• Korean Journal of Food Science and Technology
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• v.48 no.3
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• pp.201-207
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• 2016

This study was conducted to produce oligosaccharides from buckwheat hull by using commercial enzymes. Yields of oligosaccharides obtained by enzymatic hydrolysis of the cellulose and hemicellulose fractions were 132.37 and 393.04 g/kg, respectively. Xylose, glucose, fructose, xylobiose, xylotriose, cellobiose, and cellotriose were detected in the hydrolysate produced from buckwheat hull. Antioxidant activity of oligosaccharide from cellulose fraction (OSC) reduced with increasing hydrolysis time; however, the antioxidant activity of oligosaccharide from hemicellulose fraction (OSF) increased as the hydrolysis time was prolonged. OSF and OSC showed higher increase in viable counts compared to the control. As a result, oligosaccharides produced from buckwheat hull by enzymatic hydrolysis showed antioxidant activity and prebiotic effects. It is suggested that utilization of oligosaccharides produced from buckwheat hull as functional food materials may be improved when hydrolysis time and conditions are controlled for this purpose.

• Biotechnology and Bioprocess Engineering:BBE
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• v.1 no.1
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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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• v.26 no.3
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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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• v.32 no.6
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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.

• Microbiology and Biotechnology Letters
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• v.47 no.4
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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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• v.24 no.12
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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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• v.25 no.1
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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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• v.31 no.2
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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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• v.12 no.2
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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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• v.34 no.3
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• pp.227-235
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• 2009

Many of the protective functions of saliva can be attributed to the biological, physical, structural, and rheological characteristics of salivary glycoproteins. Therefore, the development of ideal saliva substitutes requires understanding of the rheological as well as biological properties of human saliva. In the present study, we investigated the changes of salivary enzymatic activities by saliva substitutes and compared viscosity of saliva substitutes with human saliva. Five kinds of saliva substitutes such as Moi-Stir, Stoppers4, MouthKote, Saliva Orthana, and SNU were used. Lysozyme activity was determined by the turbidimetric method. Peroxidase activity was determined with an NbsSCN assay. $\alpha$-Amylase activity was determined using a chromogenic substrate, 2-chloro-p-nitrophenol linked with maltotriose. The pH values of saliva substitutes were measured and their viscosity values were measured with a cone-and-plate digital viscometer at six different shear rates. Various types of saliva substitutes affected the activities of salivary enzymes in different ways. Stoppers4 enhanced the enzymatic activities of hen egg-white lysozyme, bovine lactoperoxidase (bLP), and $\alpha$-amylase. Saliva Orthana and SNU inhibited bLP activity and enhanced $\alpha$-amylase activity. MouthKote inhibited $\alpha$-amylase activity. Moi-Stir inhibited the enzymatic activities of bLP and $\alpha$-amylase. The pH values were very different according to the types of saliva substitutes. Stoppers4, MouthKote, and Saliva Orthana showed lower values of viscosity at low shear rates and higher values of viscosity at high shear rates compared with unstimulated and stimulated whole saliva. Moi-Stir and SNU displayed much higher values of viscosity than those of natural whole saliva. Collectively, our results indicate that each saliva substitute has its own biological and rheological characteristics. Each saliva substitute affects the enzymatic activity of salivary enzyme and finally oral health in different ways.