The $Ca^{2+}-activated$ neutral protease calpain induced proteolysis has been suggested to play a role in certain cell growth regulatory proteins. Cyclin proteolysis is essential for cell cycle progression. D-type cyclins, which form an assembly with cyclin-dependent kinases (cdk4 and cdk6), are synthesized earlier in G1 of the cell cycle and seem to be induced in response to external signals that promote entry into the cell cycle. Here we show that cyclin D3 protein levels are regulated at the posttranscriptional level by calpain protease. Treatment of human breast carcinoma MDA-MB-231 cells with lovastatin and actinomycin D resulted in a loss of cyclin D3 protein that was completely reversible by the peptide aldehyde calpain inhibitor, LLnL. The specific inhibitor of the 26S proteasome, lactacystin, the lysosome inhibitors, ammonium chloride and chloroquine, and the serine protease inhibitor, phenylmethylsulfonylfluoride (PMSF), did not block the degradation of cyclin D3 by lovastatin and actinomycin D. Results of in vitro degradation of cyclin D3 by purified calpain showed that cyclin D3 protein is degraded in a $Ca^{2+}-dependent$ manner, and the half-life of cyclin D3 protein was dramatically increased in LLnL treated cells. These data suggested that cyclin D3 protein is regulated by the $Ca^{2+}-activated$ protease calpain.
A cold mix asphalt (CMA) treatment process was proposed as a tool to recycle soils contaminated with petroleum hydrocarbons. Experimental studies were conducted to characterize performances of the CMA process in treating soils contaminated with diesel or diesel compounds. From the screening experiments, it was found that performances of five types of asphalt emulsions that contained a cationic or an anionic or a nonionic surfactant were not substantially different. In consideration of higher affinity for soils and higher sorption coefficients obtained, an emulsion containing Lauryl Dimethyl Benzyl Ammonium Chloride (LDBAC) was selected as a promising asphalt emulsion for treating diesel-contaminated soils. When the asphalt emulsion LDBAC was applied to treat three compounds that originated from diesel, the removal efficiencies obtained in the order of decreasing efficiencies were as follows: docosane > pentadecane > undecane. Leaching experiments on the specimen formulated by the emulsion LDBAC found that the selected treatment method could treat soils with diesel concentrations as high as 10,000 mg/kg. Leaching of the diesel from the specimen was controlled by diffusion for the first four days and then leaching rate diminished substantially. The latter behavior was characterized as depletion, which represents that the contaminant released amounts to more than $50\%$ of the total amount of the contaminant that can be leached. The amounts of three diesel compounds leached from the specimen in the order of decreasing amount were undecane, pentadecane, and docosane. The curing of the soil contaminated with pentadecane was relatively slow.
Liquid livestock manure (LLM) has been used as a nitrogen fertilizer source for horticulture plants. LLM contains organic nitrogen (N), ammonium, nitrate, and nitrite. The amount of nitrate and nitrite in LLM are usually small compared to the amount of ammonium in it and so they can be negligible if total nitrogen (N) concentration in LLM is higher than $1,000mg\;L^{-1}$. However, if total N concentration in LLM is less than $1,000mg\;L^{-1}$, the amount of nitrate and nitrite may affect total N concentration in LLM. Currently, Kjeldahl digestion method is mainly used for ammonium-N in LLM. Therefore, it is ineffective to analyze nitrate-N and nitrite-N. The objective of this study was to evaluate whether the total N concentrations are affected by the amount of nitrate-N and nitrite-N with diverse LLMs by Kjeldahl method (with and without Devarda's alloy after Conc. sulfuric acid digestion). Five liquid livestock manure samples were collected at swine farms in Ansung and Icheon. All LLM samples were stored at $25^{\circ}C$, subsampled at every $15^{th}$ day for 90 days, and analyzed for total N, ammonium-N, and nitrate-N. At the $90^{th}$ day, LLM samples were analyzed with and without Devarda's alloy after Conc. sulfuric acid digestion. Potassium nitrate, ammonium nitrate, and ammonium chloride were used to determine the N recovery percentages. Total N concentration ranged from 560 to $4,230mg\;L^{-1}$. Nitrate-Ns were found in all LLM samples, ranged from 21 to $164mg\;L^{-1}$. N recovery percentages with potassium nitrate were 0 % without Devarda's alloy and 100% with Devarda's alloy because adding Devarda's alloy facilitated nitrate-N into ammonium-N conversion. Total Ns were significantly different between two methods, with and without Devarda's alloy. Total N concentrations were $210mg\;L^{-1}$ at LLM 4 and $370mg\;L^{-1}$ at LLM 5 without Devarda's alloy and $290mg\;L^{-1}$ at LLM 4 and $490mg\;L^{-1}$ at LLM 5 with Devarda's alloy. These results suggest that if total N of LLM is less $1,000mg\;L^{-1}$, additional procedure such as adding Devarda's alloy can be used to estimate the total N and inorganic N better.
Kim, Bieong-Kil;Doh, Kyung-Oh;Bae, Yun-Ui;Seu, Young-Bae
Journal of Microbiology and Biotechnology
/
v.21
no.1
/
pp.93-99
/
2011
Amongst a number of potential nonviral vectors, cationic liposomes have been actively researched, with both gemini surfactants and bola amphiphiles reported as being in possession of good structures in terms of cell viability and in vitro transfection. In this study, a cholesterol-based diquaternary ammonium gemini surfactant (Chol-GS) was synthesized and assessed as a novel nonviral gene vector. Chol-GS was synthesized from cholesterol by way of four reaction steps. The optimal efficiency was found to be at a weight ratio of 1:4 of lipid:DOPE (1,2-dioleoyl-L-${\alpha}$- glycero-3-phosphatidylethanolamine), and at a ratio of between 10:1~15:1 of liposome:DNA. The transfection efficiency was compared with commercial liposomes and with Lipofectamine, 1,2-dimyristyloxypropyl-3-dimethylhydroxyethylammonium bromide (DMRIE-C), and N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTAP). The results indicate that the efficiency of Chol-GS is greater than that of all the tested commercial liposomes in COS7 and Huh7 cells, and higher than DOTAP and Lipofectamine in A549 cells. Confirmation of these findings was observed through the use of green fluorescent protein expression. Chol-GS exhibited a moderate level of cytotoxicity, at optimum concentrations for efficient transfection, indicating cell viability. Hence, the newly synthesized Chol-GS liposome has the potential of being an excellent nonviral vector for gene delivery.
Approximately twelve hundred strains of Actonomycetes isolated from domestic soli were tested for their ability to produce extracellular phytase. Of all these isolates a strain, YB-26, that had the highest potential for phytase activity was chosen. The nucleotide sequence of 16S rDNA of the isolate YB-26 showed the highest similarity to that of strains beloning to genus Streptomyces. The partially purified extracellular phytase was obtained from the culture filtrate of Streptomyces sp. YB-26 grown on GSM broth by ammonium sulfate precipitation (15-70%), DEAE-Sepharose column and Q-Sepharose column chromatography. The partially purified enzyme showed the maximum activity for hydrolysis of phyate at $60^{\circ}C$ and pH 7.0, and retained 90% of its maximum activity at the range of pH $6.0{\sim}8.0$. It was thermolabile and its thermostability did not increase in the presence of calcium chloride.
Quantitative analytical conditions for chromium using solvent extraction followed by atomic absorption spectrometry was studied. Trioctylamine(TOA) in tertiary amine or Trioctylmethylammoniumchloride(TOMAC) in quaternary ammonium salt, both containing octyl group was used as an anion exchangers. Absorbance were measured for the different kinds of acid added and as changing the concentration of acid by graphite furnace atomic absorption spectrometer. The maximum absorbance was obtained at the concentrations of HCl, 0.1M to 0.3M for TOA and 0.03M to 0.1M for TOMAC. Mole ratios over 1:1 of TOA or TOMAC dissolved in MIBK solution to chromium in sample shows optimum extraction efficiency while HCl was added to the MIBK. As a result of scrutinizing the extraction process, the methods employed in this experiment turned out to be better extraction efficiency for chromium, compared to similar extraction methods.
The optimum conditions for the analysis of the difenoconazole fungicide on soil and crops were investigated and the residues of that in apple and soil were identified by using the gas chromatography. The extract with acetonitrile was separated with saturated NaCl and n-hexane solution after filtered, and concentrated. Obtained fungicide residues were transfered to the florisil column and eluted with acetone and n-hexane mixed solution for the analysis by GLC(ECD). From the standard addition experiments with 0.20 and 1.0ppm, the average recoveries were 86~92% and the detection limit was 0.01 ppm. It seems to be safely used when difenoconazole is treated three times until 15 days before harvest of apple. In this case residual amounts of difenoconazole in apple was from 0.037ppm to 0.044ppm. The soil samples extracted with methanol and ammonium hydroxide mixed solution were partitioned with dichloromethane and saturated sodium chloride solution. The organic phase was concentrated and redissolved with toluene and analyzed with GLC(FID) after cleaned with Sep-Pak column. From the standard addition experiments with 0.10, 0.50 and 1.0ppm, the average recoveries were 101.2~103.7% and the detection limit was 0.025ppm.
To establish an agar-gel immunodiffusion (AGID) test for detection of antibodies to Aujeszky's disease virus(ADV) in swine, the precipitating antigens were prepared by four procedures using the Aujeszky's disease virus, NYJ-1-87 strain isolated from the affected piglets in Korea. The optimal condition for AGID test and the properties of the antigens were investigated. To determine the optimal concentration of antigens, four antigens were experimentally prepared by concentrating the viral fluids by 1/30 to 1/200. It was proved that the antigen precipitated with ammonium sulfate at concentration of 1/100 was the most efficient to detect ADV antibodies by AGID test. When the relationship between the concentration of the antigens and the size of precipitating in radial immunodiffusion test was investigated, a high correlation coefficiency at r=0.95 (y=0.23x+23.4) was estimated, In study on the effects of various buffered salt solutions and agars on the sensitivity of AGID test by using the experimental ADV antigens, it was found that 0.05M tris buffer without sodium chloride at pH 7.2 induced the most distinctive precipitating lines, and that there was no significant differences in the sensitivity between the agarose and Noble's special agar. When the efficiency of AGID test was compared with serum neutralization(SN) test, the sensitivity of AGID test was 100% in SN titer over 1 : 16, 91.7% in SN titer of 1 : 8 and 57.1% in SN titer of 1 : 4. The specificity of AGID test compared with the sera with SN titer under 1 : 2 was 98.4%. Protein analysis of the antigens by SDS-PAGE indicated that antigen I and antigen III showed a specific band of polypeptides with molecular weight of 116 K in comparison with the control antigen. Antigen IV, treated with tween-80 and ammonium sulfate, revealed specific polypeptides bands at the molecular weights 45K, 98K and 150 K.
Journal of Korea Technical Association of The Pulp and Paper Industry
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v.31
no.4
/
pp.58-68
/
1999
This experiment was to investigate decoloization characteristics of E1 effluents from the bleaching plant of pulp mill with three white-rot fungi(Trametes versicolor, Ganoderma appanatum and Pleurotus ostreatus).In addition, the effect of carbon and nitrogen resources was discussed on its decolorization. The color removal of E1 effluent during shaking and stationary cultures were 72% and 80%, respectively. Stationary culture was more effective on decolorization of E1 effluent compared to the shaking culture. The optimum inoculum weight was 1.0g based on dry weight of mycelia . The decolorization medium I showed 88% of the color removal of E1 effluent with in one day cultivation of T.versicolor and P.ostreatus . Color removal was increased from 87% to 90%. T.versicolor and P.ostreatus by the addition of 0.5% glucose. By addition of nitorgen sources(ammonium sulfate and ammonium choride), medium was much higher than that of carbon source. With 0.1% ammoniumm sulfate, P.ostreatus and T.versicolor showed 94% and 92% of the color removal within one day of cultivation , respectively. On decolorization medium II, T.versicolor and P.ostretus were 94% of oclor removal with addition of carbon source. The addition of nitrogen source was much more efficient than that of carbon source. With 0.1% amminium chloride, T.versicolor and P.ostreatus showed 95% of its color removal . The decolorization medium II was higher color removal than medium I, and also MnP and laccase were produced. However, the decolorization medium I produced a little MnP and laccase activity. It could be suggested that MnP and laccase may play an important role in decolorization of E1 effluent.
We examined some properties of yeast cell wall lytic enzyme produced by Dicyma sp. YCH-37. Several metal ions, reducing reagents, and chemical modifiers have little effects on the lytic activity, except guanidine-HCl. Yeast cells of early log phase were more susceptible to the enzyme than those of stationary phase, and heat-treated cells were more easily lysed than intact living ones. Yeast cells pretreated with organic solvents such as butanol and acetone were more susceptible to the enzyme than intact living ones. Yeast cells cultured in Yeast extract-Malt extract medium containing 0.5 M ammonium sulfate were easily lysed by the lytic enzyme, and yeast cells cultured without shaking were more easily lysed by the enzyme than those with shaking. When SDS, ${\beta}-mercaptoethanol$, Triton X-100, sodium sulfite, and KCl were added to enzyme reaction mixture each, lysis of yeast cells was more effective.
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