide (ZrOPr) was used as a sol-gel precursor for the preparation of zirconia/Nafion composite film and the performance of the resulting glucose biosensor was tuned by controlling the water content in the acid-catalyzed hydrolysis of sol-gel stock solution. The presence of Nafion polymer in the sol-gel-derived zirconia in the biosensor resulted in faster response time and higher sensitivity compared to those obtained at the pure zirconia- and pure Nafion-based biosensors. Because of the nanoporous nature of the composite film, the glucose biosensor based on the zirconia/Nafion composite film can reach 95% of steady-state current less than 5 s. In addition, the biosensor responds to glucose linearly in the range of 0.03-15.08 mM with a sensitivity of 3.40 $\mu$A/mM and the detection limit of 0.037 mM (S/N = 3). Moreover, the biosensor exhibited good sensor-to-sensor reproducibility (~5%) and long-term stability (90% of its original activity retained after 4 weeks) when stored in 50 mM phosphate buffer at pH 7 at 4 ${^{\circ}C}$.

• Journal of Yeungnam Medical Science
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• v.7 no.1
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• pp.39-50
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• 1990

The one event on signalling mechanism is the cleavage by adenyl cyclase of ATP into second messenger, cyclic AMP. The other transfer system of inositol metabolism. it is widely recognized that hydrolysis of the minor membrane lipid phosphoinositide bisphosphate($PIP_2$) initiated by occupation of certain receptors and catalyzed by phospholipase C, lead to toe generation of the two intracellular messengers, inositol triphosphate($IP_3$) and diacylglycerol(DG). $IP_3$ is converted to inositol tetrakisphosphate($IP_4$) by $IP_3$ kinase. In the present study, it is that purification of calmodulin is used by phenyl-Sepharose CL-4B chromatography. it's molecular weigh, 17.000 in SDS-polyacrylamide gel electrophoresis. In order to observe the affinity between calmodulin (CaM)-Affigel 15 and $IP_3$ kinase, and isolated $IP_3$ kinase, was applied in CaM-Affigel with $Ca^{2+}$ equilibirum buffer and EGTA equilibirum buffer. We compared with binding and elution effect of $IP_3$ kinase in several condition of buffer. In affinity of binding. $Ca^{2+}$ equilibrium buffer was in the most proper condition. and elution, CaM/$Ca^{2+}$ buffer(CE1 10.36, CE2 12. 76pM/min/mg of protein) was effected much more than EGTA buffer(E2 1.48, E3 2.43pM/min/mg of protein), but CaM/$Ca^{2+}$ stimulate the activity of $IP_3$ kinase. And then, several detergents such as sodium deoxycholate, tween 20. cholic acid, polyethylene glycol, chaps were applied. The 0.2% chaps buffer(E2 23.19, E3 8.05pM/min/mg of protein) was the most effective in elution of $IP_3$ kinase.