• Journal of Microbiology and Biotechnology
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• 제19권12호
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• pp.1547-1556
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• 2009

A novel thermostable $\alpha$-glucosidase (TtGluA) from Thermoanaerobacter tengcongensis MB4 was successfully expressed in E. coli and characterized. The TtgluA gene contained 2,253 bp, which encodes 750 amino acids. The native TtGluA was a trimer with monomer molecular mass of 89 kDa shown by SDS-PAGE. The purified recombinant enzyme showed hydrolytic activity on maltooligosaccharides, p-nitrophenyl-$\alpha$-D-glucopyranide, and dextrin with an exotype cleavage manner. TtGluA showed preference for short-chain maltooligosaccharides and the highest specific activity for maltose of 3.26 units/mg. Maximal activity was observed at $60^{\circ}C$ and pH 5.5. The half-life was 2 h at $60^{\circ}C$. The enzyme showed good tolerance to urea and SDS but was inhibited by Tris. When maltose with the concentration over 50 mM was used as substrate, TtGluA was also capable of catalyzing transglycosylation to produce $\alpha$-1,4-linked maltotriose and $\alpha$-1,6-linked isomaltooligosaccharides. More importantly, TtGluA showed exclusive regiospecificity with high yield to produce $\alpha$-1,6-linked isomaltooligosaccharides when the reaction time extended to more than 10 h.

• Journal of Microbiology and Biotechnology
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• 제24권6호
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• pp.771-780
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• 2014

A putative lipolytic enzyme gene, named as est9x, was obtained from a marine microbial metagenome of the South China Sea. Sequence analysis showed that Est9X shares lower than 27% sequence identities with the characterized lipolytic enzymes, but possesses a catalytic triad highly conserved in lipolytic enzymes of the ${\alpha}/{\beta}$ hydrolase superfamily. By phylogenetic tree construction, Est9X was grouped into a new lipase/esterase family. To understand Est9X protein in depth, it was recombinantly expressed, purified, and biochemically characterized. Within potential hydrolytic activities, only lipase/esterase activity was detected for Est9X, confirming its identity as a lipolytic enzyme. When using p-nitrophenol esters with varying lengths of fatty acid as substrates, Est9X exhibited the highest activity to the C2 substrate, indicating it is an esterase. The optimal activity of Est9X occurred at a temperature of $65^{\cric}C$, and Est9X was pretty stable below the optimum temperature. Distinguished from other salt-tolerant esterases, Est9X's activity was tolerant to and even promoted by as high as 4 M NaCl. Our results imply that Est9X is a unique esterase and could be a potential candidate for industrial application under extreme conditions.

• 한국산학기술학회논문지
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• 제10권1호
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• pp.194-199
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• 2009

우수한 항산화제로 알려진 astaxanthin을 함유한 H. pluvialis 균체로부터 작용기작이 상이한 exe형과 endo형의 단백질 분해 효소와 복합 다당류 분해 효소를 이용하여 식품용 haematococcus추출물을 효율적으로 제조할 수 있는 방법을 조사하였다. 상업용 단백질 분해 효소로는 Alcalase (endo형)와 Flavourzyme (exe형)을 복합 다당류 분해효소로는 Viscozyme을 사용하였다. 단백질 분해 효소는 endo형과 exe형을 병용하는 것이 추출물의 astaxanthin 함량을 증가시켰다. Viscozyme과 함께 2종류의 단백질 분해 효소를 사용하는 경우에는 Alcalase와 Flavourzyme을 병용하여 1차로 처리한 후 Viscozyme을 2차로 사용하는 2단계 가수분해 방법이 적절하였다. 이 조건에서 astaxanthin 함량은 효소를 사용하지 않은 대조구에 비하여 320% ($529{\mu}g/g{\rightarrow}2,256{\mu}g/g$) 이상 향상되었다. 또한 DPPH법으로 조사한 항산화 활성은 astaxanthin 함량에 비례하여 증가하였으며, 1차로 Alcalase와 Flavourzyme을 병용하여 처리한 후 2차로 Viscozyme을 사용하는 조건에서 가장 우수하였다.

• Journal of Microbiology and Biotechnology
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• 제23권10호
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• pp.1403-1412
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• 2013

Ethanol fuel production from lignocellulosic biomass is emerging as one of the most important technologies for sustainable development. To use this biomass, it is necessary to circumvent the physical and chemical barriers presented by the cohesive combination of the main biomass components, which hinders the hydrolysis of cellulose and hemicellulose into fermentable sugars. This study evaluated the hydrolytic capacity of enzymes produced by yeasts, isolated from the soils of the Brazilian Cerrado biome (savannah) and the Amazon region, on sugarcane bagasse pre-treated with $H_2SO_4$. Among the 103 and 214 yeast isolates from the Minas Gerais Cerrado and the Amazon regions, 18 (17.47%) and 11 (5.14%) isolates, respectively, were cellulase-producing. Cryptococcus laurentii was prevalent and produced significant ${\beta}$-glucosidase levels, which were higher than the endo- and exoglucanase activities. In natura sugarcane bagasse was pre-treated with 2% $H_2SO_4$ for 30 min at $150^{\circ}C$. Subsequently, the obtained fibrous residue was subjected to hydrolysis using the Cryptococcus laurentii yeast enzyme extract for 72 h. This enzyme extract promoted the conversion of approximately 32% of the cellulose, of which 2.4% was glucose, after the enzymatic hydrolysis reaction, suggesting that C. laurentii is a good ${\beta}$-glucosidase producer. The results presented in this study highlight the importance of isolating microbial strains that produce enzymes of biotechnological interest, given their extensive application in biofuel production.

• 한국균학회지
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• 제39권2호
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• pp.126-130
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• 2011

작물의 생육촉진 및 식물 진균병의 생물방제능을 동시에 나타내는 다기능성 미생물제제를 개발하고자 토양으로부터 분리하여 보관중인 세균 120종의 활성을 검토하였다. 그 중 siderophore를 생성하고 항진균 활성을 보이는 BS11-1, BS11-2, BS11-3를 선발하였다. 이들 균주는 cellulase, protease 같은 lytic enzyme을 생산하였으며 식물성장 촉진 호르몬중의 하나인 IAA를 생성하였다. 이들 선발균들에 의한 식물 생장 촉진을 조사한 결과, BS11-1, BS11-2, BS11-3 균 배양액 관주 시 고추 유묘의 생육을 132%, 122%, 120% 증가 시켰으며, BS11-1, BS11-2 균주의 경우 뿌리의 신장 및 생육이 촉진되었음을 확인 할 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제24권1호
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• pp.19-25
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• 2014

An extracellular agarase was purified from Bacillus sp. BI-3, a thermophilic agar-degrading bacterium isolated from a hot spring in Indonesia. The purified agarase revealed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with an apparent molecular mass of 58 kDa. The optimum pH and temperature of the agarase were 6.4 and $70^{\circ}C$, respectively. The activity of the agarase was stable at high temperatures, and more than 50% activity was retained at $80^{\circ}C$ for 15 min. Furthermore, the enzyme was stable in the pH range of 5.8-8.0, and more than 60% of the residual activity was retained. Significant activation of the agarase was observed in the presence of $K^+$, $Na^+$, $Ca^{2+}$, $Mg^{2+}$, and $Sr^{2+}$; on the other hand, $Ba^{2+}$, $Zn^{2+}$, $Cu^{2+}$, $Mn^{2+}$, $Co^{2+}$, $Fe^{2+}$, and EDTA inhibited or inactivated the enzyme activity. The components of the hydrolytic product analyzed by thin-layer chromatography showed that the agarase mainly produced neoagarobiose. This study is the first to present evidence of agarolytic activity in aerobic thermophilic bacteria.

• Applied Biological Chemistry
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• 제40권5호
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• pp.395-399
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• 1997

아밀로오스 함량이 다른 옥수수전분을 2.2N HCl로 산처리하여 산가수분해 특성 및 효소저항전분의 수율을 비교하였다. 산 가수분해경향은 아밀로오스 함량이 0%인 Amioca는 7일을 경계로, 그외의 전분은 4일을 경계로 2단계로 진행되었으며, 산가수분해 속도상수는 초기에 $4.01{\sim}9.21{\times}10^{-3}hr^{-1}$, 후기에는 $1.60{\sim}5.01{\times}10^{-3}hr^{-1}$로 아밀로오스 함량이 낮을수록 옥수수전분의 가수분해 속도가 빨랐다. 생전분의 X-선 회절양상은 아밀로오스 함량이 적은 Amioca, PFP, CMS(commercial maize starch)는 A형 , 고아밀로오스 함량인 Amaizo 5, Amylomaize VII은 B형의 전형적인 결정형을 보였으며, 산처리 후에도 모든 시료의 결정형은 그대로 유지하였으나 상대적인 결정화도는 증가하였다. 효소저항전분의 수율은 Amioca, CMS, Amylomaize VII이 각각 1.8%, 20.8%, 37.9%로 아밀로오스 함량이 높을수록 수율도 증가하였으며, 가열-냉각 횟수를 4회 반복한 후 분리한 산처리 한 옥수수전분의 효소저항전분의 수율은 1일 산처리한 CMS, Amylomaize VII이 각각 4.5%, 29.1%였으며 7일 산처리시에는 1.5%와 19.4%로 산처리에 의해 효소저항전분의 수율이 감소하였다.

• 한국식품과학회지
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• 제53권6호
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• pp.695-700
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• 2021

본 연구에서는 단백질 가수분해효소 0.1%와 1% Protamex를 사용하여 저분자 콜라겐을 제조하였다. 토종 닭발의 조단백질과 콜라겐의 함량은 일반 육계에 비해 높은 함량이 나타났다. 단백질 가수분해 효소농도와 반응시간이 증가할수록 낮은 분자량의 콜라겐을 얻을 수 있는 것으로 나타났다. 특히 1% Protamex로 7시간 처리한 시료가 1,000-5,000 Da의 저분자 콜라겐 함량이 55.6%로 나타났으며, 평균 분자량은 5,390 Da로 가장 낮은 분자량이 나타났다. 이는 단백질 가수분해효소 Protamex가 고분자 펩타이드 결합을 저분자 펩타이드로 분해했기 때문이다. 효소처리 콜라겐의 조직감은 고분자 펩타이드의 콜라겐이 저분자 펩타이드로 분해되어 gel을 형성하지 못하고 sol의 형태를 유지하였다. 효소농도와 효소반응시간이 증가할수록 콜라겐의 평균분자량은 작아지나 효소반응 5시간부터 평균분자량의 감소가 미미해지는 경향이 나타났다. 따라서 저분자 콜라겐 효소반응시간은 경제적으로 볼 때 5시간에서 7시간 사이가 적합하다고 할 수 있다. 이 연구결과는 향후 산업적 효소를 이용한 저분자 콜라겐 제조 및 식품 소재 활용의 기초자료로 이용할 수 있을 것이다.

• Journal of Microbiology and Biotechnology
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• 제32권4호
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• pp.447-457
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• 2022

Notoginsenoside R1 and ginsenoside Rg1 are the main active ingredients of Panax notoginseng, exhibiting anti-fatigue, anti-tumor, anti-inflammatory, and other activities. In a previous study, a GH39 β-xylosidase Xln-DT was responsible for the bioconversion of saponin, a natural active substance with a xylose group, with high selectivity for cleaving the outer xylose moiety of notoginsenoside R1 at the C-6 position, producing ginsenoside Rg1 with potent anti-fatigue activity. The optimal bioconversion temperature, pH, and enzyme dosage were obtained by optimizing the transformation conditions. Under optimal conditions (pH 6.0, 75℃, enzyme dosage 1.0 U/ml), 1.0 g/l of notoginsenoside R1 was converted into 0.86 g/l of ginsenoside Rg1 within 30 min, with a molar conversion rate of approximately 100%. Furthermore, the in vivo anti-fatigue activity of notoginsenoside R1 and ginsenoside Rg1 were compared using a suitable rat model. Compared with the control group, the forced swimming time to exhaustion was prolonged in mice by 17.3% in the Rg1 high group (20 mg/kg·d). Additionally, the levels of hepatic glycogen (69.9-83.3% increase) and muscle glycogen (36.9-93.6% increase) were increased. In the Rg1 group, hemoglobin levels were also distinctly increased by treatment concentrations. Our findings indicate that treatment with ginsenoside Rg1 enhances the anti-fatigue effects. In this study, we reveal a GH39 β-xylosidase displaying excellent hydrolytic activity to produce ginsenoside Rg1 in the pharmaceutical and food industries.

• Fisheries and Aquatic Sciences
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• 제25권2호
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• pp.76-89
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• 2022

The Lophius litulon liver was used as raw material for the extraction of fish oil via various extraction methods. The extraction rate by water extraction, potassium hydroxide (KOH) hydrolysis and protease hydrolysis were compared and the results revealed the protease hydrolysis extraction had a higher extraction rate with good protein-lipid separation as observed by optical microscope. Furthermore, subsequent experiments determined neutrase to be the best hydrolytic enzyme in terms of extraction rate and cost. The extraction conditions of neutrase hydrolysis were optimized by single-factor experiment and response surface analysis, and the optimal extraction rate was 58.40 ± 0.25% with the following conditions: enzyme concentration 2,000 IU/g, extraction time 1.0 h, liquid-solid ratio 1.95:1, extraction temperature 40.5℃ and pH 6.5. The fatty acids composition in fish oil from optimized extraction condition was composed of 19.75% saturated fatty acids and 80.25% unsaturated fatty acids. The content of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were 8.06% and 1.19%, respectively, with the ratio (6.77:1) surpassed to the recommendation in current researches (5:1). The results in this study suggest protease treatment is an efficient method for high-quality fish oil extraction from Lophius litulon liver with a satisfactory ratio of DHA and EPA.