• 한국과학교육학회지
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• 제40권4호
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• pp.437-449
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• 2020

이 연구에서는 예비 교사들이 갖고 있는 TPACK의 실천적 역량을 측정하기 위하여 기존의 연구들에서 사용한 TPACK 측정 설문지와 수행평가를 통해 드러난 TPACK 역량의 관계를 비교하였다. 이를 위하여 이 연구에서는 예비 교사의 TPACK 설문지를 개발하였다. 또한 '달과 천체의 변화'에 관련된 수행평가 과제 및 면담을 통해 예비 교사의 TPACK 역량의 실제를 분석하였다. 설문 분석 결과에서는 예비교사들의 PCK와 TPK이 높게 나타났고, 상대적으로 TCK와 TPACK이 낮은 것으로 나타났다. 그러나 설문에서는 TPK가 낮은 것으로 나타난 예비교사의 수행평가를 분석한 결과 TPK가 높은 것으로 나타났다. 반대로 설문에서는 TPK가 높은 것으로 나타난 예비교사의 수행평가를 분석한 결과 TPK는 낮은 것으로 나타났다. 또한 설문에서 TCK가 낮은 것으로 나타난 예비교사의 수행평가를 분석한 결과, 반대로 TCK가 매우 높은 것으로 나타났다. 반대로 설문에서 TCK가 높은 것으로 나타난 예비교사의 수행평가를 분석한 결과, 실제 TCK 역량은 낮은 것으로 나타났다. TPACK가 설문에서는 높게 나타난 경우에도 수행평가의 실제 결과를 통해서는 TPACK가 낮은 것으로 나타났다. 이를 통해 TPACK 측정 도구들이 실행 역량 보다는 자기효능감을 측정하는 것임을 확인하였다.

• Journal of Periodontal and Implant Science
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• 제35권2호
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• pp.427-436
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• 2005

Objective: MMP-8 is a neutrophil enzyme and its level increases in some inflammatory diseases, including periodontal disease. We knew that the lipopolysaccharide of E.coli(E-LPS) induced MMP-8 release from human neutrophils. E-LPS is known to induce the production and release of inflammatory cytokines through CD14, Toll-like receptor(TLR). In the present study, we investigated whether MMP-8 release by E-LPS is induced via CD14-TLR pathway and the cellular mechanism of MMP-8 release in human neutrophils. Material and methods: Human neutrophils were isolated from the peripheral blood of healthy donors and pre-incubated in medium containing antibodies against CD14, anti-TLR2 and anti-TLR4 or several inhibitors of microtubules and microfilaments and then incubated with E-LPS. The cells were treated TPCK and E-LPS simultaneously. The MMP-8amount in the culture medium was determined using ELISA. Results: E-LPS increased MMP-8release from neutrophils and its induction was inhibited by anti-CD14 and anti-TLR4 but not by anti-TLR2 antibodies. The inhibitors of microtubule and microfilament polymerization significantly decreased E-LPS-induced MMP-8release. TPCK inhibited E-LPS-induced MMP-8 release. Conclusion: These results suggest that MMP-8 release is induced by E-LPS via the CD14-TLR4 signal pathway in human neutrophils and may be depedent on microtubule and microfilament systems and $NF-{\kappa}B$ pathway.

• KSBB Journal
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• 제21권1호
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• pp.20-27
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• 2006

미생물 및 동물에 비해 식물에서는 혈전용해효소에 대한 연구가 부족한 실정이며, 기존의 혈전용해효소가 가지는 혈전에 대한 비특이적, 부작용, 고가 등의 단점을 해결할 수 있는 새로운 혈전용해효소의 개발을 위하여 율무, 홍화, 아욱의 종자로부터 추출된 수용성 단백질의 혈전용해 활성을 조사하였다. 각각의 식물들로부터 추출된 조효소 용액은 기존 혈전 용해효소인 plasmin과 양성 대조군으로 하여 비교하여 fibrin 평판법으로 확인한 결과 피브린 응집을 효과적으로 분해하였다. 그 중 율무종자의 수용성 추출물의 혈전용해 활성은 양성 대조군인 plasmin과 비교하여 1.3배의 높은 활성을 나타내었다. 전체 수용성 단백질은 50-75% 에탄올을 이용하여 농축하였으며 율무의 혈전용해효소는 fibrin zymography를 수행하여 확인하고 직접 추출하였다. SDS-PAGE에 의하여 추출효소의 분자량을 측정한 결과 7.8 kDa으로 단일 polypeptide임을 확인하였으며, 효소 활성에 미치는 온도의 효과는 $50^{\circ}C$ 이상에서는 비교적 안정하였으나 더 낮은 온도에서는 급격히 효소활성이 감소하였다. 또한, 각종 단백질분해효소 저해제에 의한 영향을 조사한 결과 APMSF, PMSF, pepstatin A 그리고 TPCK에 강력하게 저해되는 것으로 보아 추출효소는 chymotrypsin과 유사한 serine protease의 하나로 생각되었다. 그러나 EGTA와 EDTA 처리에 의해서는 효소활성의 저해가 두드러지게 나타나지 않았다. 더욱이, 종자저장 중에 미생물에 의한 부패, 활력저하, 생리활성물질의 감소와 장기저장에 따른 에너지소비 증가 등이 문제가 되고 있어 저선량의 감마선 조사를 통해 율무, 홍화, 아욱의 종자로부터 혈전용해 효소활성에 미치는 효과 및 선량에 따른 차이를 조사하였는데 비조사 종자인 대조구와 비교하여 1 Gy, 4 Gy, 16 Gy, 32 Gy선량에서는 낮은 활성을 보였으면 반면에 8 Gy와 64 Gy의 선량에서는 더 높은 활성을 나타내었다. 이러한 결과는 Y선 조사가 종자의 혈전용해 활성을 향상시킬 가능성이 있을 것으로 생각된다. 이상의 모든 결과로 볼 때 율무의 추출 효소는 chymotrypsin-like serine protease에 속하는 혈전용해효소임을 확인할 수 있었다.

• 미생물학회지
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• 제20권4호
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• pp.189-194
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• 1982

Acid protease produced from Aspergillus tubingensis was pruified by ethanol fractionation, dialysis, and DEAE cellulose column chromatography. As a result of purification its specific activity increased to 5.4 times, and percent recovery was 39. The kinetic constants of the enzyme were studied. Km and Vmax was $1.5{\times}10^{-7}M\;and\;0.11{\Delta}O.D/min$ , respectively, when casein was used as substrate. The order of Km value of several proteins is : casein

• Fisheries and Aquatic Sciences
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• 제2권2호
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• pp.218-225
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• 1999

A protease without tryptic and chymotryptic activities was purified from the hepatopancreas of shrimp, Penaeus orientalis, using Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, Mono-Q, and gel chromatography. Molecular weight (M.W.) of the protease was estimated to be 27kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS­PAGE). The amino acid composition of the protease was different from that of protease from P. japonicus or trypsin from P. orientalis. The protease was completely inhibited by benzamidine, $N\alpha-p-tosyl-L-lysine$ chloromethyl ketone (TLCK), and phenylmethylsulfonyl fluoride (PMSF) and was not affected by leupeptin, pepstatin, N-tosyl-L-phenylalanine chloromethyl ketone (TPCK), iodoacetate, and ethylenediamine tetra acetate (EDTA). The enzyme did not have any activity against Na-benzoyl-DL-arginine p-nitroanilide (BAPNA) or N-benzoyl-L-tyrosine ethyl ester (BTEE) which are specific substrates of trypsin and chymotrypsin, respectively. However, the protease showed hydrolytic activity for a carboxyl terminal of Tyr, Trp, Phe, Glu, and Cys.

• 한국수산과학회지
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• 제29권6호
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• pp.797-804
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• 1996

Two trypsins were purified from shrimp hepatopancreas through ammonium sulfate fractionation, Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, and Sephacryl S-300 gel chromatography. Both enzymes had a single polypeptide chain with a molecular weight (M.W.) of 32 kDa by sodium dodecylsulfate polyacrylamide gel electrophoresis (SOS-PAGE), although trypsin A and B were estimated to be a molecular weight of 27.2 and 22.8 kDa, respectively, using Sephacryl S-300 gel filtration. Both trypsins had similar amino acid compositions and rich in glycine, valine, alanine, aspartic acid, and glutamic acid, but low in methionine and basic amino acids. Both enzymes were completely inactivated by soybean trypsin inhibitor (SBTI), phenylmethylsulfonyl fluoride (PMSF), tosyl-L-lysine chloromethyl ketone (TLCK), benzamidine, leupeptin, however, not affected by tosyl-L-phenylalanine chloromethyl ketone (TPCK) and pepstatin.

• Fisheries and Aquatic Sciences
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• 제1권2호
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• pp.209-215
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• 1998

Trypsin-like enzyme was purified from shrimp hepatopancreas through Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, and Superdex 75 gel chromatography. Purity of trypsin-like enzyme was increased 69-fold with $44\%$ yield. The enzyme consisted of a single polypeptide chain with a molecular weight (M.W.) of 32 kDa judged by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was completely inactivated by serine enzyme inhibitors such as soybean trypsin inhibitor (SBTI), tosyl-L­lysine chloromethyl ketone (TLCK), and leupeptin. However, the enzyme was not affected by tosyl-L-phenylalanine chloromethyl ketone (TPCK) which is a chymotrypsin specific inhibitor. The enzyme had no activity against benzoyl-tyrosine ethyl ester (BTEE) which is a chymotrypsin specific substrate. The enzyme showed high activity on the carboxyl terminal of Phe, Tyr. Glu, Arg, and Asp. However. no activity was detected against the carboxyl terminal of Pro, Trp, Cys, Gly, Val, and Ala.

• 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.

• 사상체질의학회지
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• 제19권1호
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• pp.160-170
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• 2007

1. Objectives This study was performed to find the activities and characteristics of purified thrombolytic enzymes from Holotrichia extracts. 2. Methods In the first time, a coarse enzyme fluid was made by using the freedried Holotrichia extracts. After manufacturing total soluble proteins and purifing enzymes, it was evauluated the activities and characteristics of this enzyme's dissolving capability to fibrin and thrombus. This study was taken using azocasein assay, fibrin-plate method, native-PAGE and fibrin zymography. 3. Results A soluble proteins were efficiently extracted form freezedried Holotrichia extracts. And, this purified enzyme had a ten times fibrinolytic capability compare with ustulation Holotrichia sample. In native PAGE and fibrin zymography, Holotrichia extracts showed the respectable fibrinolytic activity. Also, It had higher thrombolytic activities compared with general thrombolytic enzyme 'plasmin'. In experiment of various protease inhibitors of the purified enzyme from Holotrichia extracts on the azocaseinolytic activity, the enzyme was strongly inhibited by EDTA ${\cdot}$ EGTA, and weakly by APMSF ${\cdot}$ PMSF ${\cdot}$ TPCK. 4. Conclusion Holotrichia extracts has the thrombolytic activities, and it will operate directly th fibrin-clot and thrombus.

• 대한한의학회지
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• 제24권2호
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• pp.138-147
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• 2003

Objectives : In this study, we investigated the mechanism by which Chelidonium majus (CM) regulates nitric oxide (NO) production. Methods : Using mouse peritoneal macrophages, the mechanism by which CM regulates NO or tumor necrosis $factor-{\alpha}(TNF-{\alpha})$ production was examined. NO release was measured by the Griess method. $TNF-{\alpha}$ production was measured by the ELISA method. The protein extracts were prepared and samples were analyzed for the inducible NOS(iNOS) expression and nuclear factor kappa $B(NF-{\kappa}B)$ activation by Western blotting. Results : When CM was used in combination with recombinant $interferon-{\gamma}{\;}(rIFN-{\gamma})$, there was a marked cooperative induction of NO production. CM had an effect on NO production by itself. The expression of the iNOS gene was increased in $rIFN-{\gamma}$ plus CM-stimulated peritoneal macrophages and almost completely inhibited by pre-treatment with pyrrolidine dithiocarbamate (PDTC), an inhibitor of $NF-{\kappa}B$. The $NF-{\kappa}B$ activation was increased in rIFN-{\gamma} plus CM-induced peritoneal macrophages. The increased production of NO from $rIFN-{\gamma}$ plus CM-stimulated peritoneal rnacrophages was decreased by the treatment with $N^{G}-monomethyl-{_L}-arginine{\;}(N^{G}MMA){\;}N^{\alpha}-Tosyl-Phe$ chloromethyl ketone (TPCK) , and was almost completely inhibited by pre-treatment with PDTC. Furthermore, treatment with CM alone or rIFN-{\gamma} plus CM in peritoneal macrophages caused a significant increase in $TNF-{\alpha}$ production. PDTC decreased CM-induced $TNF-{\alpha}$ production significantly. After CM treatment in HT-29 or AGS cells, cell viability decreased. Conclusions : These findings demonstrate that CM increases the production of NO and $TNF-{\alpha}{\;}by{\;}rIFN-{\gamma}-primed$ macrophages and suggest that NF-B plays a critical role in mediating these effects of CM.