• Fisheries and Aquatic Sciences
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• v.3 no.3_4
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• pp.188-194
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• 2000

A protease purified from hepatopancreas of shrimp, Penaeus japonicus, had maximum activity at $70^{\circ}C$ and in neutral and alkaline pH ranges. Specific activity at optimum reaction condition of the protease was estimated to be approximately 12 U/mg/min. The protease was stable in neutral and alkaline pH ranges and activity was retained after heat treatment at $50^{\circ}C$ for 30 min. Apparent $K_m$ and $V_{max}$ value against casein substrate were estimated to be $0.29\%$ and $7.8see^{-1}$, respectively, and those against N-CBZ-L-tyrosine p-nitropheny1 ester (CBZ­Tyr-NE) were 0.38 mM and $2,400 see^{-1}$, respectively. The N-termina1 sequence of the protease showed high homology to the trypsin from same species and the proteases from shrimp. Myosin heavy chain (MHC) from shrimp tail meat was the most susceptible to the protease and actin/tropomyosin were degraded progressively during 4 hr incubation, but to a lesser degree than MHC.

• Ceramist
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• v.21 no.3
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• pp.204-215
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• 2018

Proteases are one of the most abundant classes of enzymes in living organisms and have been considered major targets for drug development. However, despite the ability to specifically cleave their substrates, many attempts to assay protease activity have generally relied upon the use of gel zymography or fluorophore-labeled peptide substrates, which is limited in rapid and multiplex analysis. Here we review the recent advances in nanoparticle (NP)-utilized assays of protease activity focused on in vitro and cell imaging-based approaches. Owing to large surface area and unprecedented physical properties of NPs, these approaches are anticipated to facilitate many applications related to protease activity-based disease diagnosis and drug discovery.

• Microbiology and Biotechnology Letters
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• v.30 no.1
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• pp.26-32
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• 2002

For commercial production of Korean fermented anchovy sauce through rapid fermentation, a bacterial strain which showed the high protease activity was isolated from a commercially fermented anchovy sauce. The isolate was Bacillus amyloliquefaciens, and named as B. amyloliquefaciens HTP-8. The incubation temperature, initial pH, and cultivation time for optimal production of protease by B. amyloliquefaciens HTP-8 were $30^{\circ}C$, 7.0, and 3 days, respectively. In jar fermenter, B. amyloliquefaciens HTP-8 showed higher protease activity when grown at pH 7.0. The protease was partially purified by 80% ammonium sulfate precipitation and CM-Sephadex C-50 ion exchange chromatography. The partially purified enzyme had specific activity of 103.3 units/mg, yield of 0.4%, and purification fold of 43.0. The optimal pH and temperature for the protease activity were 10.0 and $50^{\circ}C$, respectively. The protease was relatively stable at the pH range of 7.0~12.0 and at the temperatures below 4$0^{\circ}C$. The activity of the enzyme was inhibited by $Ag^{＋}$ /, $Ba^{2＋}$ and selectively inhibited by PMSF, suggesting that it is a serine protease.

• 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 $50^{\circ}C$.

• Journal of Sericultural and Entomological Science
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• v.35 no.1
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• pp.1-6
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• 1993

The activity change of mulberry seeds protease was compared during germination for 5 days at 28$^{\circ}C$ in the dark place after daily hormone injection of different concentration. The protease from germinated mulberry seeds for 4 days was partially purified and the enzyme characteristics was investigated. The protease activity of mulberry seeds treated by hormone was highest with 10 $\mu$m GA3 followed by 10 $\mu$M zeatin and 10 $\mu$M kinetin. The protease activity of mulberry seeds was increased by 14% with 10ml agar culture that control at 4th day of germination. The protease from mulberry seeds was purified 313 fold by DEAE-Toyo-pearl 650M, Butyl-Toyopearl, Hydrozylapatite and Toyopearl HW 55M. After purification, the specific activity of the enzyme was 175 units/mg. Optimum pH and temperature of protease from mulberry seeds was 5.0 and 37$^{\circ}C$, respectively. The protease was stable below 37$^{\circ}C$ and the enzyme activity was decreased by 50%, when incubated at 52$^{\circ}C$ for 10minutes. The protease activity of mulberry seeds was inhibited by metal ions such as mercury, iron, zinc, copper, but activited by magnesium, choromium, aluminium ions. The Km value of the protease was 0.89mM with azocasein as a subscribe.

• Journal of Microbiology and Biotechnology
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• v.21 no.1
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• pp.20-27
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• 2011

The purification and characterization of a $Mn^{2+}$-dependent alkaline serine protease produced by Bacillus pumilus TMS55 were investigated. The enzyme was purified in three steps: concentrating the crude enzyme using ammonium sulfate precipitation, followed by gel filtration and cation-exchange chromatography. The purified protease had a molecular mass of approximately 35 kDa, was highly active over a broad pH range of 7.0 to 12.0, and remained stable over a pH range of 7.5 to 11.5. The optimum temperature for the enzyme activity was found to be $60^{\circ}C$. PMSF and AEBSF (1 mM) significantly inhibited the protease activity, indicating that the protease is a serine protease. $Mn^{2+}$ ions enhanced the activity and stability of the enzyme. In addition, the purified protease remained stable with oxidants ($H_2O_2$, 2%) and organic solvents (25%), such as benzene, hexane, and toluene. Therefore, these characteristics of the protease and its dehairing ability indicate its potential for a wide range of commercial applications.

• Korean Journal of Veterinary Research
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• v.55 no.3
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• pp.175-179
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• 2015

Proteases play important roles in parasite development and host parasite interactions. The protease of Kudoa spp. has been recognized as a key factor of severe proteolysis of fish muscle post-mortem; however, there is little information available regarding the protease of Kudoa (K.) septempunctata, which was recently identified as a cause of food poisoning in humans. The present study was conducted to isolate and characterize proteases to elucidate the type of protease contained in the parasite and determine the optimal pH for protease activity. We confirmed the cysteine protease and metalloprotease produced by K. septempunctata. While the cysteine protease showed optimal activity at pH 5 that decreased rapidly with increasing pH, the optimal activity of metalloprotease was pH 7, and it remained stable from pH 6 to pH 8. These results indicate that the pH of cysteine protease is not proper for fish muscle postmortem, and that metalloprotease can act in human intestines. Overall, the present study provides important information that improves our understanding of the role of protease physiology and the subsequent food poisoning caused by K. septempunctata.

• Applied Biological Chemistry
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• v.44 no.4
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• pp.246-250
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• 2001

In order to produce alkaline protease, psychrotrophic Bacterium which have high enzyme activity, was isolated by using enrichment culture from soil samples and identified as genus Bacillus sp. The optimal pH and temperature for the enzyme activity were pH 6 and $40^{\circ}C$. The temperature range of high enzyme activity was $20{\sim}40^{\circ}C$. The optimal initial pH of culture condition for enzyme was pH 6. The most favorable carbon and nitrogen sources for the production of protease by Bacillus sp. WRD-2 were 3% maltose and 4% yeast extract, respectively.

• Korean Journal of Microbiology
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• v.10 no.2
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• pp.69-72
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• 1972

The studies of neutral protease which was obtained by passing through Sephadex A-50 had been reported not long ago. Since that time the author also conducted the research to be investigated the physical properties of acid protease absorbed by Sephadex A-50. The results are summarized as follows : 1) Cultivating Aspergillus oryza SHW-131 on a wheat bran medium, the acid protease including neutral protease is very sensitive for temperature. 3) Activity of acid protease is very sensitive for temeprature. 3) This enzyme was proved, what is called, to be a sort of weak acid protease. It's optimum pH was lied in about 4.5. 4) A range of pH for stability is far more narrow than any other protease. 5) The acid protease is dropped by EDTA solution in its activity.

• Journal of Microbiology
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• v.38 no.3
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• pp.160-168
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• 2000

A-factor I a microbial hormone that can positively control cell differentiation leading to spore formation and secondary metabolite formation in Streptomyces griseus. to identify a protease that is deeply involved in the morphological and physiological differentiation of Streptomyces, the proteases produced by Streptomyces griseus IFO 13350 and its A-factor deficient mutant strain, Streptomyces griseus HH1, as well as Streptomyces griseus HH1 transformed with the afsA gene were sturdied. In general Streptomyces griseus showed a higher degree of cell growth and protease activity in proportion to its ability to produce a higher amount of A-factor. In particular, the specific activity of the trypsin of Streptomyces griseus IFO 13350 was greatly enhanced more than twice compared with that of Streptomyces griseus HH1 in the later stage of growth. The specific activity of the metalloprotease of Streptomyces griseus HH1 was greatly enhanced more than twice compared with that of Streptomyces griseus IFO 13350, and this observation was reversed in the presence of thiostreptione, However, Streptomyces griseus HH1 transformed with the afsA gene showed a significantly decreased level of trypsin and metalloprotease activity compared with that of the HH1 strain. There was no significant difference between Streptomyces griseus IFO 13350 and HH1 strain in their chymotrypsin and thiol protease activity, yet the level of leu-amionpeptidase activity was 2 times higher in Streptomyces griseus HH1 than in strain IFO 13350 . Streptomyces griseus HH1 harboring afsA showed a similar level of enzyme activity , however, all the three protease activities sharply increased and the thiol protease activity was critically increased at the end of the fermentation. When a serine protease inhibitor, pefabloc SC, and metalloprotease inhibitor, EDTA, were applied to strain IFO 13350 to examine the in vivo effects of the protease inhibitors on the morpholofical differentiation, the formation of aerial meycelium and spores was delayed by two or three days.