• Korean Journal of Microbiology
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• v.39 no.4
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• pp.223-229
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• 2003

The purpose of this work was to perform the characterization of NAD(P)H-nitroreductase isolated from Stenotrophomonas sp. OK-5 capable of degrading 2,4,6-trinitrotoluene (TNT). Initially, NADP(H)-nitroreductase by a series of purification processes including ammonium sulfate precipitation, DEAE-sepharose, andQ-sepharose was prepared. From samples harvested from fraction collector, three different fractions (I, II & III)having the enzyme activity of NAD(P)H-itroreductase were detected. Specific activities of three fractions I, II,and III of NAD(P)H-nitroreductase were determined to approximately 5.06 unit/mg, 4.95 unit/mg and 4.86 unit/mg, and concentrated to 10.5, 9.8, and 8.9-fold compared to crude extract, respectively. Among these three fractions,the fraction I of NAD(P)H-nitroreductase demonstrated the highest specific activity in this experiment. Several factors affecting on the enzyme activity of NAD(P)H-nitroreductase (fractions I, II & III) were investigated.The optimum temperature of all NAD(P)H-nitroreductase (fractions I, II & III) was 30oC, and the optimal pH was approximately 7.5. Metal ions such as Ag+, Cu2+, Hg2+ inhibited approximately 80% enzyme activity of all NAD(P)H-nitroreductase, and the enzyme activities were decreased about 30-40% inhibition in the presence of Mn2+ or Ca2+. However, Fe3+ showed stimulatory effect on the enzyme activity. The molecular weights of NAD(P)H-nitroreductase (fractions I, II & III) were measured about 27 kDa on the SDS-PAGE.

• Korean Journal of Agricultural Science
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• v.22 no.1
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• pp.82-95
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• 1995

These experiments were carried out to isolate SIgA from human and bovine milk. The immunochemical properties of SIgA from human and bovine milk were examined by Gel filtration, DEAE and SDS-PAGE. Double Immunodiffusion, and Immunoelectrophoresis. The results obtained are as follows: 1. Human SIgA was purified from colostrum of Korean women by repeated gel filtration on Sephadex G-200 and Sepharose 6B, but bovine SigA was not cleary purified from bovine colostrum of Holstein cows by anion exchange chromatography using DEAE-Sephadex A-50 and gel filtration on Sephadex G-200 and Sepharose 6B. 2. The immunochemical properties of fractions from gel filtration on the Sephadex G-200 and Sepharose 6B column as assessed by Immunoelectrophoresis and double Immunodiffusion to identify the presence of IgM in first peak fraction, and the presence of pure SIgA in second peak fraction. However, Bovine SigA rich fraction from bovine colostrum of Holstein cows contained a large amount of $IgG_1$-dimer in addition to SIgA. 3. The fragments of reduced bovine colostrum SIgA rich fraction were estimated to have molecular weights of secretory component, heavy chain and light chain (75,000-80,000, 50,000-60,000, 25,000-27,000 daltons) by SDS-PAGE, respectively. Those were similar to the molecular weight of reduced SIgA from human milk.

• Microbiology and Biotechnology Letters
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• v.31 no.4
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• pp.368-376
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• 2003

Cyclodextrin glucanotransferase (CGTase) from Paenibacillus sp. JB-13 was immobilized on various carriers by several immobilization methods such as ionic binding, covalent linkage and ultrafiltration to improve the process performance. The ultrafiltration and covalent linkage with CNBr-activated sepharose 4B were found as the best method for immobilization of CGTase. The ability of CGTase immobilization onto CNBr-activated sepharose 4B was as high as 18,000 units/g resin when the conditions was as follows: contact time 9 hrs at $37^{\circ}C$, pH 6.0, 100 nm and enzyme loading 24,000 units/g resin. The optimum conditions for production of 2-O-$\alpha$-D-Glucopyranosyl L-Ascorbic acid by immobilized CGTase turned out to be: pH 5.0, temperature $37^{\circ}C$, 20% substrate solution containing 8% (w/v) of soluble starch and 12% (w/v) of L-ascorbic acid sodium salt, 100 rpm, far 25 hrs and with 800 units of immobilized CGTase/ml substrate solution. Moreover the CGTase activity could be stably maintained for 8 times of repetitive reactions after removing products by ultrafiltration through YM 10 membrane.

• Journal of Life Science
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• v.17 no.5 s.85
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• pp.625-633
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• 2007

We isolated a new extracellular alginate lyase-producing microorganism, which displayed alginate-depolymerizing activity in plate assays, from coastal soils in Wando, Jeollanam-do, Korea. This alginate-depolymerizing bacterium belonged to the genus Streptomyces and it was named Streptomyces sp. MET 0515. An extracellular alginate lyase(ALY1) secreted by Streptomyces sp. MET 0515, was purified to homogeneity by a combination of acetone precipitation, anion-exchange chromatography (Q-Sepharose and DEAE-Sepharose) and Sephacryl S-200 HR gel filtration chromatography. Its molecular mass was 26 kDa as determined by SDS-PACE analysis. The enzyme had an optimal temperature of $70^{\circ}C$ for its activity, and was most active at pH 7.5. The thermal and pH stability were $0-50^{\circ}C$, and pH 6.0-9.0, respectively. The enzyme activity was stimulated by 1mM $Mn^{2+}$, and inhibited by 1mM $Fe^{3+}$, 1mM EDTA and 1mM $Zn^{2+}$. Preliminary analysis of substrate specificity showed that this alginate lyase had activity on both poly-alpha 1,4-L-guluronate and poly-beta 1,4-D-mannuronate in the alginate molecule.

• Journal of Life Science
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• v.12 no.1
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• pp.87-95
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• 2002

It was observed that the hot water extract of the leaves of Capsicum annuum L., a Korean edible plant, had a potent anti-complementary activity. Crude polysaccharide fraction(CAL-0) was obtained by methanol reflux, ethanol precipitation, dialysis and lyophilization. CAL-0 contained 51.8% of total sugar, 8.2% of uronic acid and 16.8% of protein, and consisted of mainly arabinose, galactose and glucose as neutral sugars and galacturonic acid as uronic acid. The anti-complementary activity of CAL-0 decreased greatly by periodate oxidation, but was not changed by pronase treatment. Also, the anti-complementary activity of CAL-0 was reduced partially in the absence of the $Ca^{2+}$ ion. The crude polysaccharide CAL-0 was found to activate the C3 component both in the presence and in the absence of $Ca^{2+}$ through the crossed-immunoelectrophoresis suggesting that those involved in both classical and alternative complement pathway CAL-0 was further separated to an unabsorbed fraction(CAL-1) and six absorbed fractions(CAL-2longrightarrowCAL-7) on DEAE Sepharose CL-6B ion exchange column. Among them four major fractions in activity and yield were obtained, and consisted mainly of arabinose, galactose and glucose with various molar ratios. The major fraction, CAL-2, was purified to give a high molecular fraction(CAL-2-I) and a low molecular fraction(CAL-2-II) on Sepharose CL-6B column. The anti-complementary activity of CAL-2-I, a molecular weight of about 61,000, was higher than it of CAL-2-II.-II.

• Proceedings of the Korean Society for Food Science of Animal Resources Conference
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• 2001.11a
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• pp.133-133
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• 2001

• Preventive Nutrition and Food Science
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• v.1 no.2
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• pp.244-251
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• 1996

Molecular weight and partial amino acid sequence of the cis, 9-cis, 12-octadecadienoate isomerase(linoleate isomerase) of Butyrivibrio fibrisovens A-38 were determined. Linoleate isomerase was isolated from the bac-teria cultured anaerobically and purified by ultracentrifugation in conjunction with Sepharose 6B column chro-matography, Phenyl sepharose 4B column chromatography and fast performance liquid chromatography (EPLC). The isomerase was single polypeptide with 19KD of molecular weight, when determined by SDS-PAGE. Fourteen amino acids sequence of N-terminal of the linoleate isomerase was N-GEIDKYPRIIKQQ determined by Edman method.

• Journal of Microbiology and Biotechnology
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• v.11 no.4
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• pp.628-635
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• 2001

Recombinant E. coli DH5$\alpha$ harboring a dextransucrase gene (dsrB742) produced an extracellular dextransucrase in a 2% sucrose medium. The enzyme was purified by DEAE-Sepharose and Phenyl-Sepharose column chromatographies upto a 142.97-fold purification with a 11.11% recovery to near homogeneity. The enzyme had a calculated molecular mass of 168.6 kDa, which was in good agreement with the activity band of 170 kDa on a nondenaturing SDS-PAGE. An expression plasmid was constructed by inserting the dsrB742 into a pRSET expression vector. The activity after expression in E. coli BL21(DE3)pLysS increased about 6.7-fold compared to the extracellular dextransucrase from L. mesenteroides B-742CB. The expressed and purified enzyme from the clone showed similar biochemical properties (acceptor reaction, size of active dextransucrase, optimum pH, and temperature) to B-742CB dextransucrase, however, the ability to synthesize ${\alpha}$-(1$\rightarrow$3) branching decreased in comparison to that of L. mesenteroides B-742CB dextransucrase.

• Journal of the Korean Society of Food Science and Nutrition
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• v.27 no.6
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• pp.1117-1124
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• 1998

Chymosin was purified from commercial rennet with DEAE Sepharose CL 6B and immobilized on CeliteTM using glutaraldehyde. Whole casein from fresh raw milk was hydrolyzed by immobilized chymosin and total CMP was isolated by trichloroacetic acid(TCA) and ultrafiltration, and characterized. The amount of chymosin purified from 15g commercial rennet by DEAE Sepharose CL 6B was 0.16g and 18mg of chymosin was immobilized on 1g of CeliteTM by 5% glutaraldehyde. Immobilized chy mosin hydrolyzed most of casein on whole casein within 2 hours to leave para casein and casei nomacropeptide(CMP). The total CMP isolated from 10g of whole casein hydrolyzate by TCA and ultrafiltration was 0.4g and 0.1g, respectively. Results of electrophoresis, amount of sialic acid, com position of amino acid and ratio of A280 to A214 showed that total CMP by TCA was purer and had more CMP without carbohydrate than one by ultrafiltration. CMP isolated from total CMP by 12% TCA precipitation was 50% of total CMP and most of caseinoglycopeptide(CGP) was removed from total CMP, indicating less amount of sialic acid in CMP than in total CMP.

• Biomedical Science Letters
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• v.4 no.1
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• pp.35-42
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• 1998

The effects of CRP purified from human ascites fluid on phagocytic activity of the human macrophage were investigated. CRP was purified using affinity chromatography including absorption on p-diazonium phosphocholine or C-polysaccharide coupled sepharose 4B and gel filtration on hydroxylapatite column chromatography. Macrophage was separated ficoll hypaque gradient density and absorption method, and then was confirmed phagocytic uptake test using latex method. CRP was able either to inhibit or to enhance phagocytic activity of human macrophage against bacteria in vitro. The effects of CRP on phagocytic activity of human macrophage were in time and dose-dependent manners. The additional sequence of reaction mixture against bacteria in vitro shows a threshold stimulus on the activation of phagocytic response upon the CRP.