• 한국환경복원기술학회지
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• 제17권5호
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• pp.65-77
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• 2014

The ecosystem of Jaemin stream, flowing into the center of Gongju-si, had been damaged by low water quality and lack of water quantity of the steam. However, after applying the SSB (Sustainable Structured wetland Biotop) system to the flood plain and the upstream of Jaemin stream, the efficiency of ecological water purification and ecological restoration are as follows. Through the constant maintenance and monitoring from year 2009 to year 2013 after restorative design and construction the average influent concentration of BOD5 was 4.2 mg/L, and the average effluent concentration was 1.8 mg/L, reaching ecological water purification rate of 57%. As for the T-N, the average influent concentration was 9.983 mg/L, and the average effluent concentration was 6.303 mg/L, showing the rate of 37%. For the T-P, the average influent concentration was 0.198 mg/L, and the average effluent concentration was 0.098 mg/L, being the rate of 51%. The vegetation of Jaemin stream monitored for 2 years after the restoration was composed of 51 species in 28 families which show high ratio of planted native species. As for the animals in the site, 5 species in 3 families of reptiles and amphibians, 34 species of 23 families of birds, and 3 species in 2 families of mammals were monitored, indicating that the bio-diversity of the site has improved, as well.

• Journal of Microbiology and Biotechnology
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• 제25권2호
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• pp.274-279
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• 2015

Receptor activator of nuclear factor-kappa B ligand (RANKL) is a critical factor in osteoclastogenesis. It makes osteoclasts differentiate and multinucleate in bone remodeling. In the present study, RANKL was expressed as a soluble maltose binding protein (MBP)-fusion protein using the Escherichia coli maltose binding domain tag system (pMAL) expression vector system. The host cell E. coli DH5α was cultured and induced by isopropyl β-_D-1-thiogalactopyranoside for rRANKL expression. Cells were disrupted by sonication to collect soluble MBP-fused rRANKL. The MBP-fusion rRANKL was purified with MBP Trap affinity chromatography and treated with Tobacco Etch Virus nuclear inclusion endopeptidase (TEV protease) to remove the MBP fusion protein. Dialysis was then carried out to remove binding maltose from the cleaved rRANKL solution. The cleaved rRANKL was purified with a second MBP Trap affinity chromatography to separate unsevered MBP-fusion rRANKL and cleaved MBP fusion protein. The purified rRANKL was shown to have biological activity by performing in vitro cell tests. In conclusion, biologically active rRANKL was successfully purified by a simple two-step chromatography purification process with one column.

• Journal of Microbiology and Biotechnology
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• 제22권1호
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• pp.58-68
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• 2012

An investigation was conducted on the enhancement of production and purification of an oxidant and SDS-stable alkaline protease (BHAP) secreted by an alkalophilic Bacillus horikoshii, which was screened from the body fluid of a unique Korean polychaeta (Periserrula leucophryna) living in the tidal mud flats of Kwangwha Island in the Korean West Sea. A prominent effect on BHAP production was obtained by adding 2% maltose, 1% sodium citrate, 0.8% NaCl, and 0.6% sodium carbonate to the culturing medium. The optimal medium for BHAP production contained (g/l) SBM, 15; casein, 10; $K_2HPO_4$, 2; $KH_2PO_4$, 2; maltose, 20; sodium citrate, 10; $MgSO_4$, 0.06; NaCl, 8; and $Na_2CO_3$, 6. A protease yield of approximately 56,000 U/ml was achieved using the optimized medium, which is an increase of approximately 5.5-fold compared with the previous optimization (10,050 U/ml). The BHAP was homogenously purified 34-fold with an overall recovery of 34% and a specific activity of 223,090 U/mg protein using adsorption with Diaion HPA75, hydrophobic interaction chromatography (HIC) on Phenyl-Sepharose, and ion-exchange chromatography on a DEAE- and CM-Sepharose column. The purified BHAP was determined a homogeneous by SDS-PAGE, with an apparent molecular mass of 28 kDa, and it showed extreme stability towards organic solvents, SDS, and oxidizing agents. The $K_m$ and $k_{cat}$ values were 78.7 ${\mu}M$ and $217.4s^{-1}$ for N-succinyl-Ala-Ala-Pro-Phe-pNA at $37^{\circ}C$ and pH 9, respectively. The inhibition profile exhibited by PMSF suggested that the protease from B. horikoshii belongs to the family of serine proteases. The BHAP, which showed high stability against SDS and $H_2O_2$, has significance for industrial application, such as additives in detergent and feed industries.

• Archives of Pharmacal Research
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• 제29권5호
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• pp.418-423
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• 2006

Tabanus anticoagulant protein (TAP) was isolated from the whole body of the tabanus, Tabanus bivittatus, using three purification steps (ammonium sulfate fractionation, gel filtration on Bio-Gel P-60, and ion exchange chromatography on DEAE Sephadex gel). The purified TAP, with a molecular weight of 65 kDa, was assessed to be homogeneous by SDS-polyacrylamide gel electrophoresis, and an isoelectric point of 7.9 was determined by isoelectric focusing. The internal amino acid sequence of the purified protein was composed of Ser-Leu-Asn-Asn-Gln-Phe-Ala-Ser-Phe-lle-Asp-Lys-Val-Arg. The protein was activated by $Cu^{2+}\;and\;Zn^{2+}$, and the optimal conditions were found to be at pH $3\sim6\;and\;40\sim70^{\circ}C$. Standard coagulation screen assays were used to determine thrombin time and activated partial thromboplastin time. Chromogenic substrate assays were performed for thrombin and factor Xa activity. TAP considerably prolonged human plasma clotting time, especially activated partial thromboplastin time in a dose-dependent manner; it showed potent and specific antithrombin activity in the chromogenic substrate assay. Specific anti-factor Xa activity in TAP was not detected. Overall, this result suggested that TAP has significant anticoagulant activity on blood coagulation system.

• 미생물학회지
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• 제55권3호
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• pp.309-312
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• 2019

본 연구에서는 폐수처리장의 바이오필터로부터 Comamonas sp. NLF-7-7 균주를 분리하고 유전체서열을 PacBio RS II와 Illumina HiSeqXten 플랫폼을 사용하여 분석하였다. 염색체의 크기는 3,333,437 bp로 G + C 구성 비율은 68.04%, 총 유전자수는 3,197개, rRNA는 9개 및 tRNA는 49개로 구성되었다. 본 유전체는 오염물질분해와 플록형성에 관여하는 황산화 경로 유전자(SoxY, SoxZ, SoxA 및 SoxB)와 플록형성 경로 유전자(EpsG, EpsE, EpsF, EpsG, EpsL 및 glycosyltransferase)를 포함하고 있다. 이러한 Comamonas sp. NLF-7-7 균주는 폐수를 정화하는데 활용될 수 있다.

• Journal of Applied Biological Chemistry
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• 제54권4호
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• pp.231-237
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• 2011

A gene encoding a putative alanine racemase in Xanthomonas. oryzae pv. oryzae was cloned, expressed and characterized. Expression of the cloned gene was performed in Escherichia coli BL21(DE3)pLys using a pET-21(a) vector harbouring $6{\times}histidine$ tag. Purification of the recombinant alanine racemase by affinity chromatography resulted in major one band by sodium dodecyl sulfate polyacryl amide gel electrophoresis analysis, showing about 45 kDa of molecular weight. The alanine racemase gene, cloned in this experiment, appears to be constitutively expressed in X. oryzae, as analyzed by reverse transcriptase polymerase chain reaction. The enzyme was the most active toward L-alanine and secondly D-alanine, showing a racemic reaction, thus the enzyme is considered as an alanine racemase. The enzyme was considerably activated by addition of pyridoxal-5-phosphate (PLP), showing that 75% increase in activity was observed at 0.3 mM, compared with control. D-Cysteine as well as L-cysteine significantly inhibited the enzyme activity. The inhibitions by cysteines were more prominent in the absence of PLP, showing 9 and 5% of control activity at 2 mM of addition, respectively. The enzyme was the most active at pH 8.0 and more stable at alkaline pHs than acidic pH condition.

• KSBB Journal
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• 제26권5호
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• pp.393-399
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• 2011

Bacillus subtilis natto producing high level of a fibrinolytic enzyme was selected and Ultra Nattokinase$^{(R)}$ was manufactured by fermentation and purification. It was performed the evaluation of the antithrombotic effect of Ultra Nattokinase$^{(R)}$ (20,000 FU/g) with rat blood plasma. The maximum aggregation (inhibition ratio) was 71% (0%), 69% (2.8%), 62% (12.7%), 16% (77.5%) and 9% (87.3%), respectively, in the order of 0, 5, 10, 50 and 100 mg/mL of Ultra Nattokinase$^{(R)}$ solutions. Ultra Nattokinase$^{(R)}$ had antithrombotic effect, which was associated with the suppression of collagen-induced platelet aggregation. Ultra Nattokinase$^{(R)}$ in the topic of the FDP (fibrinogen degradation products) in blood coagulation tests showed a significant increasing trend. And based on the daily record of meal 39 people of ITT (what ?) group consisted with 19 people of NP (what ?) group and 20 people of PN (what ?) group except four people, two people who took vitamin K affecting the experiment and two people who took alcohol, finding to be taken Ultra Nattokinase$^{(R)}$ showed an increase in the FDP value after four weeks. In addition, FDP value of 41 people of ITT group except two people having metabolic syndrome was increased by Ultra Nattokinase$^{(R)}$.

• 인간식물환경학회지
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• 제23권2호
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• pp.179-189
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• 2020

In order to increase the indoor air purification effect of plants, plants need to be placed on 5-10% of indoor spaces. To increase the density and utilization of plants in indoor spaces, studies on bio-wall, a vertical green wall system, have been recently conducted. The purpose of this study was to investigate the growth characteristics of 7 indoor plants introduced to the system and their rooting zones at different irrigation cycles. This study was conducted to investigate a proper irrigation cycle for the continuous maintenance of bio-wall systems. The conditions of their growth environment were maintained as follows: light intensity, 20-50 μmol·m^-2·s^-1 PPFD; and temperature, 20 - 25℃. For fertilization, Hyponex diluted with water at the ratio of 1:1,000 was supplied to plants. Irrigation was treated at intervals of 1, 3, 5, and 7 days for 1 hour at a time. As a result, there was no significant difference in the growth of plants between different irrigation cycles. Dieffenbachia 'Marianne' showed a significant decrease in the number of leaves at the irrigation cycle of 7 days. In addition, the chlorophyll content was relatively low at the irrigation cycle of 7 days. In terms of the color of leaves, a decrease in L value and b value and an increase in a value were observed, resulting in changes in brightness and color. Ardisia pusilla 'Variegata' showed a slightly higher photosynthetic activity and stomatal conductance when it was watered every day and once per 5 days, while Epipremnum aureum showed a relatively higher photosynthetic activity and stomatal conductance at the irrigation cycle of 3 days. In the case of root activity, it was found that the longer irrigation cycle, the higher root activity compared to daily irrigation. The development of roots of Peperomia clusiifolia was promoted by watering at long intervals. However, in the case of Aglaonema 'Siam-Aurora', the total number of roots decreased at the interval of 7 days. In conclusion, a proper irrigation cycle for the sustainable maintenance of vertical bio-wall systems seems to be 3 days.

• BMB Reports
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• 제52권8호
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• pp.496-501
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• 2019

Conventionally, immunotoxins have been produced as a single polypeptide from fused genes of an antibody fragment and a toxin. In this study, we adopted a unique approach of chemical conjugation of a toxin protein and an antibody fragment. The two genes were separately expressed in Escherichia coli and purified to high levels of purity. The two purified proteins were conjugated using a chemical linker. The advantage of this approach is its ability to overcome the problem of low recombinant immunotoxin production observed in some immunotoxins. Another advantage is that various combinations of immunotoxins can be prepared with fewer efforts, because the chemical conjugation of components is relatively simpler than the processes involved in cloning, expression, and purification of multiple immunotoxins. As a proof of concept, the scFv of trastuzumab and the PE24 fragment of Pseudomonas exotoxin A were separately produced using E. coli and then chemically crosslinked. The new immunotoxin was tested on four breast cancer cell lines variably expressing HER2. The chemically crosslinked immunotoxin exhibited cytotoxicity in proportion to the expression level of HER2. In conclusion, the present study revealed an alternative method of generating an immunotoxin that could effectively reduce the viability of HER2-expressing breast cancer cells. These results suggest the effectiveness of this method of immunotoxin crosslinking as a suitable alternative for producing immunotoxins.

• 생명과학회지
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• 제19권5호
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• pp.671-675
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• 2009

Prostate cancer has been a critical health problem due to an increase of prostate cancer-related deaths worldwide. Also, a frequent treatment option for prostate cancer is androgen ablation, but this treatment has a limited scope, especially for hormone-refractory cancer. There is an urgent need for the identification of alternative therapeutic strategies for prostate cancer. Previously, over one hundred species of dried-plant methanol extracts were tested for inhibitory effects on proliferation. One of them, Piper longum Linn. was selected based on its potent anti-proliferation effect. The dried root of P. longum Linn. was extracted with 100% methanol for 2-3 days and its extract was fractionated using chloroform. The chloroform layer was then subjected to column chromatography on silica gel, reverse phase-18 (RP-18) and Sephadex LH-20, in turn. Finally, the pure compound was obtained and identified as pipernonaline by NMR spectroscopic and physico-chemical analysis. In this study, anti-proliferation and cell cycle arrest effects of pipernonaline on human prostate cancer PC-3 cells were investigated using the MTT and PI staining, respectively. Our findings suggest that pipernonaline represents a dose-dependent growth inhibition pattern on PC-3 cells and, moreover, its growth inhibition is associated with sub-G1 and G0/G1 cell cycle accumulation in PC-3 cells. Also, these results provide an anticancer candidate for human prostate cancer.