• Biomedical Science Letters
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• v.4 no.1
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• pp.1-9
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• 1998

A Corynebacterium diphtheriae iron-repressible gene dirA, that was homologous to TSA of Saccharomyces cerevisiae and AhpC subunit of Salmonella typhimurium alkyl hydroperoxide reductase, was amplified with PCR and expressed in E. coli. The DirA purified from the transformed E. coli crude extracts prevented the inactivation of enzyme caused by metal-catalyzed oxidation (MCO) system containing thiols but not by ascorbate/Fe$^{3+}$/$O_2$ MCO system. The DirA concentration, which inhibited the inactivation of glutamine synthetase by 50% (IC$_{50}$) against MCO system, was 0.12 mg/ml. The multimeric forms of DirA were converted to the monomeric form in SDS-PAGE under the thioredoxin system comprised of NADPH, Saccharomyces cerevisiae thioredoxin reductase, and thioredoxin. Also, DirA showed thioredoxin dependent peroxidase activity. All of these results were consistent with the characteristics of a thiol specific antioxidant (TSA) protein having two conserved cysteine residues.

• Korean Journal of Microbiology
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• v.40 no.2
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• pp.81-86
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• 2004

Among the annotated ORFs of Streptomyces coelicolor, SCO7697 was supposed to encode for phytase (myo-inositol hexakisphosphate phosphohydrolase). The DNA fragment containing SCO7697 was cloned by the PCR from the chromosomal DNA of S.coelicolor A3(2)M. The cloned fragment was introduced into E. coli expres-sion vector, pET28a(+), to yield two recombinant plasmids, pET28-SP and pET28-LP, which were designed to encode different length of proteins. When the pET28-SP and pET28-LP were introduced into E. coli BL21, the transformants successfully overexpressed recombinant proteins, but the molecular weights of the expressed pro-teins were appeared bigger than those of expected in SDS-polyacrylamide gel electrophoresis. The shift of cul-tural temperature from 37 to $30^{\circ}C$ made most of expressed protein be solubilized. The expressed protein, however, did not show any phytase activity. When the DNA fragment with its own promoter placed on the E. coli-Streptomyces vector, pWHM3, and introduced into S. lividans, the phytase activity was not detected either. These results suggest that even though the SCO7697 was annotated as a probable phytase with high probability (E value is $6e^{-89}$), the real product doest not have phytase activity.

• Microbiology and Biotechnology Letters
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• v.39 no.4
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• pp.337-344
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• 2011

Integrase MJ1 from the bacteriophage ${\Phi}FC1$ carries out recombination between two DNA sequences (the phage attachment site, attP and the bacterial attachment site, attB) in NIH3T3 mouse cells. In this study, the integration vector containing attP, attB and the integrase gene MJ, was constructed. The integration mediated by integrase MJ1 in Escherichia coli led to excision of LacZ. Therefore, the frequency of integration was measured by the counting of the white colony, which is detectable on X-Gal plates. The extrachromosomal integration in NIH3T3 mouse cells was monitored by the expression of the green fluorescent protein (GFP) as a reporter. To demonstrate integration mediated integrase MJ1 in NIH3T3 cells, vectors containing attP and attB were co-transfected into NIH3T3 cells. The integration was confirmed by fluorescent microscopy. The expression of GFP was induced in NIH3T3 cells expressing MJ1 without accessory factors. By contrast, the excision mediated by the MJ1 between attR and attL had no effect on the expression of GFP. These results suggest that integrase MJ1 may enable a variety of genomic modifications for research and therapeutic purposes in higher living cells.

• Microbiology and Biotechnology Letters
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• v.32 no.1
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• pp.67-71
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• 2004

Pata de Vaca (Bauhinia forficata) Is a tree which grows naturally in the rainforests and tropical parts of Peru and Brazil, as well as tropical zones of Asia, eastern Paraguay and northeastern Argentina. The active fraction (Pata-50) of the 70% ethanol extract from Pata de Vaca was sequentially fractionated by HP-20 Diaion column chromatography and C-18 column chromatography, and its characteristics were investigated. The growth of all cancer cells tested except for MCF-7 was Inhibited in a concentration-dependent manner by Pata-50. Its $IC_{50}$ values were estimated to be 40.4 $\mu\textrm{g}$/$m\ell$ on AGS, 51.3 $\mu\textrm{g}$/$m\ell$ on HT-29, 52.1$\mu\textrm{g}$/$m\ell$ on HepG2, 65.2$\mu\textrm{g}$/$m\ell$ on A549, and 77.5$\mu\textrm{g}$/$m\ell$ on HeLa cells. A flow cytometric analysis of HepG2 cells revealed induction of apoptosis, but cell cycle regulation was not affected. The HepG2 cell population of apoptosis region increased In a concentration-dependent manner by Pata-50.

• Microbiology and Biotechnology Letters
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• v.30 no.3
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• pp.235-240
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• 2002

Streptomycetes is a group of Gram-positive soil bacteria that growas a branching vegetative mycelium leading to the formation of spores, and display a physiological differenti-ation related to the synthesis of many secondary metabolites including antibiotics. Their complex life cycle and multicellular differentiation require various levels of regulation and types of signal transduction systems including eukaryotic-type serine/threonine protein kinases and prokaryotic-type histidine/aspartic acid protein kinases. Akt kinase that was found in cells is a sorine/threonine kinase controlling signal pathway for multi-tude of important cellular events. The activation or inactivation of Akt kinase in the cell is one of the critical regulatory points to deliver cell proliferation, differentiation, survival or apoptosis signal. To find the regula-tory protein homologous to Akt in Streptomyces, the fluorescien-labeled synthetic peptide (FITC-TRRSR-TESIT) was designed from the consensus sequence of target proteins for Akt kinase. From the difference of the mobility between the nonphosphorylated and phosphorylated synthetic peptides on Agarose gel electro-phoresis, the Akt-phosphorylating activity was monitored. The cell-free extract prepared from Streptomyces griseus IFO 13350 and the Akt homologous protein was purified by ammonium sulfate fractionation and many steps of column chromatographies such as, DEAE-Sepharose, Mono Q, Resource Phenyl-Soporose and Gel permeation column chromatographies. As a result, the protein phosphorylating the fluorescien-labeled Akt substrate was identified and it's molecular weight was estimated as 39 kDa on SDS-PAGE.

• Microbiology and Biotechnology Letters
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• v.17 no.6
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• pp.580-586
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• 1989

Penicillium sp. CB-20 was selected for strong polygalacturonase activity among various strains of molds found in soil. The optimum pH for the enzyme activity was 5.0 and optimum temperature was 4$0^{\circ}C$. The enzyme was relatively stable in acidic condition and unstable by heat treatment. The activation energy, Km and V$_{max}$ for the polygalacturonase were 2.499 Kcal/mol, 2.13$\times$10$^{-2}$mol/l, and 104.17 $\mu$mol/min. The activity of polygalacturonase was inhibited by Ag$^{+}$, Cu$^{++}$, Pb$^{++}$, Fe$^{+++}$, $Ca^{++}$, Na$^+$, Mn$^{++}$. The enzyme can be inactivated by the treatment ethylenediamintetra acetic acid, 2,4-dinitrophenol and $H_2O$$_2$. The results indicate the possible involvement of histidine, chelate and terminal amino group as active site. The enzyme was endo-type polygalacturonase.

• Korean Journal of Microbiology
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• v.30 no.6
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• pp.558-563
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• 1992

Chlorobium liimicola f. thiosulfatophilum NCIB 8327 was grown on modified Pfennig's medium using ammonium chloride. glutamine. glutamate, or dinitrogen gas as nitrogen sources. Except for the case of dinitrogen gas. the extent of gro\\1h was almost the s~me. The specific activity of glutamine synthetase in crude extracts is the highest in the cells which were grown on the medium containing glutamate. hut that of glutamate synthase is uniform for all four nitrogen sources. When the concentration of ammonium ions increases in the reaction mixture. the specific activity of glutamine synthetase in crude extract from the cells grown on glutamate decreases. hut that of glutamate dehydrogenase increases. whereas that of glutamate synthase remains unchanged. When the concentration of methionine sulfoximine increases, the activity of glutamine synthetases decreases rapidly. On the other hand. when the concentration of ammonium ions increases in the reaction mixture gradually. the activity of glutamine synthetase from the cells grown on higher concentration of ammonium ions less decreases. In the presence of light. the activity of glutamine synthetase increases. hut in the dark it decreases gradually. The production of hydrogen in intact cells depends on light. It is inhihited by adding ammonium ions. hut restores immediately hy adding methionine sulfoximine. The produclion of hydrogen in this strain can he mediated by nitrogenase only. and regulated hy glutamine synthetase.

• Ecology and Resilient Infrastructure
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• v.5 no.3
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• pp.111-117
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• 2018

Disinfection of ballast water using chlorine dioxide was investigated under various initial microorganism contents, dose concentrations and pH values. Kinetics of microorganism inactivation and byproduct generation of chlorine dioxide treatment were compared with the chlorine treatment. Results of treatments with chlorine dioxide concentrations of 0 to $10mg\;Cl_2/L$ showed that The optimum concentration of chlorine dioxide required for disinfection of ballast water was 1 mg/L. The difference among the second order reaction constants for bacterial disinfection at pH 7.2 to 9.2 for chlorine dioxide was less than 5% for both bacteria. This result implied that the bactericidal effects of chlorine dioxide was independent of the pH in the examined range. On the other hand, the inactivation kinetics of chlorine for E. coli and Enterococcus decreased by 17% and 25%, respectively, when pH increased from 7.2 to 9.2. The bactericidal power of chlorine dioxide was superior to sodium hypochlorite above pH 8.2, the average pH value of sea water. Furthermore, treatments of chlorine dioxide generated less harmful byproducts than chlorine and had a long-term disinfection effect on bacteria and phytoplankton from the results of experiment for 30 days. Chlorine dioxide would be a promising alternative disinfectant for ballast water.

• Microbiology and Biotechnology Letters
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• v.16 no.4
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• pp.310-315
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• 1988

Two isoenzymes of chorismate mutase(E.C.5.4.99.5) designated as chorismate mutase I(CM I) and chorismate mutase II(CM II), were detected and partially purified from a sp. of intrasporangium isolated from soil. CM I and CM II had pH optima of pH 6.5 and 8.0, respectively and showed the same temperature optimum of 45$^{\circ}C$. The activation energy of the enzymatic reaction was estimated to be 14.7kcal/ mole with CM I and 10.8kcal/mole with CM II. The affinity of isoenzyme CM I for substrate(Km= 1.35mM) was almost the same level as that of CM II(Km = 1.22mM). Both isoenzymes were stable at pH values ranged from pH 6.5 to 9.0, but rapidly denaturated at temperatures above 45$^{\circ}C$. CM II was activated about 7$^{\circ}C$ of its activity by $Ba^{++}$ or $Mg^{++}$ while CM I was slightly inhibited by the same metal ions. Thiol compounds were found not to be necessary for stability of the two enzymes but Co$^{++}$ and EDTA had a little stabilizing effect on CM II only. p-Chloromercuribenzoate strongly inactivated the activities of both enzymes but the reducing agents such as dithiothreitol and L-cysteine protected them against the pCMB inhibition.

• Korean Journal of Food Science and Technology
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• v.33 no.2
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• pp.226-230
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• 2001

High hydrostatic pressure was applied to Foxtail Millet Takju to investigate the effects of high pressure on inactivation of microorganisms and enzymes. Total bacteria, lactic acid bacteria and yeast in untreated Takju were $6.8{\times}10^7,\;1.3{\times}10^8\;and\;8.4{\times}10^7\;CFU/mL$, respectively. Total bacterial count in Takju reduced to $2.2{\times}10^5\;CFU/mL$, while lactic acid bacteria and yeast were sterilized completely when heated at $65^{\circ}C$ for 30 min. Lactic acid bacteria and yeast decreased with the increase of treatment pressure, and pressurization of 400 MPa for 10 min at room temperature sterilized completely the lactic acid bacteria and yeast in Takju. Total bacteria were not sterilized with pressurization of even 600 MPa at room temperature. Total bacteria were completely sterilized at $66^{\circ}C/400\;MPa/60\;min\;and\;66^{\circ}C/600\;MPa/10\;min$. Pressurization of Takju caused a partial inactivation of ${\alpha}-amylase$, and after pressurization at 600 MPa for 10 min at room temperature, 73.2% of the original activity remained. The activity of glucoamylase increased with the increase of treatment pressure. Treatment at $66^{\circ}C/400\;MPa/10\;min$ reduced the activity of ${\alpha}-amylase$ by 59.7% and glucoamylase by 20.5%. ${\alpha}-Amylase$ was inactivated to less than 1.2% of the original activity at $66^{\circ}C/600\;MPa/30\;min$.