• Microbiology and Biotechnology Letters
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• v.24 no.2
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• pp.222-226
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• 1996

The enzymatic synthesis of 3, 4-dihydroxyphenyl-L-alanine (L-DOPA) was examined for the effects of the reaction additives such as sodium borate, alcohol, and organic solvents. The enzyme used was tyrosine phenol-lyase of Citrobacter freundii KCTC 2006 produced in Escherichia coli. The amounts of tyrosine phenol-lyase and pyridoxal-5-phosphate were optimized to 2.0 units/ml and 0.1 mM, respectively, for the synthetic reaction. Sodium borate, a substance that forms a complex with pyrocatechol, reduced the enzyme deactivation by pyrocatechol although it seriously inhibited the enzyme activity. Among the organic solvents tested, dimethylsulfoxide, dimethylformamide, and alcohol increased the productivity of the L-DOPA synthesis. In a reaction system with 5% methanol, L-DOPA concentration increased up to 210 mM after 24 hours, and 77.1% of which was separated as precipitates. The L-DOPA was purified to 99.96% by solubilizing and recrystallyzing the precipitates.

• Journal of fish pathology
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• v.21 no.3
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• pp.181-188
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• 2008

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay was used to detect and identify four fish viruses, fish iridovirus, viral hemorrhagic septicaemia virus (VHSV), viral nervous necrosis virus (VNNV), hirame rhabdovirus (HRV). Four viruses were detected by PCR with each specific primers. Identification of iridovirus was achieved by digesting the PCR amplified fragment with a restriction enzyme ApaⅠ. It was possible to distinguish positive from false positive PCR amplicons of VHSV by RFLP of PstⅠ or HindⅢ restriction enzymes. VNNV was identified using RFLP of BamHⅠrestriction enzyme and HRV was identified by XbaⅠ restriction enzyme. This approach can be used for more rapid, simple and specific diagnosis of fish viral diseases.

• Journal of Environmental Health Sciences
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• v.30 no.2
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• pp.79-83
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• 2004

The oxidative degradation of BPA with laccase from Trametes versiclor was conducted in a closed, temperature controlled system containing acetate buffer for pH control. The effects of medium pH, buffer concentration, temperature and mediator on degradation of BPA were investigated. The inactivation of the enzyme by temperature and reaction product was also studied. The optimal pH for BPA degradation showed about 5. Buffer concentration did not affect BPA degradation. On the other hand, the enzyme stability was higher at low concentration buffer(25 mM). Temperature rise increased the degradation rate of BPA up to 45$^{\circ}C$. The valuable mediator of laccase for BPA was ABTS. Elevated temperature and reaction product irreversibly inactivated the enzyme.

• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.4
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• pp.257-262
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• 2003

A straightforward and effective method is presented for immobilizing enzymes on a microchip platform without chemically modifying a micro-channel or technically microfabricating a column reactor and fluid channel network. The proposed method consists of three steps: the reconstitution of a nitrocellulose (NC) membrane on a plane substrate without a channel network, enzyme immobilization on the NC membrane, and the assembly of another substrate with a fabricated channel network. As a result, enzymes can be stably and efficiently immobilized on a microchip. To evaluate the proposed method, two kinds of enzymatic reaction are applied: a sequential two-step reaction by one enzyme, alkaline phosphatase, and a coupled reaction by two enzymes, glucose oxidase and peroxidase, for a glucose assay.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.573-578
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• 2005

Pyrimidine nucleotide N-ribosidase (pyrimidine 5'-nucleotide phosphoribo(deoxyribo)hydrolase/pyrimidine 5'-nucleotide nucleosidase, EC 3.2.2.10) catalyzes the breakdown of pyrimidine 5'-nucleotide into pyrimidine base and ribose(deoxyribo)-5-phosphate. However, detailed characteristics of the enzyme have not yet been reported. The enzyme was purified to homogeneity 327.9-fold with an overall yield of $6.1\%$ from Pseudomonas oleovorans ATCC 8062. The enzyme catalyzed cytidine monophosphate (CMP) and uridine monophosphate (UMP), but not adenosine monophosphate (AMP) and guanosine monophosphate (GMP). The enzyme optimally metabolized CMP at pH 6.0 and UMP at around 8.5, and the optimum temperature for the overall enzyme reaction was found to be $37^{\circ}C$. The $K_m$ values of the enzyme for CMP (at pH 6.0) and UMP (at pH 8.5) were 1.6 mM and 1.1 mM, respectively. AMP, deoxyCMP, and deoxyUMP were very effective inhibitors of the reaction. Double-reciprocal plots obtained in the absence and in the presence of AMP revealed that this inhibitory effect was of the mixed competitive type with respect to the breakdown of CMP and of the noncompetitive type with respect to the breakdown of UMP. In the presence of AMP, the enzyme followed sigmoid kinetics with respect to each substrate.

• Applied Biological Chemistry
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• v.11
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• pp.55-61
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• 1969

This K-17 strain was not absolutely requiring xylose as an inducer for enzyme formations. The most activity of this enzyme was lost when treated at $75^{\circ}C$. for 30 hours but was not influenced at $70^{\circ}C$. for 70 hours of treatment. The activity of this enzyme was increased by the addition of magnesium ions or cobalt ions in the reaction system. In the studies, we found that the magnesium ions simply activate the enzyme reaction and the cobalt ions do not but protect the enzyme from heat inactvation. And it was also found the phosphate buffer solution was very suitable as glucose dissolving solvent on the enzyme reaction. The mixed carbon source medium containing glucose, fructose, sorbitol, xylose and sucrose was more favorable for enzyme production then a sole carbon source containing medium on the shaking culture method.

• Journal of Biosystems Engineering
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• v.32 no.6
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• pp.455-461
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• 2007

The electric current of a flow injection type enzyme-sensor was measured to confirm the stable operating conditions of the sensor. The current of the sensor was decreased as the buffer solution velocity increased. Under the limitation of the cycle time to be below 10 minutes, the effective ranges of the buffer solution velocity were suggested $0.10{\sim}0.26$, $0.12{\sim}0.24$, $0.1{\sim}0.25$ and $0.05{\sim}0.10\;cm/s$ of 1.0, 1.4, 2.4 and 3.4 mm of the electrode diameters, respectively. As the reaction time of the enzyme and the substrate was increased, the current was decreased because of the dilution between the sample and buffer solution. Therefore, it could be recommended that the reaction time was able to be selected as shortly as possible in consideration of the total cycle time. As the result of the experiments using a different volume ratio of the enzyme to substrate, it was concluded that the substrate had to be mixed with the same amount of the enzyme. The current have increased remarkably in proportion to the electrode diameter under 0.1 cm/s of the buffer solution velocity but there was no difference over 0.1 cm/s of the buffer solution velocity. The cross type arrangement of the electrode was highly suggested for application and machining of the sensor.

• Journal of the Korean Society of Food Science and Nutrition
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• v.24 no.6
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• pp.943-947
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• 1995

The sweet potato drinks were prepared with the reaction of sweet potato and complex enzyme(${\beta}-amylase,\;{\alpha}-amylase$, protease). The reducing sugar and soluble solid of sweet potato drinks were the highest on reaction of sweet potato : water(1 : 1) and complex enzyme(pH 4.5). In the color of the sweet potato drinks, hunter value(L, a, b) were the lowest on reaciton of sweet potato : water(1 : 1) and complex enzyme, and were the highest on reaciton of sweet potato : water(1 : 3) and complex enzyme(pH 4.5). In the sensory test of the sweet potato drinks, the sensory score(color, taste, flavor, texture) were the best on reaction of sweet potato : water(1 : 1) and complex enzyme(pH 4.5). These results demonstrated that the sweet potato drink was good to drink when sweet potato : water(1 : 1) were treated with complex enzyme at pH 4.5, $60^{\circ}C$ for 5 hrs.

• Journal of Microbiology and Biotechnology
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• v.12 no.2
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• pp.189-195
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• 2002

Using Streptomyces sp. YU100 isolated from Korean soil, the fermentative production of phospholipase D was attempted along with its purification and characterization studies. When different carbon and nitrogen sources were supplemented in the culture medium, glucose and yeast extract were found to be the best. By varying the concentration of nutrients and calcium carbonate, the optimal culture medium was determined as 2.0% glucose, 1.5% yeast extract, 0.5% tryptone 0.3% calcium carbonate. During cultivation, the strain secreted most of the phospholipase D in the early stage of growth within 24 h. The phospholipase D produced in the culture broth exhibited hydrolytic activity as well as transphosphatidylation activity on lecithin (phosphatidylcholine). In particular, the culture broth showed 8.7 units/ml of hydrolytic activity when cultivated at $28^{\circ}C$ for 1.5 days. The phospholipase D was purified using 80% ammonium sulfate precipitation and DEAE-Sepharose CL-6B column chromatography, which produced a major band of 57 kDa on a 10% SDS-polyacrylamide gel with purity higher than 80%. The enzyme showed an optimal pH of 7 in hydrolytic reaction, and at pH 4 in a transphosphatidylation reaction. The enzyme activity increased until the reaction temperature was elevated to $60^{\circ}C$. The enzyme was relatively stable at high temperatures and neutral pH, but significantly unstable in the alkaline range. Among the detergents tested as emulsifiers of phospholipids, the highest enzyme activity was observed when 1.5% Triton X-100 was employed. However, no inhibitory effect by metal ions was detected. Under optimized reaction conditions, the purified enzyme not only completely decomposed PC to phosphatidic acid within 1 h, but also exhibited higher than 80% conversion rate of PC to PS by transphosphatidylation within 4 h.

• Journal of Microbiology and Biotechnology
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• v.6 no.4
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• pp.238-244
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• 1996

$\gamma$-Glutamyl transpeptidase ($\gamma$-GTP; EC 2.3.2.2) present in the culture filtrate of Bacillus sp. KUN-17 was purified 400-fold through a consecutive procedure including organic precipitation and column chromatography. The enzyme has an estimated molecular weight of 70, 000 and consists of hetero-subunits with molecular weights of 42, 000 and 22, 000. In vitro optimal conditions for those transfer and hydrolysis reactions appeared to be pH 7.0 at $50^{\circ}C$ and pH 8.4 at $40^{\circ}C$, respectively. The denatured enzyme recovered most of its $\gamma$-GTP activity by removing detergents such as sodium dodecyl sulfate (SDS) or urea with dialysis. The enzyme showed higher affinities against a number of amino acids as $\gamma$-glutamyl acceptors than glycylglycine in the following order: L-valine, L-methionine, L-glutamic acid or L-as-paragine, L-alanine. Also, it was shown that L-glutamine was the most suitable $\gamma$-glutamyl donor for the transfer reaction among those tested. Amino acids generally inhibited the enzyme activity for the transfer reaction, but not for the hydrolysis reaction.