• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1291-1300
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• 2018

Objective: This experiment was conducted to investigate the effects of dietary crude protein (CP) level and exogenous protease supplementation on growth performance, serum metabolites, carcass traits, small intestinal morphology and endogenous protease activity in broiler chickens reared under a tropical climate. Methods: A total of 480 day-old male broiler chicks were randomly assigned to eight dietary treatments in a $4{\times}2$ factorial arrangement. The main effects were CP level (21.0%, 19.7%, 18.5%, or 17.2% from 1 to 21 days and 19.0%, 17.9%, 16.7%, or 15.6% from 22 to 35 days) and protease enzyme supplementation (0 ppm or 500 ppm). All experimental diets were fortified with synthetic feed-grade lysine, methionine, threonine and tryptophan to provide the minimum amino acid recommended levels for Cobb 500. Results: Reducing dietary CP linearly reduced (p<0.05) growth performance, serum albumin, total protein, and carcass traits and increased (p<0.05) serum triglycerides and abdominal fat. There was no consistent effect of reducing dietary CP on morphological parameters of the intestine and on the pancreatic and intestinal endogenous protease activity (p>0.05). Protease supplementation improved (p<0.05) feed conversion ratio, body weight gain, carcass yield and intestinal absorptive surface area. Conclusion: Protease supplementation, as measured by growth performance, intestinal morphology and carcass yield, may alleviate the detrimental effects of low protein diets in broiler chickens.

• Journal of the Korean Society of Food Science and Nutrition
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• v.42 no.9
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• pp.1461-1466
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• 2013

In this study, we ascertained the characteristics of crude protease extracted from fruits (fig, kiwifruit and pineapple) as well as traditional Korean fermentation starters (bio nuruk, traditional nuruk, meju and rice koji) to determine their suitability for industrial application. Crude protease extracted from traditional Korean fermentation starters was found to have a higher optimum temperature ($70^{\circ}C$) and salt concentration (1~3%) but a lower optimum pH (3~6) value compared to the corresponding values for the protease extracted from fruits. On comparison, the total activities of protease per gram unit follow the order: bio nuruk> rice koji> traditional nuruk> fig> pineapple> meju> kiwifruit. Based on our results, we conclude that protease extracted from traditional Korean fermentation starters has potential for application in food industry, for example, as a meat tenderizer for sausage manufacturing and as a protease for cheese production.

• Applied Biological Chemistry
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• v.38 no.6
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• pp.534-540
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• 1995

Alkalophilic coryneform bacteria TU-19 isolated from soil extracellularly produced at least three proteases (Protease I, II, and III). Investigating the cultural conditions related to the enzyme production of this bacterial cell, the optimum pH and temperature were 10.0 and $30^{\circ}C$, respectively. In order to purify these enzymes from the 2 day culture broth ammonium sulfate fractionation, gel filtration and QAE-Sephadex column chromatography were performed step by step. And then these three proteases were purified to near homogeneity by judging from SDS-PAGE pattern, and had the molecular weights of 120, 80, and 45 kilodaltons, respectively. The optimum pH and temperature for the enzyme activity of Protease I and II were 10.5 and $45^{\circ}C$, respectively, and Protease II were 11.0 and $50^{\circ}C$. And the enzymes were completely inhibited by PMSF suggesting serine protease, but not affected by pCMB. 1,10-phenanthroline, IAA, and EDTA.

• The Korean Journal of Food And Nutrition
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• v.8 no.3
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• pp.192-198
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• 1995

For the production of EMC, various professes and lipases were used to hydrolyse cheese sulk. The optimal conditions of various proteases were as follows, pronase-3$0^{\circ}C$, p14 7.0, pancreatln-4$0^{\circ}C$, pH 8.0, pacific protease-3$0^{\circ}C$, pH 7.0 and protease from Asp. sp. -5$0^{\circ}C$, pH 8.0. The optimal conditions of various lipases were as follows ; pancreatic lipase-5$0^{\circ}C$, pH 8.0, palatase ML-5$0^{\circ}C$, pH 7.0 and lipase form Candida -4$0^{\circ}C$, pH U.0. After hydrolysation under optimal conditions, the amounts of free amino acid and free fatty ac14 were increased with reaction time. Hydrolysates of pacific protease and pronase were showed high amount of free amino acid(0.67mg/ml and 0.74mg/ml). Especially EMC had high amount of glutamic acid and leucine. Lipase from Candida cylindracea produced high amount of free fatty acid (24.63 mg/ml) Butyric acrid, palmitic acid, stearic acid and oleic acid among free fatty acids were showed high amounts. Sensory evaluation of various MC were tasted nth 8 panelist. EMC produced with pancreatic lipase was most bitterness and EMC produced with palatase ML was best acceptable cheese flavor.

• Asian-Australasian Journal of Animal Sciences
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• v.29 no.7
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• pp.998-1003
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• 2016

Although exogenous protease enzymes have been used in poultry diets quite extensively, this has not been the case for pig diets. In general, due to their better gut fermentative capacity and longer transit time, pigs have greater capacity to digest dietary proteins than poultry. However, in early-weaned piglets, the stress brought about by weaning adversely affects the digestion of dietary proteins. Therefore, a study was conducted to determine the effects of a commercial protease enzyme in weanling pigs. Indices of growth, nutrient digestibility, blood profiles, fecal microflora, fecal gas emission and fecal scores were measured during the study. A total of 50 weanling pigs ($6.42{\pm}0.12kg$) at 28 d of age were randomly assigned to receive 1 of 2 dietary treatments: i) control diet (corn-soy based) with no supplemental protease (CON), and ii) control diet+200 g/ton protease (PROT) for 42 d. A completely randomized design consisting of 2 treatments, 5 replicates, and 5 pigs in each replicate was used. Growth performance in terms of body weight ($27.04{\pm}0.38kg$ vs $25.75{\pm}0.39kg$; p<0.05) and average daily gain ($491{\pm}7.40g$ vs $460{\pm}7.46g$; p<0.05) in PROT fed pigs were increased significantly, but gain per feed ($0.700{\pm}0.01$ vs $0.678{\pm}0.01$; p>0.05) was similar between treatments at d 42. Relative to CON pigs, PROT fed pigs had increased (p<0.05) apparent total tract digestibility ($84.66%{\pm}0.65%$ vs $81.21%{\pm}1.13%$ dry matter and $84.02%{\pm}0.52%$ vs $80.47%{\pm}1.22%$ nitrogen) and decreased (p<0.05) $NH_3$ emission ($2.0{\pm}0.16ppm$ vs $1.2{\pm}0.12ppm$) in the feces at d 42. Except for a decreased (p<0.05) in blood creatinine level, no differences were observed in red blood cell, white blood cell, lymphocyte, urea nitrogen, and IgG concentrations between treatments. Fecal score and fecal microflora (Lactobacillus and E. coli) were also similar between CON and PROT groups. Overall, the supplementation of protease enzyme in weanling pigs resulted in improved growth rate and nutrient digestibility. Exogenous protease enzyme reduced fecal $NH_3$ emission, thus, potentially serving as a tool in lowering noxious gas contribution of livestock production in the environment.

• Applied Biological Chemistry
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• v.41 no.1
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• pp.6-12
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• 1998

To extract insoluble proteins and to improve functional properties of abolished proteins, a protease producing Aspergillus sp. MS-18 was isolated from soil. The enzyme was purified and its enzymological characteristics were investigated. It was found that production of protease reached to the maximum when the wheat brae medium containing, 3% arabinose, 0.5% polypepton, 0.1% $(NH_4)_2SO_4$ and 0.2% magnesium chloride was cultured for 3 days. Protease was purified 16.9 folds after ion exchange chromatography and gel filtration and the specific activity was 340.4 unit/mg. Purified enzyme was confirmed as a single band by the polyacrylamide gel electrophoresis. The molecular weight of protease was estimated to be 30,000. Crystalization form of purified protease was a stick shape with rounding edges. The optimum pH and temperature for the protease activity were 9.0 and $60^{\circ}C$, respectively. The enzyme was stable in pH 7.0-12.0 at $50^{\circ}C$. The activity of purified enzyme was inhibited by $Hg^{2+}$, $Cu^{2+}$, $Zn^{2+}$ and $Pb^{2+}$, whereas it was activited by $Na^+$, $Mg^{2+}$ and $Mn^{2+}$. The activity of the protease was inhibited by the treatment with ethylenediaminetetraacetic acid and phenylmethane sulfonyl fluoride. The result suggests that the purified enzyme is a serine protease with metal ion at active site. Km and Vmax of purified protease were $29.33\;{\mu}mole/L$ and $5.13\;{\mu}g/min$, respectively.

• Microbiology and Biotechnology Letters
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• v.25 no.1
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• pp.56-61
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• 1997

An alkaline protease producing microorganism was isolated from korean traditional Meju and identified as Scopulariopsis brevicaulis. The optimum culture condition of Scopulariopsis brevicaulis for the production of alkaline protease was as follow: 2% soluble starch, 0.2$, tryptophan, 0.1% (NH$_{4}$) $_{2}$S$_{2}$O$_{8}$ 0.2% NaHPO$_{4}$, pH 7.5, 35$\CIRC $C. The optimum pH and temperature for the enzyme activity of alkaline protease producing Scopulariopsis brevicaulis were pH 9.0 and 50$\circ $C, respectively. The enzyme was relatively stable at pH 6.0~11.0 and at temperature below 40$\circ $C. The activity of the enzyme was inhibited by Hg$^{2+}$ whereas Cu$^{2+}$ gave rather activating effects on the enzyme activity. Phenylmethanesulfonyl fluoride inhibited the enzyme activity. This result indicates that serine is very important role in this enzyme. Km value for casein was 1.2410$^{4}$ M/L, V$_{max}$ value for casein was 25.99 $\mu $g/min. This enzyme hydrolyzed casein more rapidly than the hemoglobin.

• Microbiology and Biotechnology Letters
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• v.19 no.4
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• pp.383-389
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• 1991

In order to produce alkaline protease, psychrotrophic bacterium which have high enzyme activity at low temperature, was isolated by using enrichment culture from various samples and identified as genus alkalopsychrotropic Pseudomonas sp. RP-222. The optimal culture conditions for enzyme production were pH- 10.0, temperature-$20^{\circ}C$ and culture time-4 days. The optimum pH and temperature for the enzyme activity were pH 10.5 and $40^{\circ}C$, respectively and the enzyme was relatively stable at pH 7.0~13.0 and below $50^{\circ}C$. The enzyme was inhibited by ethylenediaminetetraacetate and phenylmethylsulfonylfluoride, indicating that the enzyme was a serine metalloenzyme, but considerably stable in the presence of surface active agents. Activity of the enzyme was increased by the addition of 0.05% Na-$\alpha$-olefin sulfonate.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.330-336
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• 2004

Fifty-two pathogenic Vibrio parahaemolyticus strains were isolated from the environments of Busan and Yeosu, Korea. Forty-three of these strains showed protease activities, whereas 4 strains showed $\alpha / \beta$ hemolysin activities and 6 strains had urease activities. Their pathogenic factors were not overlapping except one strain, which had both protease and hemolysin activities. The 6 urease-positive strains (V. parahaemolyticus YKB4, YKB14, S25, YFB20, YFO21, and YFO22) showed the same biochemical characteristics as a reference strain [V. parahaemolyticus KCTC 2471 (urease-negative)], except for urease production. The 6 urease-positive strains showed different urease activities in their culture supernatant during the growth. The urease activity of S25 increased sharply at the late exponential phase, and was the highest at the initial stationary phase and was kept until the late stationary phase. The other 5 isolates, except C25, showed urease activities at the mid-stationary phase and increased steadily until the late stationary phase, when the urease activity was maximal. To compare the degree of virulence of V. parahaemolyticus with different pathogenic factors, hemolysin, protease, or urease-positive strains were injected into groups of 10 each of ICR mice (7- to l0-week-old males). The lethal rates of urease-positive V. parahaemolyticus, YKB14, YKB4, and S25, were significantly high, being 50, 70, and 80%, respectively. Protease-positive V. parahaemolyticus strains FM39 and FM50 showed 40% and 60% of lethal rate, respectively. Hemolysin-positive V. parahaemolyticus strains S34 and S72 had no mortality, similar to nonpathogenic V. parahaemolyticus FM12.

• Journal of Life Science
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• v.17 no.2 s.82
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• pp.272-278
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• 2007

Proteases are enzymes that break peptide bonds between amino acids of other proteins and occupy a crucial position with respect to their applications in both physiological and commercial fields. In order to screen new source of protease, bacteria producing extracellular proteases at low temperature were isolated from deep sea water of East Sea, Korea. A bacterium showing the best growth rate and production of an extracellular protease at low temperature was designated HJ 47. The DNA sequence analysis of the 16S rRNA gene, phenotypic tests and morphology led to the placement of this organism in the genus Pseudoalteromonas. Although maximal growth was observed at $37^{\circ}C$, enzyme production per culture time was maximum at $20^{\circ}C$. At this temperature, extracellluar protease production was detected from the end of the exponential phage to stationary phase, and maximal at 15 hours after initial production. The optimum temperature and pH of the protease were found to be $35^{\circ}C$ and 8.