• Microbiology and Biotechnology Letters
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• v.1 no.2
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• pp.93-98
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• 1973

The acid protease was isolated from the culture broth of the Penicillum sp. grown in the wheat bran media. 'quot;The crude purification of this enzyme was carried out by extraction with distilled water and precipitated with saturated ammonium sulfate. " The activity of this enzyme was found to be very strong by Folin′s colorimetric method. The results were as follows. 1. The optimum pH of the enzyme activity was at 3.0 and its optimum temperature was 5$0^{\circ}C$. 2. Although the enzyme activity to hydrolyze casein was maximal at 5$0^{\circ}C$, its activity decreased rapidly by about 50％, treated at 5$0^{\circ}C$ for 30 min. When treated at 4$0^{\circ}C$ for 60 min, the enzyme activity decreased to 75％ of original value and did not decrease any more. 3. The enzyme was stable at pH 2.0 to 6.0. 4. This enzyme activity was not effected by metal ions; C $d^{＋＋}$, Z $n^{＋＋}$, $Co^{＋＋}$, H $g^{＋＋}$, M $n^{＋＋}$, P $b^{＋＋}$, $Mg^{＋＋}$, L $i^{＋}$, C $u^{＋＋}$, $Ba^{＋＋}$, A $g^{＋}$, $Al^{＋＋＋}$, $Ca^{＋＋}$, F $e^{＋＋}$, and F $e^{＋＋}$ 5. Also, it was not effected by treatemnt of EDTA.

• The Journal of Korean Society of Virology
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• v.29 no.3
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• pp.175-182
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• 1999

An attractive target for anti-herpes chemotherapy is the herpes simplex virus 1 (HSV-1) protease encoded by the UL26 gene. HSV-1 protease is essential for DNA packaging and virus maturation. To perform high throughput for potent inhibitors, the efficient production of larger amounts of highly purified enzyme and protease activity assay method must be established. In this report, expression in E. coli and purification of the protease gene of HSV-1 strain F was investigated. The protease gene was cloned pET28, and the nucleotide sequence of protease catalytic domain of HSV-1 compared strain F with other strains (KOS and CL101). In these results the F strain was different in base sequence. However, the amino acid sequence was identifical. The HSV-1 protease was purified with His-tagged affinity column. The analysis of HSV-1 protease activity was performed by high performance liquid chromatography.

• Microbiology and Biotechnology Letters
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• v.7 no.4
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• pp.217-223
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• 1979

Six strains of mold which had high saccharifying and acid-productive ability were isolated from wild sources. The strain M-80 among them and Aspergillus kawachii, which was generally used as rice makkulli koji were used for this studies. The results obtained were summarized as follows. 1) $\alpha$-amylase activities of the strain M-80 and Asp, kawachii showed similarly as about 140W. V., while $\beta$-amylase activity of M-80 was 54 A. U. and Asp. kawachii was 40 A. U. 2) Acid protease activity of M-80 was higher then Asp. kawachii but alkaline protease activity was lower Asp. kawachii respectively. 3) The contents of total acid, ethanol and fusel oil in makkulli brewing with M-80 were higher those of Asp. kawachii and methanol contents of the tested two strains were about 33mg/％. 4) Fifteen kinds of free amino acid were detected from makkulli brewing of two strains, and free amino acid contents of M-80 were 10％ higher than those of Asp. kawachii.

• Korean Journal of Microbiology
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• v.47 no.3
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• pp.255-262
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• 2011

Six protein-degrading bacteria were isolated from digestive organ of Harmonia axyridis. These isolates were categorized as Staphylococcus sciuri subsp. sciuri (3 strains), Bacillus subtilis (1 strain), and Bacillus thuringiensis (2 strains) by 16S rRNA gene sequence analysis. The Staphylococcus sp. CB2-3 was selected as a protease-producing bacterium which showed the highest protease activity of 58.5 U/ml at the pH 5.0 medium. The optimal pH and temperature of protease activity were pH 5.0 and $40^{\circ}C$, respectively. This acid protease had a relatively high stability of 80% between $30-50^{\circ}C$ at broad temperature range. The opimal medium compositions of carbon, nitrogen and mineral source for cell growth and protease activity were investigated. When sorbitol (0.5%) was used as carbon source, enzyme activity was increased about 2 times than that of the basal medium. When skim milk (0.5%) was used as nitrogen source, activity was increased about 2.5 times than that of the control. Cell growth and enzyme activity were increased by mineral source such as KCl, $K_2HPO_4$, $FeSO_4$, but was completely inhibited by divalent ions such as $Co^{2+}$, $Zn^{2+}$, $Mn^{2+}$, $Cu^{2+}$.

• Analytical Science and Technology
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• v.26 no.3
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• pp.182-189
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• 2013

The purpose of this research is to separate and quantitate selenium species in some food samples with HPLC-ICP-MS. Cation exchange chromatography showed efficient separation only for inorganic Se species while reversed phase ion pair chromatography showed good separation for both inorganic and organic Se species. $C_8$ column ($Symmetryshield^{TM}\;RP_8$, 3.5 ${\mu}m$, $4.6{\times}150$ mm) was used with optimum condition of 5% methanol mobile phase, 0.05% of nonafluorovaleric acid ion pairing reagent. Five standard Se species of Se(IV), Se(VI), SeCys(selenocystein), SeMet(selenomethionine) and Se-M-C(seleno methyl cystein) were separated successfully under the optimum condition (mobile phase; 5% methanol, ion-pairing reagent; 0.05% nonafluorovaleric acid, flow rate; 0.9 mL $min^{-1}$). To extract Se species, microwave assisted and enzyme-assisted extraction methods were studied. In enzyme-assisted extraction method, protease I for garlic, protease I plus trypsin for pork and mackerel, and protease XIV for tuna showed the best extraction efficiency. With the optimum condition for each sample, it was found that mostly inorganic Se, SeCys and SeMet are present in the sample studied ranging from few ${\mu}g$ $g^{-1}$ to few tens of ${\mu}g$ $g^{-1}$.

• Parasites, Hosts and Diseases
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• v.52 no.1
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• pp.117-120
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• 2014

Several studies have reported that the citrus red mites Panonychus citri were an important allergen of citrus-cultivating farmers in Jeju Island. The aim of the present study was to purify and assess properties of a cysteine protease from the mites acting as a potentially pathogenic factor to citrus-cultivating farmers. A cysteine protease was purified using column chromatography of Mono Q anion exchanger and Superdex 200 HR gel filtration. It was estimated to be 46 kDa by gel filtration column chromatography and consisted of 2 polypeptides, at least. Cysteine protease inhibitors, such as trans poxy-succinyl-L-leucyl-amido (4-guanidino) butane (E-64) and iodoacetic acid (IAA) totally inhibited the enzyme activities, whereas serine or metalloprotease inhibitors did not affect the activities. In addition, the purified enzyme degraded human IgG, collagen, and fibronectin, but not egg albumin. From these results, the cysteine protease of the mites might be involved in the pathogenesis such as tissue destruction and penetration instead of nutrient digestion.

• KSBB Journal
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• v.17 no.3
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• pp.276-282
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• 2002

Symbiotic bacteria with highly effective insecticidal activities were isolated and compared with their physiological characteristics from seven species of entomopathogenic nematodes belong to Steinernamatidae and Heterorhabditidae sp., and three of them were identified as Xenorhabdus nematophilus. Culture characteristics, insecticidal activities, pretense activities and fatty acid contents of various symbiotic bactierial isolates were also examined. In the case of cell growth and insecticidal activity, XR-PC and XR-MK were superior to other species when cultured in vitro. The insecticidal activity were highest at the early exponential growth phase, and gradually decreased with time. The protease activity of XR-DR was remarkable compared to other species. In the case of HE-HY, however the pretense activity increased in parallel with cell growth. Interestingly, the fatty acid patterns of Xenorhabdus nematophilus isolated from different emtomopathogenic nematode, showed remarkable differences in their contents of 12:0, 14:0, 16:1 cia 5 and 17:0 cyclo and hydroxy and branch factty acids were varied from 2% to 15% among total fatty acid contents.

• Fisheries and Aquatic Sciences
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• v.25 no.2
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• pp.76-89
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• 2022

The Lophius litulon liver was used as raw material for the extraction of fish oil via various extraction methods. The extraction rate by water extraction, potassium hydroxide (KOH) hydrolysis and protease hydrolysis were compared and the results revealed the protease hydrolysis extraction had a higher extraction rate with good protein-lipid separation as observed by optical microscope. Furthermore, subsequent experiments determined neutrase to be the best hydrolytic enzyme in terms of extraction rate and cost. The extraction conditions of neutrase hydrolysis were optimized by single-factor experiment and response surface analysis, and the optimal extraction rate was 58.40 ± 0.25% with the following conditions: enzyme concentration 2,000 IU/g, extraction time 1.0 h, liquid-solid ratio 1.95:1, extraction temperature 40.5℃ and pH 6.5. The fatty acids composition in fish oil from optimized extraction condition was composed of 19.75% saturated fatty acids and 80.25% unsaturated fatty acids. The content of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were 8.06% and 1.19%, respectively, with the ratio (6.77:1) surpassed to the recommendation in current researches (5:1). The results in this study suggest protease treatment is an efficient method for high-quality fish oil extraction from Lophius litulon liver with a satisfactory ratio of DHA and EPA.

• International Journal of Industrial Entomology and Biomaterials
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• v.22 no.2
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• pp.83-93
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• 2011

Biochemical and enzymatic characterization for extracellular protease isolated from Cordyceps militaris cultivated on rice bran medium was investigated. C militaris produced proteolytic enzymes from 10 days after inoculation, maximum enzyme production was found at 25 days. The optimum temperature and pH of proteases production was at $25^{\circ}C$ and pH 7.0, respectively. The protease activity was observed in the four peaks (Pro-I, Pro-II, Pro-III, and Pro-IV) separated through Sephadex G-100 column chromatography. The separated protease was optimally active at $25^{\circ}C$. Optimum pH of the protease was between 7 and 8. Enzyme was also stable over at $30-80^{\circ}C$. The enzyme was highly stable in a pH range of 4-9. Protease activity was found to be slightly decreased by the addition of $Mg^{2+}$, $Mn^{2+}$, $Zn^{2+}$, $Fe^{2+}$ and $Cu^{2+}$, whereas inhibited by the addition of $Ca^{2+}$ and $Co^{2+}$ Protease activity was inhibited by protease inhibitor PMSF. On the other hand, the partially purified protease was investigated on proteolytic protease activity by zymogram gel electrophoresis using three substances (casein, gelatin and fibrin). Four active bands (F-I, FII, F-III, and F-IV) of fibrin degradation were revealed on fibrin zymogram gels. Both of F-II and FIII showed caseinolytic, fibrinolytic and gelatinolytic activities in three gels. Thermostability, pH stability, and pH-thermostability of the enzyme determined the residual fibrinolytic activity also displayed on fibrin zymogram gel. The only one enzyme (F-II) displayed over a broad range of temperature at $30-90^{\circ}C$. The FII displayed fibrinolytic activity in the pH range 3-5, but was inactivated in the range of pH 6-11. The F-I and F-III showed enzyme activity in the pH range of 6-11. In the pH-thermostability, the F-II only kept fibrinolytic activity after heating at $100^{\circ}C$ for 10, 20 and 30 min at pH 3 and pH 7, respectively. On the other hand, the F-II was retained activity until heating for 10 min under pH 11 condition. By using fibrin zymogram gel electrophoresis, extracellular fibrinolytic enzyme F-II from C. militaris showed unusual thermostable under acid and neutral conditions.

• Parasites, Hosts and Diseases
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• v.43 no.4 s.136
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• pp.157-160
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• 2005

A 29kDa cysteine protease of Taenia solium metacestodes was purified by Mono Q anion-exchanger and Superose 6 HR gel filtration chromatography. The enzyme was effectively inhibited by cysteine protease inhibitors, such as iodoacetic acid (IAA) and trans-epoxy-succinyl-L-leucyl-amido (4-guanidino) butane (E-64) while inhibitors acting on serine- or metallo-proteases did not affect the enzyme activity. The purified enzyme degraded human immunoglobulin G (IgG), collagen and bovine serum albumin (BSA), but human IgG was more susceptible for proteolysis by the enzyme. To define the precise biological roles of the enzyme, more detailed biochemical and functional studies would be required.