• 한국미생물·생명공학회지
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• 제31권3호
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• pp.284-291
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• 2003

본 연구는 우리나라의 전통발효식품인 재래간장에서도 혈전용해효소 생산 미생물이 존재할 가능성이 있다고 판단하여, 지역의 재래간장으로부터 혈전용해효소 생산능이 우수한 균주 K42를 최종 선발하였다. 선발된 균주는 B. amyloliquefaciens로 동정되었으며, l2% NaCl 함유된 배지에서도 균생육이 양호하였으며 10% NaCl 농도까지 효소생산이 유지되는 내염성 세균임을 알 수 있었다. 혈전용해효소의 정제는 40% ammonium sulfate로 분별 염석 후 DEAE-Sephadex A-50, Sephadex G-100, Sephadex G-75를 이용하여 단일단백의 효소로 정제하였다. 효소의 활성도는 배양여액에 비해 17.1배 증가하였고, 회수율은 1.1%로 나타났다. Disc-electrophoresis와 SDS-PAGE를 실시하여 분자량 45,000 Dalton의 단일 단백질로 확인하였다. 혈전용해효소의 특성조사에 있어서는 최적 pH는 8.0의 약알칼리성을 나타내었고 안정성에 있어서는 pH 4.0에서 pH 10.0 범위에서 80% 이상의 잔존활성을 나타내었으며, 최적반응 온도는 $50^{\circ}C$로 조사되었고 열 안정성에 있어서는 3$0^{\circ}C$의 온도에서 50분간 열처리시 80%이상 잔존 활성을 보였다. 금속이온of B. amyloliquefaciens K42가 생산하는 fibrinolytic enzyme의 활성에 미치는 영향을 검토한 결과 $Mg^{2+}$ $Cu^{ 2+}$에 대해서는 효소 활성이 증가하는 양상을 보였다. 또한 B. amyioliquefaciens K42가 생산하는 fibrinolytic enzyme은 metallo enzyme의 활성저해제인 EDTA등에 대해서는 15%이하로 효소 활성이 감소되었는데 이러한 효소 활성 저해로 보아 metallo enzyme이거나 활성에 금속이온이 필요한 것으로 추정된다. 그리고 기질인 fibrin과의 친화성을 알아보기 위해 Km값을 측정하였는데 Km값은 2.03 mg/ml 나타났다.

• 한국미생물·생명공학회지
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• 제26권6호
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• pp.507-514
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• 1998

국내 전통식품인 청국장으로부터 강력한 fibrinolytic activity를 갖는 K-54주를 분리, 동정한 결과 B. subtilis로 확인되었으며 이 균으로부터 강력한 갖는 효소를 정제하였다. 정제도는 약 66.9배 증가하였으며 약 10.1%의 수율을 보였다. B. subtilis K-54에서 분리한 fibrinolytic enzyme의 최적 활성 pH가 8-12사이였으며 50-75$^{\circ}C$사이에서 최적 활성을 보였다. 정제된 효소의 분자량은 약 29 kDa이었고, 등전점은 8.67로 나타났으며 Vmax값은 47.16%, Km값은 0.31(3.1 mg/$m\ell$)였다. N-말단 아미노산 서열 분석 결과 B. subtilis K-54주에서 정제한 fibrinolytic enzyme는 Ala-Gly-Ser-Val-Pro-Try-Gly-Ser이었으며 serine protease저해제인 PMSF에 강력한 활성 저해를 보여 serine protease계열의 효소임을 확인할 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제24권2호
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• pp.245-253
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• 2014

A fibrinolytic enzyme was produced by an edible mushroom of Pleurotus ostreatus using submerged culture fermentation. The enzyme was purified from the culture supernatant by applying a combination of freeze-thaw treatment, ammonium sulfate precipitation, hydrophobic interaction, and gel filtration chromatographies. The enzyme was purified by a 147-fold, with a yield of 7.54%. The molecular masses of the enzyme an determined by gel filtration and SDS-PAGE were 13.6 and 18.2 kDa, respectively. The isoelectric point of the enzyme was 8.52. It hydrolyzed fibrinogen by cleaving the ${\alpha}$ and ${\beta}$ chains of fibrinogen followed by the ${\gamma}$ chains, and also activated plasminogen into plasmin. The enzyme was optimally active at $45^{\circ}C$ and pH 7.4. The enzyme activity was completely inhibited by EDTA, whereas protease inhibitors of TPCK, SBTI, PMSF, aprotinin and pepstatin showed no inhibition on its activity. The partial amino acid sequences of the enzyme as determined by Q-TOF2 were ATFVGCSATR, GGTLIHESSHFTR, and YTTWFGTFVTSR. These sequences showed a high degree of homology with those of metallo-endopeptidases from P. ostreatus and Armillaria mellea. The purified enzyme can also be applied as a natural agent for oral fibrinolytic therapy or prevention of thrombosis.

• Korean Chemical Engineering Research
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• 제53권6호
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• pp.667-671
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• 2015

Anode는 효소를 이용한 효소전극과 cathode는 PEMFC용 전극을 이용해 효소연료전지를 구동하였다. 효소 anode는 graphite 분말과 효소로서 글루코스 산화제, 전자매개체로 ferrocene을 혼합해 압축해서 만들고 Nafion 이오노머로 코팅하였다. Anode 제조조건을 변화시키며 성능을 측정해 효소 anode 제조 최적조건을 찾았다. 효소 anode 압축 시 최적 압력은 8.89 MPa이고, 효소 anode의 graphite 성분비가 60%일 때 최고의 출력밀도를 나타냈다. Anode 기질 용액의 최적 glucose 농도는 1.7mol/l이었다. 효소 anode는 Nafion 용액에 1초, 2회 침지에 의해 안정화되었다.

• Journal of the Korean Wood Science and Technology
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• 제39권2호
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• pp.187-195
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• 2011

In this study, dilute acid pretreatment of $Liriodendron$ $tulipifera$ was performed for enzymatic hydrolysis. As the pretreatment temperature was increased, enzymatic hydrolysis and enzyme adsorption yield also increased. The highest enzymatic hydrolysis yield was 57% (g/g) and enzyme adsorption was 44% (g/g). Enzymatic hydrolysis yield was determined with weight loss of pretreated biomass by enzyme, and enzyme adsorption was a percentage of enzyme weight attaching on pretreated biomass compared with input enzyme weight. When $L.$ $tulipifera$ was pretreated with 1% sulfuric acid at $160^{\circ}C$ for 5 min., hemicellulose was significantly removed in pretreatment, but the lignin contents were constant. Other changes in surface morphology were detected on biomass pretreated at $160^{\circ}C$ by a field emission scanning electron microscope (FESEM). A large number of spherical shapes known as lignin droplets were observed over the entire biomass surface after pretreatment. Hemicellulose removal and morphological changes improved enzyme accessibility to cellulose by increasing cellulose exposure to enzyme. It is thus evidence that enzyme adsorption is a significant factor to understand pretreatment effectiveness.

• Journal of Microbiology and Biotechnology
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• 제11권5호
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• pp.845-852
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• 2001

A microorganism producing fibrinolytic enzyme was isolated from Korean traditional soybean paste and identified as Bacillus sp. KDO-13. The fibrinolytic enzyme was purified to homogeneity by ammonium sulfate fractionation, ion-exchange chromatography on DEAE-celluose, and gel chromatography on Sephadex G-100 of the culture supernatant of Bacillus sp. KDO-13. The molecular weight of the purified enzyme was estimated to be 44,000 by SDS-PAGE. The optimum pH and temperature for the enzyme activity were pH 8.0 and $50{\circ}C$, respectively. The enzyme activity was relatively stable at pH 7.0-9.0 and temperature below $50{\circ}C$. the activity of the enzyme was inhibited by $AI^{3+}$ and $Hg^{2+}$, but activated by $Co^{2+}$\;and\;Ni^{2+}. In addition, the enzyme activity was potently inhibited by EDTA and 0-phenanthroline. The purified enzyme could completely hydrolyze a fibrin substrate within 6 h in vitro, and had a low $K_m$ value for fibrin hydrolysis. It was concluded that the purified enzyme was a metalloprotease with relatively high specificity for fibrinolysis, and thus, could be applied as an effective thrombolytic agent.

• 한국균학회지
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• 제14권4호
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• pp.273-280
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• 1986

Rhizorus oryzae의 생장 및 autolysis에 따른 여러가지 autolytic enzymes의 활성도 변화와 이 autolytic enzyme system을 이용한 원형질체 생성에 관하여 조사하였다. Autolytic·phase의 culture filtrate 내에 함유된 autolytic enzyme은 Rh. oryzae 균사체로부터의 원형질체 생성에 효율적인 세포벽 분해효소로 작용하였으며 Autolytic enzyme중 원형질체 생성은 proteolytic 및 chitosanase activity와 가장 긴밀하게 관련되어 있었다. 이와같은 autolytic enzyme을 이용한 원형질체 생성은 10시간 배양한 균사체에 0.5M mannitol을 사용하였을 때 최고의 수율을 보였으며 원형질 생성의 최적온도 및 pH는 각각 $25{\sim}30^{\circ}C$ 및 $6.0{\sim}6.5$로 나타났다 한편 18시간 배양된 균사체를 osmotic stabilizer와 aut-olytic enzyme을 1 : 1이 되도록 처리하고 5시간 동안 $30^{\circ}C$에서 incubation하여 얻은 원형질체를 주사전자현미경으로 조사한 결과 효소에 의하여 가수분해되는 세포벽 주변으로부터 원형질체가 형성되는 것을 확인하였다.

• 한국미생물·생명공학회지
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• 제18권3호
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• pp.251-259
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• 1990

생전분 분해력이 강력한 glucoamylase를 생산하는 균주로서 Asp.usamii IAM 2185를 선정하였다. 밀기울배지에서의 효소생산의 최적 initial pH는 6.0-8.0, 최적 배양온도는 25-$30^{\circ}C$, 최적 배양시간은 72시간이고, 밀기울배지에 ammonuim nitrate와 albumin의 첨가는 효소의 생산을 약간 증가시켰다. 황산암모늄분획, CM-cellulose와 DEAE-cellulose column chromatography에 의하여 효소를 정제하였고, 정제효소의 specific activity는 34.3U/mg.protein, 수율은 10.3 이었다.

• 대한수의학회지
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• 제29권1호
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• pp.21-25
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• 1989

This experiment was carried out to determine the progesterone concentration of bovine plasma by liquid phase double antibody enzyme immunoassay. The optimum conditions of assay-system, enzyme conjugate and gelatin were invested. The sensitivity, recovery rate and reproducibility by this assay were also analyzed. The results obtained were as follows: 1. The suitable supplementation level of gelatin was 0.2%. As the gelatin level increased to 1%, coefficient variation of interassay was shown to be irregular. 2. The optimum dilution rate of enzyme conjugate was 30 times. Enzyme activity was greatly fluctuated depending on the minor condition of enzyme conjugate technique. Therefore, it was considered to be checked when determined. 3. The sensitivity of the assay was 12 pg/tube. 4. Recovery rate was decreased when the amount of sample was too little or too much, but the recovery rate was high as 97.8% when the amount of sample between 50 and $200{\mu}l$. 5. Mean intra-assay and inter-assay coefficient of variation was 4.5% and 5.9%, respectively. By using liquid phase double antibody enzyme immunoassay, progesterone in plasma can be detected, and also this assay system is applicable to study on physiological function of progesterone and to diagnosis of reproductive disorders.

• 한국식품과학회지
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• 제15권4호
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• pp.404-408
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• 1983

Malo-alcohol 발효(醱酵)에 관여하는 분열효모균(分裂酵母菌), Schizos-accharomyces japonicus var. japonicus St-3가 생성(生成)하는 malic enzyme(EC 1.1.1 40)의 몇가지 성질(性質)을 조사하였다. Malic enzyme의 생성(生成)은 배양(培養) 24시간(時間)에 최대(最大)에 이르렀고 효소반응(酵素反應)의 최적(最適) pH는 10.0, 온도(溫度)는 $25^{\circ}C$였다. 본(本) 효소(酵素)는 pH $7.0{\sim}8.4$에서 안정(安定)하였으며 $60^{\circ}C$, 10분간(分間) 열처리(熱처理)로 50% 실활(失活)되었다. $Mn^{2+}$ 첨가(添加)는 효소활성(酵素活性)을 촉진(促進)시켰으며 유기산(有機酸), 아마노산(酸) 및 ethanol의 첨가(添加)는 효소활성(酵素活性)에 아무런 영향(影響)이 없었다.