• Korean Journal of Food Science and Technology
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• v.30 no.5
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• pp.1169-1178
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• 1998

This study was performed to purify fig pectinesterase(F-PE) and characterize its in situ activity. Three kinds of F-PE were partially separated by using ammonium sulfate fractionation, Q-Sepharose column, CM-cation exchanger column chromatography, and HPLC. One of those was anionic protein and the others were cationic proteins. All of them had approximate molecular weight of 27,000 and lost rapidly their activity during storage. Therefore alternative crude enzyme was prepared by suspending the freeze dried and milled fig powder in 0.1 M NaCl at pH 7.5. F-PE had the optimum pH of 8.5, the optimum temperature of $50^{\circ}C$ with activation energy of 7,671 cal $mol^{-1}K^{-1}$ and stability up to $55^{\circ}C$ with 10 minutes heating. Optimum activity was obtained in $0.2{\sim}0.4$ M NaCl with optimum solubility at above 0.8 M NaCl.

• Korean Journal of Food Science and Technology
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• v.34 no.2
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• pp.343-345
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• 2002

The study was investigated hot-water soluble on waxy black and waxy rice flours according to gelatinization temperatures and heating time. The hot-water soluble contents of both samples were increased during heating time at $90^{\circ}C$. The waxy black and waxy rice flours solutions according to gelatinization temperatures ($80^{\circ}C,\;90^{\circ}C,\;95^{\circ}C\;and\;98^{\circ}C$) were heated for 10, 20 and 30 min. Determination of elution patterns of rice flours was used by Sepharose CL-2B column. As a result, the elution patterns were eluted the most in void volume ($V_0$) and the elution patterns of the hot-water solubles were increased according to heating time and gelatinization temperatures.

• Journal of Microbiology
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• v.39 no.4
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• pp.279-285
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• 2001

Vibrio parahaemolyticus is a halophilic bacterium associated with seafood gastroenteritis. An unusual strain of Kanagawa-positive urease producing Vibrio parahaemolyticus O1:K1 was isolated from the environment and identified . A polymerase chain reaction assay revealed that this strain harbored both the tdh and the genes. The urease from this strain was studied. Maximum urease production was induced in LB medium containing 0.2% urea, 0.5% glucose, 2% NaCl and pH 5.5 with 6h of culti-vation at 37$\^{C}$ under aeration. Purification of urease was achieved by the process of whole cell lysate, 65% ammonium sulphate precipitation, DEAE-cellulose ion exchange column chromatography, Sepharose CL-6B gel filtration and oxirane activated Sepharose 6B-urea affinity chromatography with 203 fold purification and 2.2% yield. Analysis of the purified enzyme by SDS-PAGE demonstrated the presence of the subunits with a molecular weight of 85kDa, 59kDa, 41kDa and the molecular weight for the native enzyme by nondenaturing PAGE and gel filtration chromatography was 255kDa. The purified urease was stable at pH 7.5 and the opeimal pH in HEPES buffer was 8.0 The enzyme was stable at 60$\^{C}$ for 2 h with a residual activity of 32% . The addition of 10$\mu$M if NiCl$_2$maintained stability for 30 min. The Km value of the purified enzyme was 35.6 mM in urea substrate. The TD$\_$50/(median toxic dose) of the purified urease was 2.5$\mu\textrm{g}$/ml on human leukemia cells.

• Korean Journal of Microbiology
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• v.42 no.4
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• pp.294-298
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• 2006

An immunoaffinity chromatography for the specific binding of Streptomyces somaliensis phospholipase D (PLD) that is considered as an industrially potential enzyme was developed. By using the protein structure prediction programs and the X-ray crystal structure of a Streptomyces PLD, 5 different epitopes with high antigenicity that are predicted to locate on the surface of the S. somaliensis PLD were selected and then synthesized for the preparation of antipeptide antibodies. Each purified rabbit IgG was coupled with NHS-activated Sepharose to prepare the immunoaffinity resins. After one-step purification of the culture concentrate on the antipeptide IgG-coupled Sepharose column, SDS-PAGE and the Western blot analysis of the purified samples showed that purification of PLD on the affinity columns was satisfactory, indicating that the peptide design using the structural information of Streptomyces PLDs was rational. However, the purified PLD in the solution aggregated rapidly, which resulted in poor specific activity and low purification yield.

• Journal of fish pathology
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• v.17 no.2
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• pp.131-137
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• 2004

In the present study, phosphatidylcholine (PC) hydrolyzing enzyme had been isolated from membrane of flounder liver. PC hydrolyzing enzyme solubilized in 1% Triton X -100 from membrane was partially purified by sequential chromatography on Heparin Sepharose CL-6B and Heparin-5PW columns. The products by membrane-bound hydrolyzing enzyme were identified as phosphatidic acid and choline, but in the presence of primary alcohol, phosphatidylethanol was produced at the expense of phosphatidic acid. These data suggest that membrane-bound enzyme may be a PC-phosphoipase D (PLD) type. The enzyme had pH optimum at below 6.0 and temperature optimum at $37^\circ{C}$. The activity of PC-PLD was dose-dependently increased by $Ca^{2+}$ but not $Mg^{2+}$. The activity of PC-PLD was stimulate by PC, PIP2 and PE.

• Journal of Microbiology and Biotechnology
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• v.15 no.2
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• pp.395-402
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• 2005

Biologically active recombinant human follicle stimulating hormone (rhFSH) was produced in Chinese hamster ovary cells and purified by a series of chromatographic steps. The chromatographic steps included anion-exchange chromatography (DEAE Sepharose F/F, Q Sepharose F/F), hydrophobic interaction chromatography (Source 15 PHE), and hydroxyapatite chromatography (Macro-Prep ceramic hydroxyapatite type I). A distinctive step of the purification process developed was the use of ZnCl$_2$ for the removal of non-glycosylated or lowly-glycosylated FSH and impurities through co-precipitation with Zn$^{2+}$. Purified rhFSH was identified and characterized by several physicochemical and biological methods such as gel electrophoresis, high-performance liquid chromatography, amino acid analysis, carbohydrate analysis, and biological activity. The overall yield of the purification was ~$30\%$. The rhFSH preparation obtained showed high purity (>$99\%$) and high in vivo potency (>16,000 IU/mg). Carbohydrate analysis suggested that the purified rhFSH contained approximately $40\%$ (w/w) carbohydrate with di­or tri-antennary structure on average, which is somewhat more heavily sialylated than commercially available rhFSH. In conclusion, the results of these analyses established an identity of the purified rhFSH with natural FSH from human pituitary glands, and furthermore, the purified rhFSH preparation showed higher in vivo potency and was slightly more heavily sialylated than commercially available rhFSH.

• Journal of Food Hygiene and Safety
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• v.18 no.3
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• pp.146-152
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• 2003

Pyruvate dehydrogenase phosphatase (PDP) is a mitochondrial protein serine/threonine phosphatase that catalyzes the dephosphorylation and concomitant reactivation of the pyruvate dehydrogenase component of the pyruvate dehydrogenase complex (PDC). PDP consists of a catalytic subunit (PDPc, Mr 52,600) and regulatory subunit (PDPr, Mr 95,600). In the presence of $Ca^{2+}$, PDPc binds to the dihydrolipoamide acetyltransferase (E2) component of the pyruvate dehydrogenase complex in proximity to its substrate, the phosphorylated E1 component, thereby increasing the rate of dephosphorylation. PDPc possesses and intrinsic $Ca^{2+}$ binding site and a second $Ca^{2+}$ site is generated in the presence of E2. Using the unique interaction, highly pure PDPc was produced by the GSH-Sepharose-GST-L2 matrix with a specific activity of approx. 1000 U/mg and a yield of about 80%.

• Journal of Life Science
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• v.20 no.11
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• pp.1582-1588
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• 2010

A new alginate lyase gene of marine bacterium Streptomyces sp. M3 had been previously cloned in pColdI vector and transformed into E. coli BL21 (DE3). In this study, M3 lyase protein without signal peptide was overexpressed by induction with IPTG and purified with Ni-Sepharose affinity chromatography. The absorbance at 235 nm of the reaction mixture and TLC analysis showed that M3 alginate lyase was a polyG-specific lyase. When M3 lyase was assayed with substrate for 10 min, optimum pH and optimum temperature were pH 9 and $60^{\circ}C$. For the effect of 1mM metal ion on M3 lyase activity, $Ca^{++}$ and $Mn^{++}$ ions increased the alginate degrading activity by two-fold, whereas $Hg^{++}$ and $Zn^{++}$ ions inhibited the lyase activity completely. $Mg^{++}$, $Co^{++}$, $Na^+$, $K^+$, and $Ba^{++}$ did not show any strong effects on alginate lyase activity.

• The Journal of the Korea Contents Association
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• v.12 no.9
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• pp.244-249
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• 2012

This study was carried out to investigate effect of serum lipid and liver tissue in lectin from Viscum album on rats. The lectin was purified by sepharose 4B affinity chromatography and gel filtration using sephadex G-150 with plant material from Viscum album. After 72 h of $CCl_4$ injection, there was a significant increase in serum total cholesterol and triglycerige levels relative to the control group. However, treatment of both Viscum album and purified lectin were significantly decreased lipid parameters against the $CCl_4$-induced. Histological observation of the liver section of rats challenged with $CCl_4$ produced a marked increase of cytoplasmic vacuoles in number, while rats administered olive oil alone did not alter the normal hepatic architecture. Histological observation of the liver section in rats treated 72 h with either Viscum album purified lectin or $CCl_4$-induced liver lipogenesis showed decreased numbers of cytoplasmic vaculoes and necrotic cells. These results suggest that Viscum album lectin has a significantly decreased lipid in serum or histological observation of the liver section decreased in lipogenesis in rats.

• KSBB Journal
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• v.20 no.5 s.94
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• pp.355-358
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• 2005

Bacteroides is the most predominant bacteria in the human large intestinal tract. Bacteroides fragilis RoidB isolated from a Korean faecal sample showed high $\alpha-fucosidase$ activity compared to other bacterial species. The optimized meidum for the production of $\alpha-fucosidase$ from 8. fragilis contained BHI 37g, hemin 10mg, cysteine 0.5g, resazurin 1mg, vitamin K1mg, and starch 5g per 1 L. The crude $\alpha-fucosidase$ obtained through DEAE-sepharose and CM- cellulose chromatography showed optimum temperature and pH at $40^{\circ}C$ and 7.0, respectively. Among several metals, $Co^{++}\;and\;Zn^{++}$ showed strong inhibition on enzyme activity.