• Journal of Microbiology and Biotechnology
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• v.28 no.1
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• pp.1-11
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• 2018

An antioxidant peptide derived from Pinctada fucata meat using an Alcalase2.4L enzymatic hydrolysis method (named AOP) and identified by LC-TOF-MS has promising clinical potential for generating cosmetic products that protect skin from sunshine. To date, there have been few published studies investigating the structure-activity relationship in these peptides. To prepare antioxidant peptides better and improve their stability, the design and expression of an antioxidant peptide from Pinctada fucata (named DSAOP) was studied. The peptide contains a common precursor of an expression vector containing an ${\alpha}$-helix tandemly linked according to the BamHI restriction sites. The DNA fragments encoding DSAOP were synthesized and subcloned into the expression vector pET-30a (+), and the peptide was expressed mostly as soluble protein in recombinant Escherichia coli. Meanwhile, the DPPH radical scavenging activity, superoxide radical scavenging activity, and hydroxyl radical scavenging activity of DSAOP $IC_{50}$ values were $0.136{\pm}0.006$, $0.625{\pm}0.025$, and $0.306{\pm}0.015mg/ml$, respectively, with 2-fold higher DPPH radical scavenging activity compared with chemosynthesized AOP (p < 0.05), as well as higher superoxide radical scavenging activity compared with natural AOP (p < 0.05). This preparation method was at the international advanced level. Furthermore, pilot-scale production results showed that DSAOP was expressed successfully in fermenter cultures, which indicated that the design strategy and expression methods would be useful for obtaining substantial amounts of stable peptides at low costs. These results showed that DSAOP produced with recombinant Escherichia coli could be useful in cosmetic skin care products, health foods, and pharmaceuticals.

• Journal of Animal Science and Technology
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• v.48 no.4
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• pp.541-554
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• 2006

Present review is to introduce an omasal sampling technique in rumen proteolysis and to consider some information on the omasal sampling technique with particular emphasis on methodological aspects. Use of the omasal sampling technique provides a new opportunity for accurate estimation of rumen metabolism with overcoming limitations of previous in vivo, in vitro and/or in situ methods. The potential advantages of the present technique compared with post-ruminal sampling techniques include following points; 1) only rumen cannulated animals are required, 2) less endogenous nitrogen (N) is contaminated in omasal digesta and 3) omasal digesta are devoid of exposure to acid peptide hydrolysis occurring in the abomasum. Estimates of soluble non-ammonia N (SNAN) in omasal digesta indicate that the assumptions underlying the in situ method that rapidly degradable N fraction can be degraded at an infinite rate and only insoluble dietary N escapes the rumen may be not valid. Quatitatively higher peptide concentration rather than free amino acid and soluble protein in escapable SNAN suggests that hydrolysis of peptide to amino acid may be the rate-limiting step in rumen proteolysis.

• Journal of Life Science
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• v.27 no.11
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• pp.1324-1330
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• 2017

Salt, or sodium chloride (NaCl), is a critical ingredient in many foods. It has roles in the flavor profiles of food products, textures of foods and preservation of foods against microbes. However, it increases risks of hypertension and is closely related to the development of cardiovascular disease. In recent years, health concerns related to sodium intake caused an increased demand for salt-reduced products in worldwide; it became necessary to develop natural salt-alternative products that are globally competitive. In a recent study, researchers succeeded in obtaining a natural salt enhancer through the hydrolysis of vegetable- and animal-matter mixtures. This study used various methods to identify and quantify peptide-containing arginine as a salt-alternative peptide (SAP) in an optimum combination. Arginine, or dipeptide-containing arginine, was analyzed as a standard substance using an NMR spectroscopy. The NMR carbon signal of the guanidine group of the standard substance was verified in a similar location (the L-arginine (Arg) was 156.8 ppm, the Arg-Alanine was 156.4 ppm and the Arg-Serine was 156.4 ppm). The results suggested that it is possible to analyze peptide-containing arginine quantitatively through the hydrolysis of vegetable- and animal-matter mixtures.

• Journal of the Korean Society of Food Science and Nutrition
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• v.32 no.5
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• pp.745-751
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• 2003

This study was carried out to investigate whether peptide produced from wheat protein by enzyme hydrolysis can be used as a natural antimicrobial agent. Antimicrobial peptide was obtained from wheat protein hydrolyzed by 7 of pretense. The produced antimicrobial peptide was purified through ultrafiltration, membrane filtration and HPLC and molecular weight and amino acid sequence of the purified antimicrobial peptide were determined. Among hydrolysate produced from wheat protein by 7 of protease, antimicrobial activity was observed for the peptide obtained from Asp. saito protease. The Asp. saito protease did produce antimicrobial hydrolysate showing the highest antimicrobial activity at reaction condition of 37$^{\circ}C$ and pH 6.0, but not at reaction condition above 5$0^{\circ}C$. Wheat protein hydrolysate was fractionated by membrane filtration and showed antimicrobial activity between molecular weight 1,000~3,000. The antimicrobial activity fraction obtained by membrane filtration was separated through HPLC and showed antimicrobial activity in the peak of retention time 31.1~31.8 min. We could convince this hydrolysate as heat-stable peptide since antimicrobial activity was maintained after treated with heat for 15 min at 121$^{\circ}C$. Molecular weight of antimicrobial peptide identified by MALDI-mass was 1,633. Amino acid sequence of antimicrobial peptide was cysteine, glycine, prolin, prolin, prolin, valine, valine, alanine, alanine and arginine.

• Korean Journal of Agricultural Science
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• v.29 no.2
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• pp.78-90
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• 2002

This study was carried out to investigate whether peptide produced from wheat protein by enzyme hydrolysis can be used as a natural antimicrobial agent. Antimicrobial peptide was obtained from wheat protein by protease of 7 species. The produced antimicrobial peptide was purified through ultrafiltration, membrane filtration and HPLC, and molecular weight and amino acid sequence of the purified antimicrobial peptide were determined. Among hydrolysate produced from wheat protein by protease of 7 species, antimicrobial activity was observed for the peptide obtained from Asp. saito protease. The Asp. saito protease did production antimicrobial hydrolysate showing the highest antimicrobial activity at reaction condition of $37^{\circ}C$ and pH 6.0, but not at reaction condition above $50^{\circ}C$. Wheat protein hydrolysate was fractionated by membrane filtration and showed antimicrobial activity between molecular weight 1,000 - 3,000. The antimicrobial activity fraction obtained by membrane filtration was separated through HPLC and showed antimicrobial activity in the peak of retention time 31.1 - 31.8 min. Since after wheat protein protease hydrolysate was heated during 15 min at $121^{\circ}C$, antimicrobial activity was maintained, we could be conviction as heat-stable peptide. Molecular weight of antimicrobial peptide identified by MALDI-mass was 1,633. Amino acid sequence of antimicrobial peptide was cysteine, glycine, prolin, prolin, prolin, valine, valine, alanine, alanine and arginine.

• Food Science of Animal Resources
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• v.23 no.1
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• pp.63-69
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• 2003

This experiment was carried out to study changes in solubility and emulsifying properties of whey protein. Whey protein hydrolysates were obtained from tryptic hydrolysis of whey protein concentrate at pH 8.0 and 37$^{\circ}C$ for 6 hours. Emulsifying activity of whey protein hydrolysate was highest at 4 hours of hydroysis and at 5.50% of DH. During hydrolysis of whey protein concentrate with trypsin, ${\alpha}$-lactalbumin was not easily broken down. But ${\beta}$-lactoglobulin was hydrolysed rapidly from the early stage of hydrolysis, producing several low molecular weight peptides, which have to participate in increasing emusifying activity. The solulbility of hydyolysates tended to increase depending on hydrolysis time; however, there was a gradual decrease after 5 hours. The hydrolysate had a minimum solubility near the isoelectric point range (pH 4∼5). The more hydrolysed the whey protein concentrates, the more soluble they are near the pl. They aye also more soluble above pH 6. Emulsifying activity of hydrolysates showed similar results to solubility. Creaming stability gradually increased when hydrolysis increased, increasing rapidly above pH 8 after 4 hours of hydrolysis.

• Journal of the Korean Applied Science and Technology
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• v.33 no.1
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• pp.176-185
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• 2016

Yeonsan Ogae has been known as supporting health and high efficacy of treatment. In recent days, as the efficacy of functional peptides has known, the optimization of oligo peptides production and its characteristics from Ogae legs has been performed. Response surface method was used to perform the optimizaion of enzyme hydrolysis. The range of processes was temperature ( 40, 50 and $60^{\circ}C$), pH( pH 6.0, 7.0 and 8.0 ), and enzyme( 1, 2 and 3% ). The degree of hydrolysis, amino acids, molecular weight of products were analyzed. The optimum process of enzyme hydrolysis were determined as temperature $58^{\circ}C$, pH 7.5, and enzyme concentration 3%. At optimum conditions, the degree of hydrolysis after 2 h reaction was 75-80%. The amino acid and were 168.131 mg/100 g, respectively. The molecular weight of products by using MALDI-TOF was ranged from 300 to 1,000 Da.

• Journal of Microbiology and Biotechnology
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• v.6 no.2
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• pp.92-97
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• 1996

Portage kinetic constants of peptide transport can be measured by competitive spectrophotometry. The kinetic constants of L-Glu-L-Glu transport in Escherichia coli were ascertained using L-Phe-L-3-thia-Phe (PSP) as a detector. Since the production of thiophenol upon intracellular hydrolysis of PSP was competitively inhibited by L-Glu-L-Glu, it was able to compute the kinetic constants of L-Glu-L-Glu using this method. The resulted data were in agreement with the values obtained by the method of Michaelis-Menten kinetics. The potential of this method was examined against dipeptide transport systems in various microorganisms. These results strongly suggest that the overall properties of individual systems for dipeptide transports can be easily characterized by competitive spectrophotometry.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2001.06a
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• pp.40-45
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• 2001

Raw starch-digesting amylase (BF-2A, M.W. 93, 000 Da) from Bacillus circulans F-2 was converted to two components during digestion with subtilisin. Two components were separated and designated as BF-2A' (63, 000 Da) and BF-2B (30, 000 Da), respectively. BF-2A' exhibited the same hydrolysis curve for soluble starch as the original amylase (BF-2A). Moreover, the catalytic activities of original and modified enzymes were indistinguishable in $K_{m}$, Vmax for, and in their specific activity for soluble starch hydrolysis. However, its adsorbability and digestibility on raw starch was greatly decreased. Furthermore, the enzymatic action pattern on soluble starch was greatly different from that of the BF-2A. A smaller peptide (BF-2B) showed adsorb ability onto raw starch. By these results, it is suggested that the larger peptide (BF-2A') has a region responsible for the expression of the enzyme activity to hydrolyze soluble substrate, and the smaller peptide (BF-2B) plays a role on raw starch adsorption. A similar phenomenon is observed during limited proteinase K, thermolysin, and endopeptidase Glu-C proteolysis of the enzyme. Fragments resulting from proteolysis were characterized by immunoblotting with anti-RSDA. The proteolytic patterns resulting from proteinase K and subtilisin were the same, producing 63- and 30-kDa fragments. Similar patterns were obtained with endopeptidase Glu-C or thermolysin. All proteolytic digests contained a common, major 63-kDa fragment. Inactivation of RSDA activity results from splitting off the C-terminal domain. Hence, it seems probable that the protease sensitive locus is in a hinge region susceptible to cleavage. Extracellular enzymes immunoreactive toward anti-RSDA were detected through whole bacterial cultivation. Proteins of sizes 93-, 75-, 63-, 55-, 38-, and 31-kDa were immunologically identical to RSDA. Of these, the 75-kDa and 63-kDa proteins correspond to the major products of proteolysis with Glu-C and thermolysin. These results postulated that enzyme heterogeneity of the raw starch-hydrolysis system might arise from the endogeneous proteolytic activity of the bacterium. Truncated forms of rsda, in which the gene sequence encoding the conserved domain had been deleted, directed the synthesis of a functional amylase that did not bind to raw starch. This indicates that the conserved region of RSDA constitutes a raw starch-binding domain, which is distinct from the active centre. The possible role of this substrate-binding region is discussed.d.

• Food Science and Preservation
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• v.10 no.1
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• pp.80-88
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• 2003

In order to establish the enzymatic hydrolysis system improving of taste and flavor in the preparation of soy protein hydrolysates using the enzymes with excellent hydrolytic ability and different hydrolysis pattern of soy protein, Degree of hydrolysis(DH) and surface hydrophobicity under the optimal conditions of enzyme reaction, hydrolysis patterns by the SDS electrophoresis and sensory evaluation of soy protein hydrolysates by enzyme reactions were investigated. Four enzyme reactions were highly activated at pH 7.0, 45$^{\circ}C$ under the optimal conditions. As result of changes on the pattern of soy-protein hydrolysates by SDS-electrophoresis, high molecular peptides of hydrolysates by No. 5(Mucor circinelloides M5) and No. 16(Bacillus megaterium B16) enzymes were slowly decrease and 66KD band of these were remained after 3hours reaction. Production of low molecular peptides of hydrolysates by No. 4(Aspergillus oryzae M4) and No. 95(Bacillus subtilis YG 95) enzymes were remarkably detected during the proceeding reactions. As results of HPLC analysis, low molecular peptides of 15∼70KD were mainly appeared during the proceeding enzyme reactions. And, the more DH was increased, the more SDS-surface hydrophobicity was decreased. Hydrolysates by No. 4 enzyme was not only the highest DH of all hydrolysates, but the strongest bitter taste in a sensory evaluation. Sweat taste among the hydrolysates showed little difference. But, when combinative enzymes were treated, combinative enzyme of No. 4(Aspergillus oryzae M4)and No. 16(Bacillus megaterium B16) showed the strongest sweat taste. In conclusion, we assumed that it will be possible to prepare the hydrolysates having functionality when soy-protein were hydrolyzed by these specific enzymes.