• Korean Journal of Microbiology
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• v.43 no.3
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• pp.222-226
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• 2007

Proteases are degradative enzymes which hydrolyze a peptide bond between amino acids and they are abundantly applied to commercial field. In order to screen new source of pretense, bacteria secreting extracellular pretense were isolated by enrichment culture from deep sea water samples of East Sea, Korea. A bacterium, named as HJ19, showed the best growth and the largest clear zone in plates supplemented skim milk at $30^{\circ}C$. The partial DNA sequence analysis of the 16S rRNA gene, phenotypic tests and morphology identified that this strain was In genus Micrococcus. The strain HJ19 could not grow at $10^{\circ}C$ but it started growth and showed pretense activity at $20^{\circ}C$. The optimal growth was at $37^{\circ}C$ and the maximal protease activity at $30^{\circ}C$ was about 480unit/ml.

• Korean Journal of Microbiology
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• v.28 no.3
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• pp.199-203
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• 1990

To study distribution of bacterial flora and their biochemical characteristics in the Antarctic soilecosystem, these experiments were performed during the austral summer(Feb., 1989) on the King George Island, Antarctica. The numbers of heterotrophic bacterial colonies and extracellular enzyme actibities were estimated from the Antarctic terrestrial soils which were sampled from 17 different locations near Sejong station (Korea) and Teniente Jubany station (Argentina) on the King George Island. The numbers of heterotrophic bacterial colonies were extremely variable with sampling sites and incubation temperatures. Arithmetric average numbers were $2.5\times 10^{4}$, $2.7\times 10^{7}$ , $6.9\times 10^{5}$ CFU/$cm^{3}$ soil at the incubation temperature of $37^{\circ}C$, $25^{\circ}C$ and $4^{\circ}C$, respectively. The activities of extracellular $\alpha$-glucosidase, $\beta$-glucosidase and N-acetyl-$\beta$-glucosaminidase were shown as similar mean percentage in the colonies obtained at different temperatures. Mean value of protease activities, however, was remarkably higher (92%) in the colonies grown at $4^{\circ}C$,.

• Journal of the Korean Society of Food Science and Nutrition
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• v.30 no.2
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• pp.222-227
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• 2001

The members of the genus Vibrio include harmless aquatic strain as well as strains capable of causing infections in human and fish. Pathogenic mechanisms are only understood for Vibrio cholerae O1 and O139 and not for the majority of Vibrio species. Twelve clinical and nonclinical strains were examined by in vitro and in vivo experiments for the importance of extracellular enzymes as a virulence determinant of Vibrio species. In vivo cytotoxicity assay was performed by injecting approximately $10^{8}$ cells/mL into mice (BALB/c). V. harvyi and V. vulnificus showed 100% lethality within 3hr after bacterial injection. V. fluvialis and four strains of V. parahaemolyticus showed 50% lethality within 4hr. V. mimicus, V. alginolyticus and V. furnissii revealed 30% lethality within 9hr. Nonclinical strains, V. campbellii and V. ordalii, did not show any lethality. In vitro protease and hemolytic activities were also good indicators for clinical and nonclinical strains of Vibrio species. The clinical strains showed much higher activities than nonclinical strains. The activity of some clinical strains of re-isolates was evidently increased. Most clinical strains had $\beta$ hemolytic activity. The results demonstrate that the prevalent distribution of extracellular proteases in pathogenic Vibrio sp. implies their importance as a virulence determinant.

• CELLMED
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• v.4 no.1
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• pp.6.1-6.12
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• 2014

Anthrax is the deadly disease for human being caused by Bacillus anthracis. Instantaneous research work on the mode of infection of the organism revealed that different proteases are involved in different steps of pathogenesis. Present study reports the in silico characterization and the detection of pathogenic proteases involved in anthrax infection through protein-protein interaction. A total of 13 acid, 9 neutral, and 1 alkaline protease of Bacillus anthracis were selected for analysing the physicochemical parameter, the protein superfamily and family search, multiple sequence alignment, phylogenetic tree construction, protein-protein interactions and motif finding. Among the 13 acid proteases, 10 were found as extracellular enzymes that interact with immune inhibitor A (InhA) and help the organism to cross the blood brain barrier during the process of infection. Multiple sequence alignment of above acid proteases revealed the position 368, 489, and 498-contained 100% conserved amino acids which could be used to deactivate the protease. Among the groups analyzed, only acid protease were found to interact with InhA, which indicated that metalloproteases of acid protease group have the capability to develop pathogenesis during B. anthracis infection. Deactivation of conserved amino acid position of germination protease can stop the sporulation and germination of B anthracis cell. The detailed interaction study of neutral and alkaline proteases could also be helpful to design the interaction network for the better understanding of anthrax disease.

• Korean Journal of Microbiology
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• v.35 no.4
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• pp.296-301
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• 1999

A Bacillus strain from Korean soil was isolated and identified to be Bacillus licheniformis B1 through various biochemical tests, VITEK, and MIDI system analysis. The strain produced extracellular amylase and protease. Whether or not the strain can perform Chungkookjang fermentation with autoclaved soybean and Kanjang fermentation was determined in this study. In Chungkookjang fermentation, browining materials of strong anti-oxidant increased 8-fold, and 2-fold in Kanjang, compared with initiation material for fermentation. Maximal protease activity in Chungkookjang was observed one day after inoculation. Protease activities in Kanjang decreased to the half, and then maintained constant values during fermentation, probably due to the inhibitory effect of salt on protease activities. High molecular mass of nucleic acids was identified in Chungkookjang and Kanjang. Since the nucleic acids were not observed in autoclaved soybean, they seem to be originated from B. licheniformis B1. This study demonstrated successive fermentation of Chungkookjang and Kanjang by B. licheniformis B1 isolated from nature, and suggested possible development of food rich in browing and nucleic acids.

• Journal of Food Hygiene and Safety
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• v.16 no.1
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• pp.33-36
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• 2001

V. parahaemolyticus possessed an extracellular alkaline protease activity during the stationary growth phase. Various factors such as initial pH of medium, incubation temperature and shaking rate were investigated far optimizing the production of alkaline protease from V. parahaemolyticus ATCC 17802. Maximal activity of the protease was obtained when the bacteria were grown in 2% skim milk medium in 0.1M tris/HCl buffer (pH 7.6). Maximal activity of the protease was obtained when the bacteria were growls at initial pH of 7.6, incubation temperature 37$^{\circ}C$ and shaking rate of 250 rpm.

• Parasites, Hosts and Diseases
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• v.36 no.2
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• pp.133-142
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• 1998

A 40 kDa cysteine protease was purified from the crude extract of adult worms of GMnnophalloines seoi by two consecutive steps: Sephacryl S-200 HR and DEAE- Sephacel chromatography. Enzyme activities were completely inhibited by cysteine protease inhibitors, L-lorans-epoxysuccinylleucylamido (4-guanidino) butane (E-64) and iodoacetic acid, strongly suggesting that the purified enzyme belongs to the cysteine family of proteases. The enzyme was maximally acive at pH 4.5 in 0.1 M of buffer, and its activity was greatly potentiated in the presence of 5 mM dithiothreitol. The protease degraded macromolecules with differential capabilities : it degraded extracellular matrix proteins, such as collagen and fibronectin, with a stronger activity against collagen than fibronectin . However, the enzyme digested hemoglobin and human immunoglobulins only slightly. leaving them nearly intact after an overnight reaction. Our results suggest that the cysteine protease of G. seoi adults is potentially significant in the nutrient uptake from the host intestine.

• Journal of Microbiology
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• v.41 no.4
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• pp.289-294
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• 2003

Gelatinase is a proteolytic enzyme that hydrolyzes gelatin. Gelatinolytic activity was detected from culture broths of Streptomyces griseus IFO13350 and HH1 by paper disc assays on 0.5% agar plates containing 1% gelatin. The concentrated extracellular protein from the S. griseus was analyzed by SDS polyacrylamide gel, and two proteins, with molecular weights of 30 and 28 kDa, respectively, were identified to have gelatinase activity by gelatin zymography. The protein with a molecular weight of 28 kDa was confirmed to be S. griseus trypsin (SGT). The effects of metal ions and metal chelators on the protease activity of the SGT were studied. Of the metal ions tested, only manganese was found to enhance the protease activity, 2.6 times, however, $Co^{2+},\;Cu^{2+},\;and\;Zn^{2+}$, and metal chelators, such as EDTA and EGTA, inhibited the SGT activity. When the protease activity of the SGT was measured at various pHs, in the presence of 5 mM $MnCl_2$, its highest activity was at pH 11.0, whereas only 60% of the maximum activity was observed between pHs 4.0 and pH 6.0, and almost 80% activity between pHs 7.0 to pH 10.0. The protease activity was measured at various temperatures in the presence of 5 mM $MnCl_2$. The SGT was found to be stable up to $60^{\circ}C$ for 30 min, while only 16% of the enzyme activity remained at $60^{\circ}C$, and at $80^{\circ}C$ almost all the activity was lost. The optimal temperature for the protease activity was $50^{\circ}C$.

• Biomedical Science Letters
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• v.6 no.3
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• pp.209-221
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• 2000

The purpose of this study was to investigate the practical availability of pretense production that can be used at home after isolating Serratia sp.2000-1 which produced extracellular pretense from clinical specimen. Basic production conditions and partial enzymatic characteristics of pretense produced by Serratia sp. 2000-1 was as follows: The kind and concentration of carbohydrate, nitrogen and metal salts for optimal enzyme production condition were each identified as the concentration of 1.5% glucose, 2.0% CSP, and 0.1% CaCl$_2$, and the optimal temperature, time and initial pH for culture were each 3$0^{\circ}C$, 72 hours, and pH 8.0. The final enzymatic yeild that was purified by 3 steps with ammonium sulfate precipitation (45~80%), DEAE-cellulose column chromatography, and Sephadex G-200 gel chromatography was 14.4%, and enzyme inactivity rate increased approximately 291314s. The optimal temperature and pH for purified pretense activity were 35$^{\circ}C$ and pH 7.0~8.0, and purified pretense activity was relatively stable by 4$0^{\circ}C$ at pH 6~10 for 30 min, however heating at 6$0^{\circ}C$ for 30 min, it liminated detectable pretense activity. The pretense activity was activated by $Mg^{2+}$, $Ba^{2+}$, $Ca^{2+}$, Mn$^{2+}$, but inactiviaed by Hg$^{2+}$, Ag$^{2+}$, Cu$^{2+}$, and the pretense activity was inhibited strongly by SDS among enzyme activity inhibitors. Further study is required to evaluate the practical availability of pretense production that can be used at home by isolating Serratia sp. from more clinical specimen and examining pretense more in details.

• The Korean Journal of Mycology
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• v.39 no.2
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• pp.99-104
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• 2011

Shiitake breeding requires the procedures of mating of two different parental strains and selection of hybrid strains that have good traits for the mushroom production. In this study, we tested the possibility of the use of chromogenic plate-based assay for extracellular enzyme production in order to assess and find good biochemical properties-possessed hybrid strains that were generated from genetic cross of the monokaryotic strains derived from two different parental strains of Lentinula edodes Sanjo-101ho and Sanjo-108ho. We observed that there was difference in the ability of producing ${\beta}$-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease among the monokaryotic strains. We could also comparatively assess that the ability of the seven extracellular enzymes production in the hybrid strains depended on the mating combination of the monokaryotic strains. Our results demonstrate that the assessment method for extracellular enzyme production using chromogenic plate assay could be usefully applied to the assessment of the hybrid strains derived from the breeding procedure of L. edodes.