• 대한화학회지
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• 제40권1호
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• pp.72-80
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• 1996

Chitinase를 생성하는 세균인 serratia marcescens JM을 해안 갯벌 시료로부터 분리하여, ammonium sulfate precipitation, affinity adsorption, hydroxylapatite와 Sephadex G-200 column chromatography를 통하여 정제하였다. 정제된 chitinase는 7.1% 회수율과 4.22의 정제도를 나타내었으며, 전기영동시 단일밴드를 얻을 수 있었고, SDS-PAGE에 의해 측정된 분량은 59,000으로 나타났다. 정제된 chitinase의 $K_m$과 $V_{max}$는 5.71mg/mL과 39.8 unit/mL로 나타났다. Chitinase의 최적활성 pH와 온도는 7과 50$^{\circ}C$였고 최적안정pH는 7.0이며 50$^{\circ}C$이하에서는 안정하였다. $Cu^{2+}\;Ca^{2+}$ 및 $Mg^{2+}$는 효소활성을 증가시켰으나 $Hg^{2+}$와 $I_2$는 효소 활성을 억제시켰다. 또한 cysteine은 효소활성을 증가시키나 EDTA, MIA, PCMB, 및 SDS는 효소활성을 억제시켰다. 해수 음이온 중 $MG^{2+},\;Ca^{2+},\;K^+$는 효소활성을 약간 증가시켰으나 $Na^{2+}$ 이온은 1mM이상농도에서 활성이 억제되었다. 본 논문에서 정제된 chitinase는 여러가지 특이점이 있는 serratia효소였다.

• 한국미생물·생명공학회지
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• 제24권3호
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• pp.282-289
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• 1996

2,3-DHBP dioxygenase was purified from E. coli CK1092 carrying the pcbC gene, which was cloned from 4-chlorobiphenyl-degrading Pseudomonas sp. P20. Purification of this enzyme was done by acetone precipitation, DEAE- Sephadex A-25 ion exchange chromatography, and preparative gel electrophoresis. The molecular weight of subunit was 34 kDa determined by SDS-PAGE, and that of native enzyme was about 270 kDa. It suggests that this enzyme consist of eight identical subunits. This enzyme was specifically active against only 2,3-DHBP as a substrate with 18 $\mu$M of Km value, but not catechol, 3-methylcatechol, 4-methylcatechol and 4-chlorocatechol. The optimal pH and temperature of 2,3-DHBP dioxygenase were pH 8.0 and 40-60$\circ$C. The enzyme was inhibited by Cu$^{2+}$, Fe$^{2+}$ and Fe$^{3+}$ ions, and was inactivated by H$_{2}$0$_{2}$2 and EDTA. The lower concentrations of some organic solvents such as acetone and ethanol don't stabilize the activity of 2,3-DHBP dioxygenase. The enzyme was completely inactivated by adding the reagents such as N-bromosuccinimide, iodine and p- diazobenzene sulfonic acid.

• Journal of Microbiology
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• 제37권3호
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• pp.148-153
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• 1999

Laccase produced by Coriolus hirsutus was purified to electrophoretic homogeneity by acetone precipitation, Sephacryl S-2000 HR chromatography, DEAE Sepharose CL-6B chromatography, and Mono Q HR 5/5 chromatography. The purification of laccase was 46.6-fold with an overall yield of 23.7%. Laccase from this fungus was a monomeric glycoprotein with 16% carbohydrate content, and has an isoelectric point of 4.2, and molecular mass of 78 kDa, respectively. The N-terminal amino acid sequence of the enzyme showed significant homology to hoste of laccases from Coriolus versicolor, Pycnoporus cinnabarius, and an unidentified basidiomycete, PM1. The highest rate of 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS) oxidation by laccase was reached at 45$^{\circ}C$, and te pH optima of the enzyme varied depending on the substrate in the range of 2.0 to 4.5. The enzyme was stable at 60$^{\circ}C$ for 5 h and lost 80% activity at 80$^{\circ}C$ in 30 min. The enzyme oxidized a variety of usual laccase substrates including lignin-related phenol, and had the highest affinity toward ABTS. Under standard assay conditions, the apparent Km value of the enzyme toward ABTS was 8.1 ${\mu}$M. The enzyme was completely inhibited by L-cysteine and sodium azide, but not by potassium cyanide, SDS, ad thiourea.

• 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.

• Geomechanics and Engineering
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• 제17권5호
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• pp.421-427
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• 2019

Coastal erosion is becoming a significant problem in Greece, Bangladesh, and globally. For the prevention and minimization of damage from coastal erosion, combinations of various structures have been used conventionally. However, most of these methods are expensive. Therefore, creating artificial beachrock using local ureolytic bacteria and the MICP (Microbially Induced Carbonate Precipitation) method can be an alternative for coastal erosion protection, as it is a sustainable and eco-friendly biological ground improvement technique. Most research on MICP has been confined to land ureolytic bacteria and limited attention has been paid to coastal ureolytic bacteria for the measurement of urease activity. Subsequently, their various environmental effects have not been investigated. Therefore, for the successful application of MICP to coastal erosion protection, the type of bacteria, bacterial cell concentration, reaction temperature, cell culture duration, carbonate precipitation trend, pH of the media that controls the activity of the urease enzyme, etc., are evaluated. In this study, the effects of temperature, pH, and culture duration, as well as the trend in carbonate precipitation of coastal ureolytic bacteria isolated from two coastal regions in Greece and Bangladesh, were evaluated. The results showed that urease activity of coastal ureolytic bacteria species relies on some environmental parameters that are very important for successful sand solidification. In future, we aim to apply these findings towards the creation of artificial beachrock in combination with a geotextile tube for coastal erosion protection in Mediterranean countries, Bangladesh, and globally, for bio-mediated soil improvement.

• 한국미생물·생명공학회지
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• 제20권5호
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• pp.559-564
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• 1992

섬유소 자원의 효율적 이용을 위하여 강력한 xylan 분해효소 생산 세균을 분리 및 선발하였으며, 형태학적, 배양학적 생리학적 특성을 확인한 결과 Bacillus sp. N25로 동정하였다. 이 균주에 의해서 생산된 효소를 ammonium sulfate precipitation, DAEA-sephadex A-50, Sephadex G-100 column으로 부분 정제하여 얻은 효소의 열안정성은 $50^{\circ}C$에서 30분간 처리시 80의 역가가 잔존 했으며, pH 안정은 pH 6-8 범위에서 30'C에서 10시간 방치 후에도 안정했으며, cellulose 분해능력은 없었으며, $Hg^{2+}$, $Ag^{2+}$, $Mn^{2+}$에 의해 저해되었다. 그리고 최종분해산물은 주로 xylose이므로 exo-type xylanase로 추정된다.

• Preventive Nutrition and Food Science
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• 제7권1호
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• pp.43-47
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• 2002

Crude caseinomacropeptide (CMP) was prepared from Na-caseinate using a commercial renneting enzyme. Most of the crude CMP was released from the Na-caseinate by hydrolyzing with the enzyme for 40 min. The hydrolysis of the k-casein with carbohydrate was slower than that of the k-casein without carbohydrate, as shown by the analyses of the sialic acid content and the tricine-SDS-polyacrylamide gel electrophoresis. The yield of crude CMP from Na-caseinate was 3.7%. Cation exchange chromatography showed that the crude CMP consisted of 40.5% CMP and 59.5% caseinogylcomacropetide (CGP). The effect of the TCA concentration on the solubility of CMP and CGP was determined by using crude CMP. The amounts of crude CMP and sialic acid decreased in the proportion to the increase of trichloroacetic acid (TCA) concentration from 2 to 12%, suggesting that the CGP containing carbohydrate, as well as the CMP having no carbohydrate, was precipitated in a range of 4 to 12%, depending on the TCA concentration. This result supports the hypothesis that the different non-glycosylated and glycosylated forms of CMP have different sensitivities to TCA precipitation.

• The Korean Journal of Ecology
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• 제9권4호
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• pp.231-241
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• 1986

The annual decomposition of cellulose and hemicellulose by microorganism and distribution of soil microbial flora were investigated in the humus horizon of a Pinus rigida stand in Mt. Kwanak. The cellulase activity was the lowest, 142$\mu$g glucose/g/hr from Dec. 1985 to Mar. 1986 and the highest, 760~1, 072$\mu$g glucose/g/hr in Jul. and Aug. 1985. The xylanase activity was 47% higher than the cellulase activity and was the lowest, 211~275$\mu$g xylose /g/hr from Feb. to Mar. 1986 and the highest as 799~1, 322$\mu$g xylose/g/hr from Jun. to Aug. 1986. The vertical distribution of the enzyme activity was decreased with the order of F, H, L, and A1 in both enzymes and the activities were exponentially decreased below L horizon, which suggests that most decomposition be done in F and H horizons with lots of organic matters. The SEM study slowed that the main decomposers of litters were fungi and initial attack into litters was also made by them. The enzyme activities of soil had strong correlations with the temperature and the precipitation. The correlation coefficients were 0.813 and 0.886 in the cellulase, and 0.673 and 0.626 in the xylanase for the temperature and the precipitation, respectively.

• 한국지반공학회논문집
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• 제37권4호
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• pp.39-47
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• 2021

최근 효소 기반 탄산칼슘 침전(EICP) 기법은 시멘트 기반 지반보강공법의 대안 중 하나로 간주되어 왔다. 하지만 EICP 기법에서 발생하는 환경 유해 부산물인 암모늄 이온의 배출에 대한 문제는 해결되지 않고 있다. 따라서 본 연구에서는 칼슘 치환 제올라이트를 사용하여 환경 유해 부산물이 없는 EICP(Ze-EICP)의 실험적 연구를 수행하고자 한다. 실험결과는 칼슘 치환 제올라이트를 사용하는 Ze-EICP가 염화칼슘을 사용하는 EICP와 비교하여 암모늄 이온은 96.96%가 제거되었으며, 거의 동일한 양의 탄산칼슘이 침전되었음을 보여주었다. 또한 Ze-EICP는 제올라이트의 조밀화와 탄산칼슘의 고결화로 인해 EICP 대비 높은 강도증진 효과를 보여주었다.

• Journal of Applied Biological Chemistry
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• 제48권2호
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• pp.75-78
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• 2005

Halotolerant protease produced by Halomonas marisflava KCCM 10457 was partially purified through ammonium sulfate precipitation and Sephacryl S-200HR gel permeation chromatography. Optimal pH and temperature of protease were 11.0 and $45^{\circ}C$, respectively. Enzyme activity was inhibited by $Cu^{2+}$, $Hg^{2+}$, $Fe^{2+}$, and $Fe^{3+}$, and selectively inhibited by p-chloromercuribenzoic acid (PCMB), suggesting this enzyme is cysteine protease. The enzyme is halotolerant, because it retained 77% of original activity in presence of 3.33 M NaCl. The protease showed broad substrate specificity to various natural proteins; BSA, casein, egg albumin, gelatin, and hemoglobin.