• Journal of Microbiology and Biotechnology
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• 제28권2호
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• pp.284-292
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• 2018

A novel ${\beta}$-agarase, AgaJ5, was identified from an agar-degrading marine bacterium, Gayadomonas joobiniege G7. It belongs to the glycoside hydrolase family 86 and is composed of 805 amino acids with a 30-amino-acid signal peptide. Zymogram analysis showed that purified AgaJ5 has agarase activity. The optimum temperature and pH for AgaJ5 activity were determined to be $30^{\circ}C$ and 4.5, respectively. AgaJ5 was an acidic ${\beta}$-agarase that had strong activity at a narrow pH range of 4.5-5.5, and was a cold-adapted enzyme, retaining 40% of enzymatic activity at $10^{\circ}C$. AgaJ5 required monovalent ions such as $Na^+$ and $K^+$ for its maximum activity, but its activity was severely inhibited by several metal ions. The $K_m$ and $V_{max}$ of AgaJ5 for agarose were 8.9 mg/ml and 188.6 U/mg, respectively. Notably, thin-layer chromatography, mass spectrometry, and agarose-liquefication analyses revealed that AgaJ5 was an endo-type ${\beta}$-agarase producing neoagarohexaose as the final main product of agarose hydrolysis. Therefore, these results suggest that AgaJ5 from G. joobiniege G7 is a novel endo-type neoagarohexaose-producing ${\beta}$-agarase having specific biochemical features that may be useful for industrial applications.

• Journal of Microbiology and Biotechnology
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• 제7권2호
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• pp.114-120
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• 1997

Microorganisms producing xylanase were screened for the enzymatic production of xylo-oligo saccharides from xylan. One of the bacteria isolated from compost produced an endoxylanase extracellularly. The bacterium was identified as Bacillus sp. according to its taxonomic characteristics examined. Xylanase production reached upto 5 U/ml after 22 h of culture in LB medium at $30^{\circ}C$. The xylanase was purified by ammonium sulfate precipitation and gel filtration. The molecular weight of the xylanase was estimated to be 20,400 by SDS-PAGE. Optimal temperature and pH for the xylanase activity was $60^{\circ}C$ and 6.5, respectively. The enzyme was stable at temperatures upto $40^{\circ}C$ and pH values from 4 to 10. The xylanase was completely inhibited by the addition of 2 mM mercury ion. Apparent $K_m$ and $V_max$ values for oat spelt xylan were 9.2 mg/ml and 1954 U/mg protein, respectively. For birchwood xylan, the values were 6.3 mg/ml and 1009 U/mg protein. The predominant products of the xylan hydrolysis were xylobiose, xylotriose and xylotetraose, indicating that the enzyme is an endoxylanase. Upto $85{\%}$ of the initially added enzyme (2 U/ml) was bound to 50 mg/ml of the insoluble fraction of oat spelt xylan after incubation at $30^{\circ}C$ for 30 min.

• Journal of Microbiology and Biotechnology
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• 제17권9호
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• pp.1521-1526
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• 2007

A maltogenic amylase gene from Lactobacillus gasseri ATCC33323 (LGMA) was expressed in Lactococcus lactis MG1363 using the P170 expression system. The successful production of recombinant LGMA (rLGMA) was confirmed by the catalytic activity of the enzyme in liquid and solid media. The N-terminal amino acid sequencing analysis of the rLGMA showed that it was Met-Gln-Leu-Ala-Ala-Leu-, which was the same as that of genuine protein, meaning the signal peptide was efficiently cleaved during secretion to the extracellular milieu. The optimal reaction temperature and pH of rLGMA ($55^{\circ}C$ and pH 5, respectively) and enzymatic hydrolysis patterns on various substrates (${\beta}$-cyclodextrin, starch, and pullulan) supported that rLGMA was not only efficiently secreted from the Lactococcus lactis MG1363 but was also functionally active. Finally, the branched maltooligosaccharides were effectively produced from liquefied com starch, by using rLGMA secreted from Lactococcus lactis, with a yield of 53.1%.

• Journal of Microbiology and Biotechnology
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• 제18권1호
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• pp.160-166
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• 2008

A DNA fragment containing 2,079 base pairs from Bacillus circulans CGMCC 1416 was cloned using degenerate PCR and inverse PCR. An open reading frame containing 981 bp was identified that encoding 326 amino acids residues, including a putative signal peptide of 31 residues. The deduced amino acid sequence showed the highest identity (68.1%) with $endo-{\beta}-1,4-D-mannanase$ from Bacillus circulans strain K-1 of the glycoside hydrolase family 5 (GH5). The sequence encoding the mature protein was cloned into the pET-22b(+) vector and expressed in Escherichia coli as a recombinant fusion protein containing an N-terminal hexahistidine sequence. The fusion protein was purified by $Ni^{2+}$ affinity chromatography and its hexahistidine tag cleaved to yield a 31-kDa ${\beta}$-mannanase having a specific activity of 481.55U/mg. The optimal activity of the purified protein, MANB48, was at $58^{\circ}C$ and pH 7.6. The hydrolysis product on substrate locust bean gum included a monosaccharide and mainly oligosaccharides. The recombinant MANB48 may be of potential use in the feed industry.

• Journal of Microbiology and Biotechnology
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• 제14권4호
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• pp.715-721
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• 2004

Many food protein hydrolysates have been shown to have antioxidant activities, and recent research focuses on low molecular peptides produced during hydrolysis of food protein. Korean rice wine contains about 60-70% of protein at dry base and originates from raw materials. It has been suggested that the protein is transformed into low molecular weight peptides, and have antioxidant activity during fermentation. The objectives of this study were to evaluate the antioxidant activity of the pre-purified and purified peptides found in Korean rice wine and to identify the responsible peptides. The wine extract of Samhaeju, a traditional Korean rice wine made by low temperature fermentation, was evaporated at $35^{\circ}C$. The two methods employed in the evaluation of antioxidant activity were the DPPH radical scavenging method and the beta-carotene bleaching test. The pre-purified samples showed 808 AAC (Antioxidant Activity Coefficient) and 56.5% AOA (Antioxidant Activity), which were higher than $\alpha$-tocopherol (572 AAC and 78% AOA). The rice wine extract was separated by reversed-phase HPLC. The protective effect of the four most antioxidant active fractions were tested for t-butyl hydroperoxide induced oxidation of healthy human erythrocytes and the byproduct was determined by malondialdehyde formation. Fraction No.5 showed 35% lower MDA concentration as compared to the control. The peptides were further purified using consecutive chromatographic methods and 4 antioxidant peptides were isolated. The amino acid sequences of the peptides were identified as Ile-His-His, Val- Val-His(Asn), Leu-Val-Pro, and Leu(Val)-Lys-Arg-Pro. The AAC value of the synthetic form of the identified peptides was the highest for Ile-His-His.

• Journal of Microbiology and Biotechnology
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• 제16권2호
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• pp.272-279
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• 2006

The gene encoding Thermus sp. T351 alkaline phosphatase (T351 APase) was cloned and sequenced. The gene consisted of 1,503 bp coding for a protein with 500 amino acid residues including a signal peptide. The deduced amino acid sequence of T351 APase showed relatively low similarity to other Thermus APases. The T351 APase gene was expressed under the control of the T7lac promoter on the expression vector pET-22b(+) in Escherichia coli BL21 (DE3). The expressed enzyme was purified by heat treatment, and $UNO^{TM}$ Q and $HiTrap^{TM}$ Heparin HP column chromatographies. The purified enzyme exhibited high activity at extremely alkaline pHs, reaching a maximum at pH 12.0. The optimum temperature of the enzyme was $80^{\circ}C$, and the half-life at $85^{\circ}C$ was approximately 103 min. The enzyme activity was found to be dependent on metal ions: the addition of $Mg^{2+}$ and $CO^{2+}$ increased the activity, whereas EDTA inhibited it. With p-nitrophenyl phosphate as the substrate, T351 APase had a Michaelis constant ($K_{m}$) of $3.9{\times}10^{-5}M$. The enzyme catalyzed the hydrolysis of a wide variety of phosphorylated compounds.

• 한국식품과학회지
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• 제25권5호
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• pp.571-575
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• 1993

대두 펩타이드의 표면소수도가 혈청 콜레스테롤의 농도에 미치는 영향을 조사하기 위하여, 대두 단백질(ISP), 카세인(CNP), 이들 단백질을 펩신으로 가수분해하여 pH에 따른 펩타이드 침전 획분들(SHT, SH8, SH6, SH4, CHT, CH6, CH5, CH4)을 흰쥐에 섭취시키고 혈청 콜레스테롤 농도 및 분변 스테로이드이 배설량을 측정하였다. 그리고 각 펩타이드의 표면소수도를 ANS 형광법 및 SDS 결합법으로 측정하여, 이들의 상관관계를 분석한 결과, 펩타이드의 ANS 표면소수도가 높아질수록 분변으로 배설된 스테로이드량은 증가하였으며(r=0.81), 혈청 콜레스테롤 농도는 낮아졌다.(r=-0.868). 그러나 SDS 표면소수도는 그들과 상관관계가 없었다. 또한 대두 단백질은 가수분해에 의하여 ANS 표면소수도가 증가하였다. 이상의 결과는 흰쥐의 담즙염이, 소화중 생성된 높은 표면소수도의 펩타이드와 결합하여 체외로 배출되므로서 대두단백질의 섭취에 의한 혈청 콜레스테롤 농도가 낮아짐을 시사하였다.

• 상하수도학회지
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• 제31권5호
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• pp.389-395
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• 2017

In this paper, we investigate the characteristics of membrane fouling caused by water temperature in the Membrane bioreactor(MBR) process and try to derive the membrane fouling control by chemical enhanced backwashing(CEB). The extracellular polymeric substances(EPS) concentration was analyzed according to the water temperature in the MBR, and the membrane fouling characteristics were investigated according to the conditions, with sludge & without sludge, through a lab-scale reactor. As shown in the existing literature the fouling resistance rate was increased within sludge with the water temperature was lowered. However, in the lab-scale test using the synthetic wastewater, the fouling resistance increased with the water temperature. This is because that the protein of the EPS was more easily adsorbed on the membrane surface due to the increase of entropy due to the structural rearrangement of the protein inside the protein as the water temperature increases. In order to control membrane fouling, we tried to derive the cleaning characteristics of CEB by using sodium hypochlorite(NaOCl). We selected the condition with the chemicals and the retention time, and the higher the water temperature and the chemical concentration are the higher the efficiencies. It is considered that the increasing temperature accelerated the chemical reaction such as protein peptide binding and hydrolysis, so that the attached proteinaceous structure was dissolved and the frequency of the reaction collision with the protein with the chemical agent becomes higher. These results suggest that the MBRs operation focus on the fouling control of cake layer on membrane surface in low temperatures. On the other hand, the higher the water temperature is the more the operation strategies of fouling control by soluble EPS adsorption are needed.

• 한국식품과학회지
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• 제21권4호
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• pp.492-497
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• 1989

천연식품성분에 의한 항산화기작을 해명하기 위하여 어육단백질, 탈지대두박, egg albumin 및 casein등의 단백질 가수분해물을 사용하여 linoleic acid 와의 모델계 반응을 통하여 이들의 항산화능과 ${\alpha}-tocopherol$과의 상승작용 및 금속이온 봉쇄능 등에 대하여 연구 검토하였는데 그 결과 각 단백질 가수분해물은 항산화능이 매우 큰 것으로 나타났으며. 특히 egg albumin가수분해물의 항산화능이 매우 강한 것으로 나타났다. 또한 천연항산화제인 ${\alpha}-tocopherol$과의 상승작용도 우수한 것으로 나타났는데 단백질 가수분해물간의 차이는 거의 없는 것으로 나타났다. 금속이온 봉쇄능 또한 높게 나타났으며 단백질 가수분해물의 금속이온 봉쇄능은 금속이온의 종류에 따라 다소 차이가 있는 것으로 나타났다.

• 한국축산식품학회지
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• 제37권1호
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• pp.62-70
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• 2017

The aim of this study was identifying a suitable food grade enzymes to hydrolyze whey protein concentrates (WPCs), to give the highest bioactivity. WPCs from ultrafiltration retentate were adjusted to 35% protein (WPC-35) and hydrolyzed by enzymes, alcalase, ${\alpha}-chymotrypsin$, pepsin, protease M, protease S, and trypsin at different hydrolysis times (0, 0.5, 1, 2, 3, 4, and 5 h). These 36 types of hydrolysates were analyzed for their prominent peptides ${\beta}-lactoglobulin$ (${\beta}-Lg$) and ${\alpha}-lactalbumin$ (${\alpha}-La$), to identify the proteolytic activity of each enzyme. Protease S showed the highest proteolytic activity and angiotensin converting enzyme inhibitory activity of IC50, 0.099 mg/mL (91.55%) while trypsin showed the weakest effect. Antihypertensive and antioxidative peptides associated with ${\beta}-Lg$ hydrolysates were identified in WPC-35 hydrolysates (WPH-35) that hydrolyzed by the enzymes, trypsin and protease S. WPH-35 treated with protease S in 0.5 h, responded positively to usage as a bioactive component in different applications of pharmaceutical or related industries.