• KSBB Journal
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• 제32권2호
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• pp.133-139
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• 2017

Rheum palmatum has traditionally been used as a preventive agent and medication against fever and infection. The aim of this study was to isolate and characterize an antibacterial substance from R. palmatum that is effective against bacterial vaginosis. A methanol extract from R. palmatum showed antibacterial activity against Lactobacillus vaginalis KC TC 3515, Chryseobacterium gleum KCTC 2904, and Sphingomonas paucimobilis KCTC 2834, which cause bacterial vaginosis. After extraction and pH control of the methanol extract from R. palmatum, we found that acidic and alkaline extracts did not show antibacterial activity. A neutral extract (50 mg/mL) displayed an inhibitory zone of 18 mm on a nutrient agar plate with C. gleum KCTC 2904. Fractions No. 11 and 12 among 41 fractions obtained by silica gel column chromatography produced inhibitory zones of 10 mm on nutrient agar plates with C. gleum KCTC 2904. $R_f0.15$ and $R_f0.17$ spots produced by TLC of fraction No. 11 showed antibacterial activity against C. gleum KCTC 2904. Isolation and purification of the peak at a retention time (Rt) of 9.427 min was achieved by HPLC of $R_f0.29spots$. The peak at Rt 9.427 min showed antibacterial activity against C. gleum KCTC 2904.

• Journal of Microbiology and Biotechnology
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• 제17권9호
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• pp.1469-1476
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• 2007

Jeot-gal is a traditional Korean fermented seafood and has long been used for seasoning. We isolated 188 strains from shrimp, anchovy, and yellow corvina Jeot-gal, and screened sixteen strains that showed strong fibrinolytic activities on a fibrin plate. Among those strains, the strain that had the largest halo zone was chosen and identified as Bacillus licheniformis by using 16S rDNA sequencing and an API CHB kit. The fibrinolytic activity of Bacillus licheniformis was characterized and designated as bpKJ-31. The active component of bpKJ-31 was identified as a 37 kDa protein, designated bacillopeptidase F, by internal peptide mapping and N-terminal sequencing. The optimum activity of bpKJ-31 was shown at pH 9 and $40^{\circ}C$, with a chromogenic substrate for plasmin. It had high degrading activity for the $B{\beta}$-chain and $A{\alpha}$-chain of fibrin(ogen), and also acted on thrombin, but not skim milk and casein. The amidolytic activity of bpKJ-31 was inhibited by 1 mM phenylmethanesulfonyl fluoride, but 1 mM EDTA did not affect the enzyme activity, indicating that bpKJ-31 is an alkaline serine protease, like a plasmin. The bpKJ-31 showed approximately 14.3% higher fibrinolytic activity than the plasmin. These features of bpKJ-31 make it attractive as a health-promoting biomaterial.

• Biomolecules & Therapeutics
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• 제12권3호
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• pp.189-193
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• 2004

Over the past several years, research efforts have been directed both at economically producing valuable substances from the wood biomass and at producing lignolytic enzymes at a lower cost. In the present study, we found that Phellinus igniarius, the basidiomycetes, secreted lignin peroxidase as a main lignolytic enzyme, which was detected maximum activity at 16th day of culture and showed 37 kDa of molecular mass in identification by activity assay and purification by anion-exchange chromatography. The Phellinus igniarius-derived lignin peroxidase hydrolyzed steam-exploded wood (Quercus mongolica) powder into small molecules showing cytotoxicity against cancer cel1s (HepG2 hepatoma, SK-N-SH neuroblastoma, B16 melanoma, MBT-2 bladder cancer). In addition, the enzyme hydrlysates of lignins (ELg) that were extracted from the steam-exploded oak showed more potent cytotoxic effects on the cancer cells than the enzyme hydrolysates of wood biomass (EWp), indicating that the cytotoxic effect of EWp may be due to the enzyme-degraded products of lignin among the lignocellulosics. Furthermore, the cytotoxic effect of ELg on Chang, normal liver cells, was much less potent than that of ELg on HepG2 and B16 cancer cells, indicating that the cytotoxic effect of ELg may be specific for cancer cells. The present results suggest that Phellinus igniarius may be a useful resource for the large-scale production of lignin peroxidase and that the lignin peroxidase may be applied for the generation of valuable biodegradation products from wood lignocellulosics for medical use.

• Applied Biological Chemistry
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• 제46권3호
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• pp.176-182
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• 2003

한국재래간장으로부터 혈전용해효소를 강하게 생산하는 균주를 선발하고 이를 Bacillus subtilis K7로 동정하였다. B. subtilis K7이 생산하는 혈전용해성 protease를 정제하여 분자량을 확인한 결과 21,500 Da이었다. 정제된 효소의 최적반응조건은 $40^{\circ}C$와 pH 9.0이었으며 pH 5.0라서 12.0까지 안정하고 $50^{\circ}C$에서 20분간 열처리한 후에도 50%이상의 효소활성을 가지며 EDTA, CDTA 및 iodoacetat에 실활하는 효소이었다. 이 효소의 fibrin에 대한 Km 값은 $1.8{\times}10^{-2}$ M이었다.

• International Journal of Industrial Entomology and Biomaterials
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• 제2권2호
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• pp.161-166
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• 2001

The storage proteins of the mulberry longicorn beetle, Apriona germari Hope, were purified and characterized. Three kinds of storage protein (SP1, SP2 and Sp3) were purified from the last instar larval hemolymph of A. germari by the FPLC techniques, anion exchange chromatography and gel permeation chromatography. The SP1, SP2 and SP3 have a native molecular weight of 480, 440 and 420 kDa, respectively. In the SDS-polyacrylamide gel electrophoresis analysis, these storage proteins are composed of a single protein subunit with molecular weight of 90, 85 and 80 kDa, respectively. This result showed that the storage proteins are hexameric protein. The SP1 and SP2 were stained with Schiffs reagent, but SP3 was not stained. It can be assumed that SP1 and SP2 are glycoprotein. Western blot analyses using the each of polyclonal antiserum against purified SP1, SP2 and SP3 showed that the three antibodies reacted with the each of SP bands, respectively. Also, antibodies against SP1 and SP3 cross-reacted with the SP3 and SP1, respectively. However, SP2 was not cross-reacted with these two antibodies. Also, antiserum against SP2 did not cross-reacted with the SP1 and SP3.

• 대한미생물학회지
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• 제35권4호
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• pp.273-282
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• 2000

dTDP-rhamnose provides L-rhamnose to the bridge-like structure between mycolyl arabinogalactan and peptidoglycan of the mycobacterial cell wall. dTDP-rhamnose is composed of glucose-l-phosphate and dTTP by four enzymes encoded by rmlA-D. To determine the region(s) of RmlC protein essential for its dTDP-4-keto-6-deoxyglucose epimerase activity, we overexpressed both whole (202 amino acids) and three different truncated (N-terminal 106 or 150 or C-terminal 97 amino acids) RmlC proteins of Mycobacterium tuberculosis. The RmlC enzyme activity in the soluble lysates of ${\Delta}rmlC$ E. coli strain $S{\Phi}874$ (DE3 PlysS) expressing the wild type or truncated rmlC genes was initially analyzed by three sequential reactions from dTDP-glucose to dTDP-rhamnose in the presence of purified RmlB and RmlD. All three soluble lysates containing the truncated RmlC proteins showed no enzyme activity, while that containing the wild type RmlC was active. This wild type RmlC was then overexpressed and purified. The incubation of the purified RmlC enzyme so obtained with dTDP-4-keto-6-deoxyglucose resulted in the conversion of dTDP-4-keto-rhamnose. The results show that the truncated regions of the RmlC protein are important for the RmlC enzyme activity in M. tuberculosis.

• 미생물학회지
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• 제31권1호
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• pp.63-71
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• 1993

A xylanase (xylanase I) was purified 11.9-fold from the culture filtrate of Trichoderma koningii ATCC 26113 by the column chromatography on Sephadex G-75, SP-Sephadex C-50, DEAE-Sephadex A-50 and Sephadex G-50 with an overall yield of 8.2%. The molecular mass determined by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis was found to be a monomeric polypeptide of ca. 35 kDa. The isoelectric point of the enzyme was estimated to be 9.3. The optimal reaction pH and temperature are 5.8 and 55.deg.C, respectively. The enzyme is stable up to 60.deg.C, while 78% of its activity is lost after the incubation for 10 min at 70.deg.C. The enzyme hydrolyzes sylan with relatively high activity, as well as carboxymethyl cellulose and avicel. The $K_{m}$ values of the enzyme for oat-spelf sylan, larchwood xylan and Avicel were 3.5, 1.6 and 10. 1 mg/ml, respectively. The enzyme hydrolyzed oat-spelt sylan to sylose, sylobiose, sylotriose and arabionoxylobiose, while it degraded larchwood xylan to xylose, xylobiose, xylotriose and arabionoxylobiose, while it degraded larchwood xylan to xylose, xylobiose and xylotriose as the major products. The hydrolysis patterns indicate that xylanase I is endo-enzyme.e.

• Bulletin of the Korean Chemical Society
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• 제28권6호
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• pp.929-935
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• 2007

The objective of this study was to develop a rapid, simple, and qualitative acetylcholinesterase (AChE)- detection kit, based on a modification of the Ellman and ELISA methods, for the detection of organophosphorus (OP) and carbamate (CB) pesticide. The developed kits were used to screen a large number of agricultural samples (spiked and real) for OP and CB pesticide residues. AChE was extracted from the heads of honeybees (Apis mellifera L.) using Triton X-100, and was purified through 3 steps: diethylaminoethylcellulose chromatography (DEAE), affinity chromatography and membrane filtering, and Mono-Q column chromatography. Epoxy-activated Sepharose 6B affinity chromatography was used for large-scale purification. The presence of OP and CB pesticide residues in agricultural samples was assayed on the basis of AchE inhibition value. The presence (6 bands) or absence of some colored bands on the test line indicated a negative or positive result, respectively. The limits of detection for measured organophosphorus (OP) and carbamates (CB) pesticide residues in standard pesticide solutions and fortified samples were ranged from 0.50 to 2.50 ppm and 0.50 to 4.75 ppm, respectively.

• Archives of Pharmacal Research
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• 제28권7호
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• pp.799-803
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• 2005

In the course of screening anti-dementia agents from natural products, two $\beta$-secretase (BACE1) inhibitors were isolated from the ethyl acetate soluble fraction of Sanguisorbae Radix by the activity-guided purification using silica gel, Sephadex LH-20, and RP-HPLC. They were identified as 1,2,3-trigalloyl-4,6-hexahydroxydiphenoyl-$\beta$-D-glucopyranoside (Tellimagrandin II, 1) and 1,2,3,4,6-pentagalloyl-$\beta$-D-glucopyranoside (2) and were shown to non-competitively inhibit $\beta$-secretase (BACE1) with the $IC_{50}$ values of $3.10{\times}10^{-6}M\;and\;3.76{\times}10^{-6}M$, respectively. The Ki values of 1 and 2 were $6.84{\times}10^{-6}M\;and\;5.13{\times}10^{-6}M$. They were less inhibitory to asecretase (TACE) and other serine proteases such as chymotrypsin, trypsin, and elastase, suggesting that they were relatively specific inhibitors of BACE1.

• 생명과학회지
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• 제16권1호
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• pp.1-5
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• 2006

본 연구는 Penibacillus polymyxa 균주의 CFTase 유전자를 결손 변이시킨 고효율 효소 CFT108을 이용하여 cyclofructan (CF)의 대량생산 조건을 검토하고 생산된 CF의 순수 분리 정제 공정을 개발하였다. CF의 대량생산 조건은 $2\%$의 inulin 기질과 40 unit/g inulin의 기질에서 3시간 반응시켰을 때 최대 생산량 9.5 g/l의 수율을 달성할 수 있었으며, 이때의 in-ulin의 CF 전환율은 $47.5\%$였다. 생산된 CF를 순수 분리 정제하기 위해서 CFTase 반응액 을 exoinulinase 1 unit/ml로 6시간 처리하여 미 반응 inulin을 단당화시켰으며, 유리된 fructose는 $3\%$ CaO로 $CO_2$가스 하에서 $80^{\circ}C$, 10분간 3회 흡착제거 시킴으로써 순수 정제하였다. 정제된 CF를 HPLC로 확인한 결과 정제도는 $95\%$ 이상이였으며, $CF_6,\;CF_7,\;CF_8$ 비율이 72 : 27 : 1 이였다.