• 한국식품과학회지
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• 제48권3호
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• pp.201-207
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• 2016

메밀껍질은 메밀 알곡보다 유효성분이 많음에도 불구하고 식품학적 가치가 떨어져 대부분 폐기되고 있다. 따라서 이를 기능성 식품소재로 활용하기 위해 효소분해하여 기능성 올리고당을 생산하고 이들의 특성을 분석하였다. 올리고당 생산을 위한 최적 가수분해조건은 셀룰로스 분획의 경우 pH 5.0, $40^{\circ}C$, 기질농도 4%로 결정되었으며, 헤미셀룰로스 분획은 pH 5.0, $40^{\circ}C$, 30 unit으로 결정되었다. 최적 분해조건을 이용하여 72시간 효소분해 후 얻은 올리고당의 생산량을 측정한 결과, 셀룰로스와 헤미셀룰로스 분획으로부터 얻은 올리고당의 수율은 각각 132.37 g/kg 및 393.04 g/kg이었다. 또한 각 분획의 올리고당 함량을 측정한 결과, 포도당, 자일로스, xylobiose, xylotriose, cellobiose 및 cellotriose가 검출되었다. 올리고당(OSC, OSH)의 산화방지 활성을 측정한 결과, OSC는 분해시간이 증가할수록 산화방지 활성은 감소한 반면 OSH는 증가하였다. 또한 메밀껍질로부터 생산된 올리고당 첨가 시, 모든 비피더스 균주의 생육이 control에 비해 증가하여 프리 바이오틱 효과가 있음을 알 수 있었다. 이상의 연구결과, 효소분해에 의해 메밀껍질로부터 올리고당을 생산할 수 있었으며, 산화방지와 프리바이오틱 효과가 있음을 확인하였다. 따라서 목적에 따라 효소의 처리시간 및 방법을 다양화한다면 기능성식품으로의 활용이 더 높아질 것으로 예상된다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제1권1호
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• pp.46-50
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• 1996

In order to design industrial scale reactors and proceises for multi-phase biocatalytic reactions, it is essential to understand the mechanisms by which such systems operate. To il-lustrate how such mechanisms can be modeled, the hydrolysis of the primary ester groups of triglycerides to produce fatty acids and monoglycerides by lipased (glycerol-ester hydrolase) catalysis has been selected as an example of multiphase biocatalysis. Lipase is specific in its behavior such that it can act only on the hydrolyzed (or emulsified) part of the substrate. This follows because the active center of the enzyme is catalytically active only when the substrate contacts it in its hydrolyzed form. In other words, lipase acts only when it can shuttleback and forth between the emulsion phase and the water phase, presumably within an interphase or boundary layer between these two phases. In industrial applications lipase is employed as a fat splitting enzyme to remove fat stains from fabrics, in making cheese, to flavor milk products, and to degrade fats in waste products. Effective use of lipase in these processes requires a fundamental understanding of its kinetic behavior and interactions with substrates under various environmental conditions. Therefore, this study focuses on modeling and simulating the enzymatic activity of the lipase as a step towards the basic understanding of multi-phase biocatalysis processes.

• KSBB Journal
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• 제26권3호
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• pp.243-247
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• 2011

Using Bacillus subtilis spore display system, with cotG as an anchoring motif, levansucrase from Zymomonas mobilis, was displayed on the outer surface of Bacillus subtilis spore. Flow cytometry of DB104 (pSDJH-cotG-levU) spore, proved the surface localization of CotG-LevU fusion protein on the spore compared to that of DB104. Enzymatic activity of DB104 (pSDJH-cotG-levU) spore showed more than 1.5 times higher levansucrase specific activity compared to that of the host spore, which is a remarkable increase of enzymatic activity considering the existence of sacA (sucrase) and sacB (levansucrase) in the Bacillus subtilis chromosome. The spore integrity, revealed by sporulation frequency test after heat and lysozyme treatment of spore, did not changed at all in spite of the CotG-LevU fusion protein incorporation into the spore coat layer during spore formation process. These data prove again that Bacillus subtilis spore could be considered as good live immobilization vehicle for efficient bioconversion process.

• BMB Reports
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• 제32권6호
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• pp.541-546
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• 1999

When we compared the chitinase activity of various plant sources using colorimetric or active gel-staining assay methods, the specific activity of pumpkin leaves was the highest among the samples we analyzed. The highly active chitinase from pumpkin leaves (designated PL-ChtIII) was purified to homogeneity using affinity chitin gel and HPLC Mono-Q anion-exchange cloumn chromatographies. In contrast to other members of the class III chitinase family, PL-ChtIII showed a strong binding affinity to the regenerated chitin gel column. The apparent molecular weight of PL-ChtIII was estimated to be 29 kDa on SDS-PAGE gel, while its optimum pH and temperature were shown to be pH 6.0 and $60^{\circ}C$, respectively. Analyzing the reaction products of PL-ChtIII with swollen chitin as substrate, the dimer and tetramer of N-acetylglucosamine were produced as major products in the first hour of the enzymatic reaction along with a small amount of monomers and trimers. As the reaction time increased, dimeric N-acetylglucosamine became the predominant form of reaction product.

• 한국미생물·생명공학회지
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• 제47권4호
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• pp.546-554
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• 2019

This study, for the first time, reports the functional expression of lipase B derived from the yeast Candida antarctica (CALB) in Corynebacterium strain using the Escherichia coli plasmid PK18. The CALB gene fragment encoding a 317-amino-acid protein was successfully obtained from the total RNA of C. antarctica. CALB was readily produced in the Corynebacterium strain without the use of induction methods described in previous studies. This demonstrated the extracellular production of CALB in the Corynebacterium strain. CALB produced in the Corynebacterium MB001 strain transformed with pEC-CALB recombinant plasmid exhibited maximum extracellular enzymatic activity and high substrate affinity. The optimal pH and temperature for the hydrolysis of 4-nitrophenyl laurate by CALB were 9.0 and 40℃, respectively. The enzyme was stable at pH 10.7 in the glycine-KOH buffer and functioned as an alkaline lipase. The CALB activity was inhibited in the presence of high concentration of Mg²⁺, which indicated that CALB is not a metalloenzyme. These properties are key for the industrial application of the enzyme.

• Journal of Microbiology and Biotechnology
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• 제24권12호
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• pp.1699-1706
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• 2014

A lichenase gene (mt-lic) was identified for the first time through function-based screening of a soil metagenomic library. Its deduced amino acid sequence exhibited a high degree of homology with endo-${\beta}$-1,3-1,4-glucanase (having both lichenase and chitosanase activities), encoded by the bgc gene of Bacillus circulans WL-12. The recombinant lichenase overexpressed and purified from Escherichia coli was able to efficiently hydrolyze both barley ${\beta}$-glucan and lichenan. The enzyme showed maximal activity at a pH of 6.0 at $50^{\circ}C$, with Azo-barley-glucan as the substrate. The metal ions $Mn^{2+}$, $Mg^{2+}$, $Ca^{2+}$, and $Fe^{2+}$ enhanced the enzymatic activity, whereas the $Cu^{2+}$ and $Zn^{2+}$ ions inhibited the enzymatic activity. The $K_m$ and $V_{max}$ values of the purified lichenase were determined to be 0.45 mg/ml and 24.83 U/min/mg of protein, respectively.

• ALGAE
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• 제25권1호
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• pp.45-56
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• 2010

In this study, we assessed the antioxidant properties of tidal pool microalgae, Halochlorococcum porphyrae and Oltamannsiellopsis unicellularis, from Jeju Island, Korea. Specifically, the antioxidant activity of fractions isolated from 80% methanol extract, and digests produced from five proteases and carbohydrases, were investigated. Almost all the fractions and the 80% methanol extract exhibited higher effects on 1,1-Diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging. The ethyl acetate fraction showed the highest superoxide anion scavenging activity, while both n-hexane and chloroform fractions exhibited higher $H_2O_2$ scavenging activity. Among the enzymatic digests from H. porphyrae and O. unicellularis, all the digests exhibited remarkable DPPH scavenging activities. In nitric oxide inhibition, all the digests recorded significantly higher effects than those of the commercial antioxidants (p < 0.05). Flavozyme and Neutrase digests from H. porphyrae, and Termamyl and Alcalase digests from O. unicellularis, showed significant effects in metal chelating. Lipid peroxidation was significantly inhibited in the ethyl acetate fraction, in the Celluclast and Protamex digests from H. porphyrae, and in the chloroform fraction from O. unicellularis. These findings suggest that the two tidal pool microalgae tested in this study are rich in potential antioxidative compounds, the specific properties of which can be considered for use in the food and pharmaceutical industries.

• KSBB Journal
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• 제31권2호
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• pp.134-143
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• 2016

Lycii fructus has been traditionally used as a preventive and therapeutic medicine to treat enervation and diverse chronic diseases. In this study, we investigated whether fermentation of Lycii fructus extract (LE) increases the enzymatic activity of the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH). The fermentation of LE by Bacillus subtilis subsp. subtilis and Saccharomyces cerevisiae IFO 2376 was shown to increase the enzymatic activity of ADH and ALDH. TLC analysis of LE and fermented LE (FLE) showed that ADH and ALDH activities increased in different spots. Fraction No. 66 of LE and fraction No. 68 of FLE by Silica gel chromatography showed increased ADH activity of 129.1% and 148.9%, respectively. Fractions No. 128 of LE and FLE by Silica gel chromatography showed increased ALDH activity of 134.1% and 148.1%, respectively. The fraction No. 68 of FLE obtained by HPLC showed new peaks at $R_t$ 11.938min, $R_t$ 22.072min and $R_t$ 28.842min, indicating that ADH activity was increased. The LE and FLE fractions with the greatest increases in ADH activity peaked at the same time ($R_t$ 13min),whereas the LE and FLE fractions with the greatest increases in ALDH activity peaked at different times ($R_t$ 16.307min and $R_t$ 36.640min, respectively).

• Toxicological Research
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• 제12권2호
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• pp.155-161
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• 1996

The enzymatic properties for NADPH-P450 reductase domain of a fusion protein between human cytochrome P450 1A1 and rat NADPH-P450 reductase expressed in Escherichia coli were investigated. The fusion plasmid pCW/1A1OR-expressed E. coli membrane showed high NADPH-cytochrome c reductase activity ($830.1\pm 85.8 nmol\cdot min^{-1}\cdot mg protein^{-1}$), while pCW control vector and P 450 1A1 expression vector pCW/1A1 showed relatively quite low activity ($4.35\pm 0.49, 3.27\pm 0.50 nmol\cdot min^{-1}\cdot mg protein^{-1}$, respectively). The kinetic curves for NADPH-cytochrome c reductase followed typical Michaelis-Menten kinetics. The $K_{max}$ and $V_{max}$ for NADPH-dependent reductase activity were $8.24\pm 2.61\mu $and $817.9\pm 60.8 nmol\cdot min^{-1}\cdot mg protein^{-1}$, respectively, whereas those for cytochrome c-dependent reductase activity were $19.97\pm 2.86\mu M$ and $1303.5\pm 67.1 nmol\cdot min^{-1}\cdot mg protein^{-1}$. The reductase activities were also compared with those of rat, porcine and human liver microsomes. The activity of pCW/ 1A1OR-expressed E. coli membrane was 15.2-fold higher than that of rat liver microsome. Treatment with benzo(a)pyrene, 7-ethoxyresorufin and $\alpha$-naphthofiavone which are known as specific substrates or inhibitor for human P450 1A1 increased NADPH-cytochrome c reductase activity of fusion protein in E. coli membrane dose-dependently. These results demonstrate that the membrane topology of fused enzyme may be important for activity of its NADPH-P450 reductase domain.

• Journal of Oral Medicine and Pain
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• 제34권3호
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• pp.227-235
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• 2009

타액의 보호작용은 주로 타액 당단백질의 생물학적, 물리적, 구조적 성질 및 유동학적 성질과 관련이 있다. 그러므로 이상적인 타액 대체제의 개발을 위해서는 인체 타액의 생물학적 성질 뿐만 아니라 유동학적 특성을 이해하여야 한다. 본 연구의 목적은 타액 대체제가 인체 타액에 존재하는 효소의 활성에 미치는 영향을 파악하고 다양한 타액 대체제의 점도와 인체 타액의 점도를 비교하기 위해서 시행되었다. Moi-Stir, Stoppers4, MouthKote, Saliva Orthana 및 서울대학교치과병원 타액 대체제(SNU)를 사용하였으며, lysozyme 활성은 turbidimetric 법으로, peroxidase 활성은 NbsSCN 법으로, $\alpha$-amylase 활성은 maltotriose와 결합된 2-chloro-p-nitrophenol를 사용하여 시행하였다. 타액 대체제의 pH를 측정하였으며 cone-and-plate 형태의 점도계를 이용하여 다양한 전단율에서 점도를 측정하였다. 본 연구에 사용된 다양한 타액 대체제는 타액 효소 활성에 각기 다른 영향을 미쳤다. Stoppers4는 hen egg-white lysozyme, bovine lactoperoxidase (bLP) 및 $\alpha$-amylase 활성을 증가시켰고, Saliva Orthana와 SNU는 bLP 활성은 저해하였으며 $\alpha$-amylase 활성은 증가시켰다. MouthKote는 $\alpha$-amylase 활성을 저해하였으며, Moi-Stir는 bLP와 $\alpha$-amylase 활성을 저해하였다. 타액 대체제의 pH는 타액 대체제의 종류에 따라 매우 달랐다. Stoppers4, MouthKote 및 Saliva Orthana는 낮은 전단율에서는 인체 타액보다 낮은 점도를 높은 전단율에서는 인체 타액보다 높은 점도를 나타내었다. Moi-Stir와 SNU는 인체 타액보다 매우 높은 점도를 나타내었다. 결론적으로 본 연구결과는 각각의 타액 대체제는 각기 다른 생물학적 기능과 유동학적 특성을 가지고 있음을 알 수 있다. 타액 대체제의 사용은 사용하는 타액 대체제의 종류에 따라 타액 효소 활성에 각기 다른 영향을 미치고 궁극적으로는 구강건강에 다른 영향을 미칠 수 있을 것이다.