• 대한미생물학회지
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• 제21권3호
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• pp.399-406
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• 1986

This study was performed to establish human plasmacytoma cell line as the partner cells for producing human hybridoma. Bone marrow cells from a multiple myeloma patient from Seoul National University Hospital, Korea were cultured and established as the cell line, named as HMC-BM4. HMC-BM4 cells were cultivated in RPMI 1640 media containing 8-azaguanine(8-AG; gradually increasing concentration from $1\;{\mu}g/ml$ to $20\;{\mu}g/ml$). 8-AG resistant cells were collected and cloned by limiting dilution. Each clone was divided and tested to die in hypoxanthine, aminopterine and thymidine (HAT) selection media. Finally one clone was selected and named as HMC-AR, which was sensitive to HAT selection media. HMC-AR cells showed typical morphology of plamacytoma in Wright staining. No cell formed the rosette with sheep erythrocytes. Surface membrane $\mu$ heavy chain was detected in 20% of HMC-AR cells and cytoplasmic $\mu$ heavy chain in 90% of them by direct immunofluorescent staining. Ia-like antigen was found in 90% of HMC-AR cells by indirect immunofluorescent staining using anti-Ia-like antigen monoclonal antibody, 1BD9-2. And about $1.0\;{\mu}g/ml$ of human $\mu$ heavy chain was detected in the 3-day culture supernatant of HMC-AR cells. 88% of cells contained 46 chromosomes. Mycoplasma was not detected in HMC-AR cells by Hoechst 33258 staining. This cell line would be used for making hybridomas secreting human monoclonal antibody.

• Animal cells and systems
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• 제2권1호
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• pp.107-111
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• 1998

Two kinds of humoral factors were observed in 2 orders, 7 families, 10 genera, and 15 different species of Korean ascidians. They are the naturally occuring hemagglutinins and/or hemolysins against human erythrocytes A, B, and 0. All but two species showed aggregative activity, although there were considerable variations in titer. The weak agglutinating and lytic activities were increased in the presence of $Ca^{++}$. Much higher activities of agglutination and/or lysis were shown in the hemolymph than extracts from tissues, and a higher response was shown in adults than in juveniles. No distinct differences from collected locations were observed. The hemolymph of Ciona intestinalis showed a strong hemolytic (cytotoxic) and weak agglutinins capacities. In addition, hemolymph of Styela plicata and Styela clava clava also showed hemoagglutining and hemolytic activities. Botryllus tuberatus had hemagglutining and weak lytic activities. Other species showed only hemagglutining activity. These agglutining activities are probably responsible for carbohydrate recognition in solitary or colonial ascidian. The lytic activity is probably responsible for antibacterial defense and nonfusion reactions between allogeneic colonial ascidians, especially the genus of Botryllus. The occurrences of humoral factors in ascidians were independent of their geographic distributions.

• Journal of Microbiology and Biotechnology
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• 제30권3호
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• pp.382-390
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• 2020

Periplanetasin-4 is an antimicrobial peptide with 13 amino acids identified in cockroaches. It has been reported to induce fungal cell death by apoptosis and membrane-targeted action. Analogs were designed by substituting arginine residues to modify the electrostatic and hydrophobic interactions accordingly and explore the effect of periplanetasin-4 through the increase of net charge and the decrease of hydrophobicity. The analogs showed lower activity than periplanetasin-4 against gram-positive and gram-negative bacteria. Similar to periplanetasin-4, the analogs exhibited slight hemolytic activity against human erythrocytes. Membrane studies, including determination of changes in membrane potential and permeability, and fluidity assays, revealed that the analogs disrupt less membrane integrity compared to periplanetasin-4. Likewise, when the analogs were treated to the artificial membrane model, the passage of molecules bigger than FD4 was difficult. In conclusion, arginine substitution could not maintain the membrane disruption ability of periplanetasin-4. The results indicated that the attenuation of hydrophobic interactions with the plasma membrane caused a reduction in the accumulation of the analogs on the membrane before the formation of electrostatic interactions. Our findings will assist in the further development of antimicrobial peptides for clinical use.

• Journal of Microbiology and Biotechnology
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• 제15권5호
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• pp.1039-1046
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• 2005

The salt resistance of antibacterial activity and synergistic effect with clinically used antibiotic agents are critical factors in developing effective peptide antibiotic drugs. For this reason, we investigated the resistance of antibacterial activity to antagonism induced by NaCl and $MgCl_2$ and the synergistic effect of P20 with cefotaxime. P20 is a 20-residue synthetic peptide derived from a cecropin A (CA)-melittin(ME) hybrid peptide. In this study, P20 was found to have potent antibacterial activity against clinically isolated methicillin-resistant Staphylococcus aureus (MRSA) strains without hemolytic activity against human erythrocytes. The combination study revealed that P20 in combination with cefotaxime showed synergistic antibacterial activity in an energy-dependent manner. We also confirmed the synergism between P20 and cefotaxime by fluorescence-activated flow cytometric analysis by staining bacterial cells with propidium iodide (PI) and bis-(1,3-dibutylbarbituric acid) trimethine oxonol (BOX). This study suggests that P20 may be useful as a therapeutic antibiotic peptide with synergistic effect in combination with conventional antibiotic agents.

• 한국미생물·생명공학회지
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• 제30권3호
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• pp.230-234
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• 2002

모기 살충성 Bacillus thuringiensis subspangiensis 21-2균주의 용혈성 내독소 단백질의 특성을 검토하고자 21-2균주의 용혈성을 가진 유전자를 Escherichia coli HBIO쎄 형질전환시켰다. 이들 중에서 형질전환 균주 47은 독소 단백질을 생산하며 2.5 kb DNA을 함유한다는 것을 효소항체법, immunoblot및 DNA전기영동법으로 확인하였다. 또한, 형질전환 균주 47-5는 2.5 kb DNA를 다시 Hind ll견 절단하여 pUC118 연결시켜서 조제하였다 그 결과형질전환 균주 47-5은 1.Bkb DNA를 함유하며, 23 kD꺼 독소 단백질을 발현하고, 발현된 독소 단백질은 Aedes aegypti모기 유충에 독성을 나타내었다. 또한 23 kDa의 내독소 단백질 그 자체로는 사람의 적혈구를 용해하지 못하였으나, proteinase K로 처리한 후에는 적혈구에 대해 용혈성을 나타내었다.

• Journal of Microbiology and Biotechnology
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• 제17권11호
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• pp.1797-1804
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• 2007

Lycopene, an acyclic carotenoid found in tomatoes (Lycopersicon esculentum) and a number off fruits, has shown various biological properties, but its antifungal effects remain poorly understood. The current study investigated the antifungal activity of lycopene and its mode of action. Lycopene showed potent antifungal effects toward pathogenic fungi, tested in an energy-independent manner, with low hemolytic effects against human erythrocytes. To confirm the antifungal effects of lycopene, its effects on the dimorphism of Candida albicans induced by fetal bovine serum (FBS), which plays a key role in the pathogenesis of a host invasion, were investigated. The results showed that lycopene exerted potent antifungal activity on the serum-induced mycelia of C. albicans. To understand the antifungal mode of action of lycopene, the action of lycopene against fungal cell membranes was examined by FACScan analysis and glucose and trehalose-release test. The results indicated that lycopene caused significant membrane damage and inhibited the normal budding process, resulting from the destruction of membrane integrity. The present study indicates that lycopene has considerable antifungal activity, deserving further investigation for clinical applications.

• Archives of Pharmacal Research
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• 제29권9호
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• pp.746-751
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• 2006

Amentoflavone is a plant biflavonoid that was isolated from an ethyl acetate extract of the whole plant of Selaginella tamariscina (Beauv.) spring. 1D and 2D NMR spectroscopy including DEPT, HMQC, and HMBC were used to determine its structure. Amentoflavone exhibited potent antifungal activity against several pathogenic fungal strains but had a very low hemolytic effect on human erythrocytes. In particular, amentoflavone induced the accumulation of intracellular trehalose on C. albicans as a stress response to the drug, and disrupted the dimorphic transition that forms pseudo-hyphae during pathogenesis. In conclusion, amentoflavone has great potential to be a lead compound for the development of antifungal agents.

• BMB Reports
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• 제36권2호
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• pp.214-222
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• 2003

A lectin named AAL has been purified from the fruiting bodies of the edible mushroom Agrocybe aegerita. AAL consisted of two identical subunits of 15.8 kDa, its pI was about 3.8 determined by isoelectric focusing, and no carbohydrate was discerned. Being treated by pyrogultamate aminopeptidase, the blocked N-terminus of AAL was sequenced as QGVNIYNI. AAL agglutinated human and animal erythrocytes regardless of blood type or animal species. Its hemagglutinating activity was unaffected by acid or alkali treatment and demetalization or addition of divalent metals $Mg^{2+}$, $Ca^{2+}$ and $Zn^{2+}$. AAL was toxic to mice: its LD50 was 15.85 mg per kilogram body weight by intraperitoneal injection. In this study, two novel activities of AAL were proved. It showed inhibition activity to infection of tobacco mosaic virus on Nicotiana glutinosa. The result of IEF suggested that AAL attached to TMV particles. Mycelia differentiation promotion was the other interesting activity. AAL promoted the differentiation of fruit body primordia from the mycelia of Agrocybe aegerita and Auricularia polytricha. AAL antiserum was prepared and immunologically cross-reactived with several proteins from five other kinds of mushrooms. These results suggested that AAL probably was a representative of a large protein family, which plays important physiological roles in mushroom.

• Molecular & Cellular Toxicology
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• 제3권1호
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• pp.68-75
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• 2007

[ $\alpha$ ]-Terpinene has been known as a repellent against the mosquito Culex pipiens pallens Coquillett based on a human forearm bioassay. $\alpha$-Terpinene showed significantly greater repellency than a commercial formulation, N, N-diethyl-m-methylbenzamide (deet). In this study, skin and eye sensitivity of $\alpha$-terpinene (2%) was examined with bioassays using white New Zealand rabbits. There were somewhat gross and histological changes observed in these treatments. Eye irritancy assays examined gross changes to cornea, iris and conjuctiva, and histological changes to smear of ocular discharge and eye tissue. Treated rabbits were divided into two cohorts, a saline washed cohort (W) or a non-washed cohort (NW). Opacity of cornea and redness, chemosis and discharge of conjuctiva were observed in both cohorts, but disappeared within 4 and 10 days in W and NW, respectively. Main components of ocular discharges were fibrin, epithelial or epitheloid cells, lymphoid cells, erythrocytes and granulocytes. These abnormal cellular components disappeared within 4 days and 10 days in W and NW, respectively. No permanent histological differences were observed between the two cohorts. However, severe irritation was determined as 57.2 of I.I.O.I value on the first day after treatment. These findings indicate a spray-type solution containing 2% $\alpha$-terpinene may serve as an alternative mosquito repellent and further studies need to reduce the eye irritation with formulation changes.

• 한국식품영양과학회지
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• 제24권4호
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• pp.587-593
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• 1995

택사로부터 황산암모늄 분별 침전, DEAE-cellulose ion exchange chromatography, Sephadex G-150 chromatography 등의 방법을 이용하여 렉틴을 분리, 정제하였다. 정제한 렉틴의 분자량은 gel filtration법에 의해 측정한 결과 90,500 dalton이었으며, SDS polyacrylamide gel 전기영동을 실시한 결과 subunit 분자량은 각각 42,000, 27,000, 22,500 dalton으로 나타나므로 택사 렉틴이 heterotrimer임을 알았다. 정제된 택사 렉틴은 사람 적혈구의 경우 모든 혈액형에 대하여 응집현상을 나타내었으며 돼지, 쥐 및 개 등의 적혈구에 대해서도 모두 응집현상이 나타나 혈구 비특이성임을 보여주었다. 또한 이 렉틴은 sialic acid, glucose, ribose, sucrose, lactose, galactose 등 당에 의해서, 그리고 $Hg^{2+},\;Fe^{2+},\;Cu^{2+}$ 이온 등에 의해 적혈구 응집력이 저해되었다.