• Korean Journal of Microbiology
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• v.39 no.4
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• pp.216-222
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• 2003

Human caspase-9, an essential apoptosis initiator protease, was excessively degraded when expressed in Escherichia coli under the conventional induction condition. To optimize the conditions for induction and develop a rapid purification method for obtaining significant amounts of wild-type procaspase-9, we expressed procaspase-9 as GST fusion in E. coli. The addition of 0.01 mM IPTG as an inducer to the bacterial culture and decreasing the culture temperature to 25oC improved the production of procasapse-9 protein by circumventing proteolytic degradation in E. coli. The wild-type procaspae-9 was purified to approximately 70% purity with relatively high yields using the method developed in this study. In addition, we found that GST-caspase-9 is autocatalytically cleaved after aspartic acid 315, which is the same site for processing in mammalian cells, during expression in E. coli.

• Fisheries and Aquatic Sciences
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• v.17 no.1
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• pp.37-46
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• 2014

Fresh sea squirt meat requires a modified processing and preservation process because it has a short shelf life due to its high moisture content and strong proteolytic enzyme activity. In this study, bottled sea squirt meat prepared in vegetable oil (BSMO) to enhance the consumer acceptability was exposed to ${\gamma}$-ray (Co60, 10 kGy/h) irradiation to extend the shelf life without the use of a heating process. Response surface methodology was used to determine the optimal mixing ratio of BSMO containing 5% dehydrated fresh meat. Texture analysis and nutritional evaluation were also performed on a control and BSMO samples. The volatile basic nitrogen (VBN) content and total cell count were measured to determine the shelf life of irradiated BSMO products during chilled storage at $4^{\circ}C$ for 60 days. According to a panel of 10 trained tasters (aged 20-29 years), the optimal mixing formulation was 80 g meat in 60 mL of mixed vegetable oil (30 mL of olive oil and 30 mL of sesame oil). The highest rated formulation, according to a panel of nine trained tasters (aged ${\geq}30$ years), was 80 g meat in 60 mL of mixed vegetable oil (42 mL of olive oil and 18 mL of sesame oil). Moisture, ash, and protein contents in BSMO did not change significantly (P < 0.05) compared with the control. A higher lipid content ($0.84{\pm}0.23$ to $2.13{\pm}0.61$; P < 0.05) was observed due to the presence of vegetable oil on the surface of BSMO. The vegetable oil raised the hardness, springiness, cohesiveness, gumminess, chewiness, and resilience of BSMO. BSMO products remained edible after 50 days of storage at $4^{\circ}C$ based on the VBN content (BSMO 1: $27.92{\pm}0.96$ mg/100 g, BSMO 2: $24.84{\pm}1.95$ mg/100 g) and total cell count (BSMO 1: $4.60{\pm}0.80$ log CFU/mL, BSMO 2: $3.65{\pm}0.20$ log CFU/mL) when compared with standard levels of VBN (25.00 mg/100 g) and total cell count (5 log CFU/mL), respectively. The results showed that irradiated BSMO products could help to expand the processed seafood market and increase the popularity of seafood among the younger generations.

• The Korean Journal of Mycology
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• v.33 no.1
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• pp.11-17
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• 2005

To increase the fibrinolytic activity and production of mycelium, extracts of 7 plant species were supplemented to the growth media of Armillaria mellea, and mycelial growth and enzymatic activity in the mycelium extracts of A. mellea were estimated. The mycelial production of A. mellea was slightly increased by adding ASH-R, UDVN or RGR extract, whereas KG extract significantly affected the growth. Supplement of ASH-S, UDVN and RGR extracts increased proteolytic activity from 36.8 to 46.1% Fibrinolytic activity was increased to $50{\sim}65%$ by supplement with RVS, ASH-S and RGR extracts, respectively. Enzyme extracts of the fungus grown with RGR extract supplement degraded all chains of fibrinogen within 2 hours, whereas control was required 3 hours. Degradation of fibrin fragments by the enzyme extracts was also observed through microscopy.

• Journal of Life Science
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• v.26 no.2
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• pp.164-173
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• 2016

Isotocin (IT), a nonapeptide homolog of oxytocin in mammals, has been suggested to be involved in physiological processes including social behaviors, stress responses, and osmoregulation in teleost fish. To study its structure and function, the gene encoding the IT precursor was cloned from the genomic DNA and brain cDNA of the cinnamon clownfish, Amphiprion melanopus. The IT precursor gene consists of three exons separated by two introns, and encodes an open reading frame of 156 amino acid (aa) residues, comprising a putative signal peptide of 19 aa, a mature IT protein of 9 aa, a proteolytic processing site of 3 aa, and 125 aa of neurophysin. Tissue-specific analysis of the IT precursor transcript indicated its expression in the brain and gonads of A. melanopus. To examine its osmoregulatory effects, the salinity of the seawater (34 ppt) used for rearing A. melanopus was lowered to 15 ppt. Histological analysis of the gills indicated the apparent disappearance of an apical crypt on the surface of the gill lamella of A. melanopus, as pavement cells covered the surface upon acclimation to the lower salinity. The level of Na⁺/K^+-ATPase activity in the gills was increased during the initial stage of acclimation, followed by a decrease to its normal level, suggesting its involvement in osmoregulation and homeostasis. The only slight increase in the level of IT precursor transcript in the A. melanopus brain upon low-salinity acclimation suggested that IT played a minor role, if any, in the process of osmoregulation.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.17 no.2
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• pp.117-124
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• 1984

Sarine scrap usually comprises about $40\%$ of the raw fish in processing. The purpose of this study is to establish the desirable methods for proteinaceous materials from the sardine scrap through autolysis or enzymatic digestion and to convert them into useful by-products such as sardine sauce. Sardine scrap was chopped and mixed with equal weight of water, and be hydrolyzed them by autolysis and/or by addition of commercial proteolytic enzyme and various concentrations of sodium chloride. The optimal conditions for hydrolysis of sardine scrap were revealed in temperature at $55^{\circ}C$ and 4 hours digestion with bromelain($0.4\%$) and commercial complex enzyme ($6.0\%$), and those conditions were also applicated in autolysis. The maximum hydrolyzing rate of protein and released amino nitrogen were $82.5\%,\;5.2\%$ through autolysis, $84.3\%,\;5.8\%$ by bromelain digestion and $92.5\%,\;5.9\%$ by complex enzyme, respectively. In the products prepared from sardine scrap through autolysis or bromelain digestion, hypoxanthine was dominant, as $17.4 {\mu}mole/g$, dry matter for autolysis and $16.0 {\mu}mole/g$, dry matter, for bromelain digestion among the nucleotidcs and their related compounds, respectively. The abundant free amino acids were leucine, glutamic acid, lysnie, valine and alanine. The contents of those amino acids were $51.3\%,\;48.3\%$ of the total free amino acids, respectively. And the contents of 5'-IMP and TMAO were negligible but total creatinine was developed in value from $9.2\%\;to\;10\%$ of total extracted nitrogen. The flavor of sardine sauce prepared from sardine scrap by autolysis or enzyme digestion were not inferior to that of traditional Korean soy sauce by sensory evaluation.

• Microbiology and Biotechnology Letters
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• v.13 no.4
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• pp.409-415
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• 1985

This study has been carried out in order to investigate the bacterial quality of fish meat paste products and the characteristics of isolated thermodurics from the products. Twenty samples of crab-flavored fish stick (Kematsal), 23 samples of plate fish meat paste (Panomuk, Kamaboko), 5 samples of fried fish meat paste (Tigimomuk), 2 samples of roasted fish meat paste (Puduromuk, Chikuwa), 20 samples of fish sausage were collected from processing plants and supermarkets in Pusan, Korea during the period from May to October in 1984. The results obtained are as follows. Amont the samples collected from supermarkets, roasted fish meat paste and fried fish meat paste marked hish counts in coliforms and fungi while very low in the samples of crab-flavored fish stick and plate fish meat paste. Salmonella was not detected in all the samples examined and Staphylococcus aureus was detected only in fried fish meat paste, Thermoduric bacteria were detected less than 10$^2$/g in the samples of crab-flavored fish stick and plate fish meat paste, which might come from subsidiary materials such as starch and seasonings. Among the isolated bacteria, distribution of the proteolytics were more than 87％ and the lipolytics were less than 20％. Gram positive bacteria was more than 70％ in crab-flavored fish stick and plate fish meat paste, 47.3％ in fried fish meat paste. And rod in shape was almost more than 90％ in all the samples. The most heat resistant bacterium isolated from the samples was identified as a Bacillus licheniformis(named B. licheniformis CR-11). The strain showed strong proteolytic activity and also grew well at above 2$0^{\circ}C$. The growth rate and generation time of CR-11 strain were 0.31 hr$^{-1}$ , 2.24 hr at 2$0^{\circ}C$, 0.64 hr$^{-1}$ , 1.09 hr at 3$0^{\circ}C$ and 0.78 hr$^{-1}$ , 0.89 hr at 35$^{\circ}C$. Heat resistance value of the spores of CR-11 strain suspended in phosphate buffer solution was D$_{85}$ $^{\circ}C$=41.9 min, D$_{90}$ $^{\circ}C$=27.9 min, D$_{95}$ $^{\circ}C$=10.2 min, D$_{100}$ $^{\circ}C$=4.3 min (Z=13.8$^{\circ}C$)

• Korean Journal of Fisheries and Aquatic Sciences
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• v.12 no.4
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• pp.235-240
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• 1979

A study on the amino acid composition of raw frozen krill, and krill solubles manufactured in forms of paste and powder has been carried out. The raw frozen krill was thawed, chopped, mixed and homogenized with same amount of water. The mixture was autolyzed or hydrolyzed by tile addition of $0.2\%$ pronase-p, a commercial proteolytic enzyme, to the weight of the raw frozen krill at $45^{\circ}C$ for 4 hours. After a thermal inactivation of enzymes at $95^{\circ}C$ for 15 minutes, the autolysate and the hydrolysate were centrifuged and filtered through gauzes, respectively, and then tile lipid layer in the supernatant was removed, The autolysate and the hydrolysate were finally concentrated under reduced atmospheric pressure in a rotary vacuum evaporator at $45^{\circ}C$ for 1 hour to produce the krill solubles in form of paste. The powdered krill solubles were prepared by the addition of $5\%$ starch to the autolysate and hydrolysate and by means of concentration in the rotary vacuum evaporator at $45^{\circ}C$ for 30 minutes and a forced air drying at $58^{\circ}C$ for 3 hours with a air velocity of 3m/sec. Among the amino acids in raw frozen krill, glutamic acid, lysine, and aspartic acid showed high values in quantity and then followed leucine, alanine, arginine, glycine and proline. The qnantity of histidine was very small and that of cystine was only in trace. The krill solubles in forms of paste and powder prepared by autolysis and hydrolysis with pronase-p revealed almost the same patterns in amino acid composition as in raw frozen krill. In case of free amino acids, a large quantity of it in raw frozen krill consisted of lysine, arginine, proline, alanine and leucine. The quantities of cystine, histidine and glutamic acid were, in contrast, very small. In the soluble krill paste prepared by autolysis, lysine, leucine, threonine and alanine existed in large quantities among the free amino acids and cystine, aspartic acid and histidine existed in small quantities. The contents of almost all of the free amino acids ill soluble krill paste perpared by hydrolysis with pronase-p were increased slightly as compared with those in soluble krill paste prepared by autolysis. In this product, the contents of cystine, histidine and serine were very low and lysine, leucine, arginine and proline were the dominant group in quantities among the free amino acids. The krill solubles in forms of paste and powder were not inferior to whole egg in the view point of its essential amino acid composition.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.8
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• pp.1265-1273
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• 2005

The purpose of this study was to prepare accelerated salt-fermented anchovy sauce using a shrimp processing byproducts (head, shell and tail) as a fermenting accelerator, and to investigate its physicochemical and enzymatic properties. Four types of sauces were prepared with 0, 10, 20, and 30$\%$ (w/w) addition of shrimp byproduct and fermented at 24$\pm$2$^{\circ}C$ for 360 days. During fermentation, all four type sauces decreased moisture content (67.5$\%$68.0$\%$ to 64.0$\∼$64.8$\%$) and pH (5.52$\∼$7.10 to 5.03$\∼$6.58), but showed increase in their crude protein (7.0$\∼$8.2 to 10.8$\%$) and volatile basic nitrogen contents (40$\∼$75 to 180$\∼$200 mg/100 g of sauce). The ratio of amino nitrogen to total nitrogen contents of control (0$\%$) and sauce with 10$\%$ shrimp byproducts (10$\%$ sauce) were maximized at 270 days, whereas 20$ \% $ and 30$\%$ added sauces were at 180 days. Endoprotease and exoprotease activities of anchovy sauces added with 20$\%$ and 30$\%$ of shrimp byproducts tend to be higher than those of control (0$\%$) and 10$\%$ addition. Proteolytic activities of sauces at pH 9 were about 2 times higher than those at pH 6. Amidolytic activities for LeuPNA decreased remarkably during fermentation, and control (0$\%$) almost lost their activity at 180 days, while additional sauces were relatively stable. These suggest that alkaline pretense of anchovy and shrimp byproducts as a endoprotease mainly contributed to the fermentation of salt-fermented sauces. The protein molecular weight distribution of sauces indicated 2 groups of peaks (peak 1,>70,000 da and peak 2, 3,000$\∼$29,000 da). As the fermentation proceeded, peak 1 tended to decrease in all of sauces, but peak 2 increased rapidly from 30 to 270 days. Optimum fermentation periods of control and 10$\%$ sauces were 270 days and those of 20$\%$ and 30$\%$ sauce were 180 days. The results suggest that shrimp byproduct can be used as accelerator of salt-fermented sauce.