• Microbiology and Biotechnology Letters
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• v.24 no.6
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• pp.678-686
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• 1996

To improve the preservation of Kimchi, we isolated Lactobacillus plantarum lytic enzyme-producing strain from soil, and the enzyme was purified and characterized. From the observation of cultural and morpho- logical characteristics, the isolated strain was identified as Metarrisium anisopliae (Metschn.) Sorok. The enzyme was purified to 75-folds with 40% yields through affinity adsorption and CM-Sephadex C-50 column chromatog- raphy. The optimum pH and temperature for lytic activity are 4.0 and 40$\circ$C, respectively, and the enzyme acitvity is stable between pH 3.0 and 9.0, and up to 50$\circ$C. The enzyme is a monomeric protein with molecular weight of 40,000 daltons by SDS-PAGE and gel filtration. The enzyme is endopeptidase which breaks the peptide linkage of Lactobacillus plantarum peptidoglycan. The lytic action spectra confirmed that Leuconostoc mesenteroides, a useful strain for the fermentation of Kimchi, is not lysed by the enzyme. The enzyme activity is inhibited by N-bromosuccinimide (NBS), which probably indicates the involvement of tryptophan residue in active site of the enzyme, and also inhibited by Ag$^{+}$. The amino acid composition analysis showed that the enzyme contains more acidic amino acids than basic ones, and composition of alanine, glycine, proline and tyrosines was very high.

• Journal of Microbiology and Biotechnology
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• v.13 no.5
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• pp.738-744
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• 2003

The gene ADH encoding NAD-dependent alcohol dehydrogenase from Bacillus stearothennophilus was cloned and overexpressed as a GST fusion protein at a high level in Escherichia coli. The expressed fusion protein was purified simply by glutathione affinity chromatography. GST fusion protein was then cleaved by thrombin, while soluble enzyme was further purified by glutathione affinity chromatography. The recombinant enzyme had the same elctrophoretic mobility as the native enzyme from Bacillus stearothennophilus. The recombinant enzyme catalyzed the oxidation of a number of alcohols and exhibited high activities towards secondary alcohols. The $K_m\;and\;V_{max}$ values of the recombinant enzyme for ethanol were 5.11 mM and 61.35 U/mg, respectively. Pyridine and imidazole notably inhibited the enzymatic activity. The activity of the recombinant enzyme optimally proceeded at pH 9.0 and $70^{\circ}C$. The midpoint of the temperature-stability curve for the recombinant enzyme was approximately $68^{\circ}C$, and the enzyme was not completely inactivated even at $85^{\circ}C$. The recombinant enzyme showed a high resistance towards denaturing agents (0.05% SDS, 0.1 M urea). Therefore, due to its stability and relatively broad substrate specificity, the recombinant enzyme could be utilized in bio-industrial processes and biosensors.

• Journal of Korea Technical Association of The Pulp and Paper Industry
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• v.35 no.2
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• pp.39-45
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• 2003

This study was performed to develop the new type enzyme immobilization sheet from carboxymethylated refiner mechanical pulp (CRMP) substrate. Enzyme immobilization was attempted to couple carboxyl groups of CRMP with amino groups of the enzyme, trypsin, through the reaction of carbodiimide reagent, 1-ethyl-3-(3-dimethyl aminopropyl)-carbodimide (EDC ). Immobilization carrier, water insoluble CRMP fraction (CRMP-IS), was successfully reacted with the enzyme, formed peptide linkage like -CONH- at 1680$cm^{-1}$ / and new ester linkage like -COO$CH_3$, methylester at 1735$cm^{-1}$ /, and produced enzyme immobilized substrate (CRMP-IST). The enzyme immobilized handsheet was prepared by mixing the above chelated enzyme immobilized substrate(CRMP-IST) with kraft pulp by paper sheet machine like papermaking process. The sheet weight and strength were increased with increasing dosage of CRMP-IST, and decreased at more than 10％ mixing of CRMP-IST, but higher than the controls. Concerning activities of immobilized trypsin(CRMP-IST) sheet by caseinolysis, the teared-off sheet with shaking was shown higher enzyme activities than sheet shape without shaking. In conclusion, this enzyme immobilized sheet would be expected easy handling for practical application and reutilization.

• Korean Journal of Food Science and Technology
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• v.25 no.3
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• pp.238-242
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• 1993

For the Purpose of utilizing tannins in the functional foods and crude drugs, the enzyme inhibition of tannins isolated from Korean green tea were determined. Acetone extract from Korean green tea showed inhibition effect against the angiotensin converting enzyme. The galloyl tannins showed higher inhibition activity against angioteosin converting enzyme than the nongalloyl tannins. In terms of stereo isomers, (-)-epicatechins had higher inhibition activity than the (+ )-catechins. The synergistic activity was also observed. Tannins isolated from Korean green tea appeared to be incompetitive inhibitor against the angiotensin converting enzyme.

• Journal of Microbiology and Biotechnology
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• v.26 no.6
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• pp.1087-1097
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• 2016

An intracellular lipase from Anoxybacillus flavithermus HBB 134 was purified to 7.4-fold. The molecular mass of the enzyme was found to be about 64 kDa. The maximum activity of the enzyme was at pH 9.0 and 50℃. The enzyme was stable between pH 6.0 and 11.0 at 25℃, 40℃, and 50℃ for 24 h. The K_m and V_max of the enzyme for pNPL substrate were determined as 0.084 mM and 500 U/mg, respectively. Glycerol, sorbitol, and mannitol enhanced the enzyme thermostability. The enzyme was found to be highly stable against acetone, ethyl acetate, and diethyl ether. The presence of PMSF, NBS, DTT and β-mercaptoethanol inhibited the enzyme activity. Hg²⁺, Fe³⁺, Pb²⁺, Al³⁺, and Zn²⁺ strongly inhibited the enzyme whereas Li⁺, Na⁺, K⁺, and NH₄⁺ slightly activated it. At least 60% of the enzyme activity and stability were retained against sodium deoxycholate, sodium taurocholate, n-octyl-β-D-glucopyranoside, and CHAPS. The presence of 1% Triton X-100 caused about 34% increase in the enzyme activity. The enzyme is thought to be a true lipase since it has preferred the long-chain triacylglycerols. The lipase of HBB 134 cleaved triolein at the 1- or 3-position.

• Journal of Microbiology and Biotechnology
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• v.14 no.4
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• pp.829-835
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• 2004

A fibrinolytic enzyme has been found in several bacteria isolated from fermented food. This study was carried out to investigate the purification and characteristics of the fibrinolytic enzyme produced by Bacillus subtilis KCK-7 originated from Chungkookjang. The fibrinolytic enzyme was purified to homogeneity from the culture supernatant using ammonium sulfate fractionation and chromatographies on DEAE-cellulose and on Sephadex G-100. The final specific activity of the purified enzyme increased 11.0-fold, and the protein amount in the purified enzyme was about 16% of that in the culture supernatant. The molecular weight of the purified enzyme was estimated to be about 45,000 by SDS-PAGE. The optimum pH and temperature for the enzyme activity were pH 7.0 and $60^{\circ}C$, respectively. The enzyme activity was relatively stable up to $60^{\circ}C$ over the pH range of 7.0-10.0. The fibrinolytic enzyme activity increased by $Ca^{2+}$ and $Cu^{2+}$, whereas it was inhibited by $Hg^{2+}$ and $Ba^{2+}$. In addition, it was severely inhibited by PMSF and DFT. It is suggested that the purified enzyme was a serine protease for the fibrinolysis. The purified enzyme could completely hydrolyze fibrin in vitro within 8 h. Hence, it is suggested that the purified enzyme can be put into practice as an effective thrombolytic agent.

• Archives of Pharmacal Research
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• v.3 no.1
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• pp.7-12
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• 1980

Ampliciline was synthesized from 6-amino-pencillanic acid (6-APA) and D-.alpha. phenylglycine methyl ester by using amplicilin synthesizing enzyme from Peudomonas melanogenum (IAM 1655). The whole cell enzyme was immobilized by entrapping it in the polyacrylamide gel lattices. The polymer used in the enzyme entrapment was made from 150 mg per ml of acrylamide monomer and 8 mg per ml of N, N'-methylenebisacrylamide. About 200 mg/whole cell enzyme was mixed in the polymer for entrapment. The maximal activity retention after immobilization was 56%. The optimal pH values for the whole cell enzyme and the immobilized whole cell enzyme were 6.0 and 5.9, respectively. The optimal temperature for the enzyme activity were the same for both type of preparations. The enzyme stabilities against pH and heat increased for immobilized whole cell enzyme. Immobilized cell was more stable especially in the acidic condition while both type were found to be very suceptible to thermal inactivation at a temperature above 4.deg.C. The kinetic constants obtained from Lineweaver-Burk plot based on two substate reaction mechanism showed somewhat higher value for immobilized whole cell enzyme as compared to the whole cell enzyme : the Km value for 6-APA were 7.0 mM and 12.5 mM while Km values for phenylglycine methyl ester were 4.5 mM and 8.2 mM, respectively. Using the immobilized whole cell enzyme packed in a column reactor, the productivity of ampiciline was studied by varying the flow rate of substrate solution. At the space velocity, SV, 0.14 hr$^{-1}$ the conversion was 45%. Operational stability found in terms of half life was 30 hr at SV = 0.2 hr.

• Archives of Pharmacal Research
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• v.23 no.6
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• pp.548-553
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• 2000

Two drug-enzyme conjugates of dexamethasone-subtilisin and dexamethasone-cellulase have been synthesized and characterized to study their drug-protein incorporation ratio, immunoreactivity, enzyme activity and stability and these studies proved that a variety of drug enzyme conjugates could also be synthesized and characterized.

• Food Science and Preservation
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• v.12 no.6
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• pp.643-649
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• 2005

This study was investigated to find out optimal medium maximizing the production of fibrinolytic enzyme by Bacillus sp. isolated from Korean fermented soybean paste, which could hydrolyze the fibrin produced through the blood coagulation mechanism in human body. Among carbon sources tested, galactose was most effective for the enzyme production, and the level of the concentration for the optimal enzyme production was $4\%$(w/v). For nitrogen sources tested, malt extract was most effective for the enzyme production, and level of the concentration for optimal enzyme production was $4\%$(w/v). For mineral sources tested, $K_2HPO_4$ was most effective for enzyme production. The enzyme was maximally produced by cultivating the organism at the liquid medium of the initial pH 6 and temperature of $40^{\circ}C$.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.10
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• pp.1457-1463
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• 2005

In this research, the possibilities of using canola meal (CM) in place of soybean meal (SBM), and also the effects of multi-enzyme and phytase supplementation on the performance of quails were investigated. For this purpose, soybean meal (44% CP), canola meal (37% CP), phytase (produced from Peniophora luci) and multi-enzyme ($\beta$-glucanases, pectinases, cellulases and hemicellulases) were used. CM was used supplying 0, 25 and 50% of CP from SBM and each of the phytase and multi-enzyme blends were added to the each level. This study was conducted with 675 day old quails (Coturnix coturnix Japonica) in 9 groups with 3 replicates including 25 birds (mixed sex) per replicate. Nine isocalaric and isonitrogenous diets were prepared. The effects of enzymes and CM levels were studied with a 3${\times}$3${\times}$3 factorial arrangement for three CM levels (0, 25 and 50%), three treatments (without enzyme, phytase enzyme and multi-enzyme) and three replicates. While the 25% CM level did not affect the liveweight gain 50% CM level decreased the liveweight gain (p<0.05). Multi-enzyme addition to the 50% CM group increased the liveweight gain compared to the other groups (p<0.05). CM levels and enzyme supplementation had no effect on feed consumption, feed conversion ratio, dressing percentage, viability, tibia ash content, Ca and P contents of tibia ash, viscera weight, gizzard weight and length of growth period. While heart weight and liver weight were not affected by CM levels, but they were affected by enzyme supplementation. CM levels and enzyme supplementation did not affect final liveweight, feed consumption, feed conversion ratio, egg yield, egg weight, shell weight and shell index during laying period. The increase in the CM level lightened the colour of the yolk (p<0.05).