• 제어로봇시스템학회:학술대회논문집
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• 제어로봇시스템학회 2003년도 ICCAS
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• pp.2356-2361
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• 2003

Proteome research in the medical field is expected to accelerate the understanding of disease mechanism, and to create new diagnostic concept. For protein profiling, this paper proposes a new methodology named CPDIB (Crude Protein Direct Blotting). In the CPDIB procedure, crude protein sample is directly immobilized on a membrane and the expression of protein molecules in the sample are analyzed quantitatively by using a special device called ImmobiChip, where the membrane is used as a field of the immune reaction. The over-all structure of the ImmobiChip is based on the conventional Slot blot device. Mechanical improvement in the air-tightness of the case holding the membrane realizes the direct blotting and results in high performance of stability in the immune reaction. In the measurement of multiple proteins, a dispensing robot is used for increasing the efficiency of handling of liquid. Cooperation of the dispensing robot with the ImmobiChip for immobilizing proteins realizes automated and stable performance of the CPDIB procedure. This paper shows the evaluation of the air-tightness of the ImmobiChip, the ability of analyzing proteins using the CPDIB procedure and the performance of the automated equipment.

• 미생물학회지
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• 제41권4호
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• pp.306-311
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• 2005

Lumazine protein은 lux operon의 하류 영역에 존재하는 riboflavin synthase와 아미노산 상동성을 보일 뿐만 아니라, riboflavin synthase의 기질인 6,7-dimethyl-8-ribityllumazine (lumazine)과 결합하여 청록색의 형광을 내게 하는 형광 단백질이다. 발광세균 Photobacterium phosphoreum의 lumazine protein을 코드하는 유전자의 염기서열을 결정하였는데, 이 유전자는 lux operon의 656 bp 상류의 영역에 존재하며, lux operon과는 서로 반대 방향으로 전사되는 것으로 나타났다. 중합효소 연쇄 반응(PCR: Polymerase Chain Reaction)의 방법으로 아미노-말단 절반 lumazine protein을 코드하게 되는 유전자(lumP-N)를 클로닝하여 형질전환의 방법으로 대장균에 유전자를 전이시켜 이들의 유전자의 발현 양상을 조사하여 보았는바, lumP 전체 유전자(lumP-W)가 삽입되어 있는 재조합 플라스미드에서는 발현이 매우 미약한 반면에 아미노 -말단(lumP-N)이 들어있는 경우는 과발현됨을 보였다.

• Journal of Microbiology
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• 제37권4호
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• pp.234-242
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• 1999

To obtain the recombinant circumsporozoite (CS) protein for the diagnosis of patients and seroepidemiology of Plasmodium vivax malaria which have been prevalent in northern part of Kyonggido, the CS protein gene was amplified by the polymerase chain reaction (PCR) from genomic DNA of the Korean vivax malaria patient. The gene consists of 1,123 nucleotides except signal peptide sequences and had an uninterrupted reading frame encoding a protein of 374 amino acids with a central region of 20 tandem repeats of the nonapeptide. The CS protein gene was expressed in Escherichia coli and purified, the molecular weight of recombinant CS protein was about 44 kDa (monomer) under denaturing purification and about 65 kDa (dimer) under native purification by SDS-PAGE. The purified recombinant CS protein which has antigenicity to malaria patients in Western blot analysis and Enzyme-linked immunosorbent assay, reacted only with the serum of P. vivax (PV210) infected malaria patients with no cross reaction to the P. falciparum malaria patient. The recombinant CS protein purified in this study will serve as a useful antigen to support the diagnosis of malaria patients and seroepidemiology.

• Archives of Pharmacal Research
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• 제11권2호
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• pp.93-98
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• 1988

To elucidate the reaction mechanism of ginseng protein on its antiradiation activity, its effects were studied on sister chromatid exchanges (SCE) induced by UV irradiation in CHO-KI cells. When cells were irradiated with 254 nm UV light at the dose of 0 to 8erg$\textrm{mm}^2$, the frequencies of CSE were increased more than two fold. However, when radio protective ginseng protein was added to the cells before the after UV irradiation, SCE frequencies were decreased significantly at all UV doses in both cases with no significant differences. As the amount of ginseng protein was varied from 100 to 500 .mu.g/ml, with UV irradiation at 60 erg$\textrm{mm}^2$, SCE frequencies dropped sharply at the first two concentrations and then reached a sort of plateau in both cases of pre-and post-treatment. When the ginseng protein was treated alone without UV irradiation, there were no changes in SCE frequencies no matter when the protein was added. There results suggest that the ginseng protein could reduced DNA damages, which may play an important role in the reaction mechanism of radioprotective activity of the protein.

• 한국식품과학회지
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• 제31권5호
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• pp.1262-1267
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• 1999

겔 형성이 어려운 유채단백질의 겔화를 위하여 미생물성 TGase의 촉매작용에 의하여 최적 겔화 반응조건을 겔강도(g)와 겔변형(mm)을 조사하여 검토하였다. 또한 형성된 겔의 고분자화를 SDS-PAGE 및 GL교차결합 함량 측정에 의하여 확인하였다. 단백질-TGase반응에서 효소 첨가량은 $45^{\circ}C$에서 60분 동안 반응 시 단백질농도가 10%에서 효소가 1 : 40([E] : [S])의 비로 첨가되었을 때 겔강도와 변형 값이 가장 높았다. 또한 이 조건에서 기질단백질 농도가 $4{\sim}l2%$로 증가함에 따라 겔강도는 비례적으로 증가함을 보였으나 겔의 변형을 고려할 때 10% 농도가 유채단백질 겔 제조에 적당한 농도로 조사되었다. 최적 활성을 나타내는 반응온도는 $45^{\circ}C$로 관찰되었고, 반응시간은 90분이면 적당한 것으로 조사되었다. 겔강도와 변형은 다른 pH와 비교할 때 pH 7에서 상당히 높은 값을 보여 pH 7이 최적조건이었다. SDS-PAGE분석에서 효소반응 에 따라 유채단백질의 본래 밴드들이 소별되거나 희미해지면서 고분자 밴드들이 형성되고 있음을 보여주었다. 또만 HPLC 분석 결과 GL교차결합 함량이 반응시간에 따라 $0\;{\mu}mol/g$에서 최고 $7.14\;{\mu}mol/g$ gel(반응 90분)까지 증가되고 있어 유채단백질의 겔화가 TGase에 의해 이루어지고 있다는 것을 확인할 수 있었다.

• BMB Reports
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• 제29권1호
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• pp.81-87
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• 1996

The relationship of S-thiolation and oxidation of glycogen phosphorylase b and peroxidation of phosphatidyl choline liposome by xanthine oxidase (XOD), 2,2'-azobis(2-amidinopropane) hydrochloride (AAPH), and 2,2'-azobis(dimethylvaleronitrile) (AMVN)-generated free radicals was investigated, Glycogen phosphorylase b was S-thiolated in the presence of glutathione and oxidized in the absence of it by XOD, AAPH and AMVN. In XOD-initiated reaction, the rates of S-thiolation and oxidation of phosphorylase were very similar and addition of liposome to the reaction mixture showed little inhibition of the modifications. In AAPH-initiated reaction, the rate of oxidation was higher than that of S-thiolation and addition of liposome increased oxidation of the protein but had no effect on S-thiolation. In AMVN-initiated reaction, S-thiolation was higher than oxidation and addition of liposome increased S-thiolation remarkably but showed no effect on oxidation. The effect of liposome on modifications of protein in AAPH and AMVN reaction seemed to be caused by certain reactive degradation products or intermediates of liposome by free radical attack. Peroxidation of liposome was not observed in XOD-initiated reaction. Liposome was gradually peroxidized by AAPH reaction. The peroxidation was inhibited by addition of GSH and phosphorylase. Peroxidation of liposome by AMVN was extreamly fast, and was not affected by GSH and phosphorylase.

• Preventive Nutrition and Food Science
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• 제2권4호
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• pp.321-327
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• 1997

High Temperature-cooking conditions of cultured fishes(loach, crucian carp, bastard halibut, and jacopever) were optimized by response surface methodology(RSM), and plastein products were prepared using enzymatic hydrolysis. Four models were proposed with regard to effects of time(t), temperature(T), and water/fish meat (w/f) ratio on the amount of 0.3M TCA soluble fractions. The model coefficients were ranged from p<0.0001 for jacopever to p<0.0433 for bastared halibut. Cooking conditions for 60% hydrolysis were optimized at 1) 14$0^{\circ}C$ except for crucian carp(136$^{\circ}C$); 2) 10.08 hours(loach), 7.25 hours(crucian carp), 9.85 hours(ba-stard harlibut), and 9.37 hours(iacopever); 3) 1:1(w/f) ratio except for the crucian carp(1.1:1). When protein hydrolyzates were employed for the plastein synthesis, optimum plastein-reaction conditions were determined to be pH 9.0 with chymotrypsin for the loach and crucian carp hydrolyzates, pH 9.0 with papain for the bastard halibut hydrolyzate, and pH 11.0 with trypsin for the jacopever hydrolyzate. Plastein reaction could be performed in water at concentration up to 20%(w/f).

• Toxicological Research
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• 제13권3호
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• pp.223-228
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• 1997

Quinones have been reported to undergo nonenzymatic reaction with thiols to generate reactive oxygens. It is therefore possible that the nonenzymatic reaction of quinones with thiols in plasma could lead to potentJared cellular toxicity or disease. When 1 mM menadione was added in plasma under pH 11.2, 7.4 and 5.0, the increase in oxygen consumption rate was the order of pH 11.2 > pH 7.4 > pH 5.0. In addition, oxygen consumption rates under plasma anticoagulated with trisodium citrate solution (pH 7.85) was significantly higher than those with acid-citrate-dextrose solution (pH 6.87). SOD and catalase reduced the rate of oxygen consumption induced by menadione in plasma. Taken together, these results suggest that the menadione-induced increased oxygen consumption was due to nonenzymatic reaction of menadione with thiols in the plasma. The presence of plasma has an additive effect on the increased oxygen consumption rates induced by the menadione treatments on our model tissue, platelets, as compared between washed platelet (WP) and platelet rich plasma (PRP). Cytotoxicity, as determined by LDH release, are well correlated with the oxygen consumption rates observed in each system and strongly suggest that menadione-induced cytotoxicity can be increased with the presence of blood plasma.

• Journal of Animal Science and Technology
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• 제64권2호
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• pp.235-246
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• 2022

In this study, we used more reliable experimental materials and methods to detect the effects of osteopontin (OPN) on boar sperm in vitro capacitation, acrosome reaction, and fertilization efficiency. We reorganized and obtained the OPN protein of the porcine source. Immunofluorescence and Western blot show the localization and expression of the OPN protein before and after sperm capacitation. To determine whether OPN can affect sperm during sperm capacitation, we examined cyclic adenosine monophosphate (cAMP) concentrations after sperm capacitation, and the results showed that OPN significantly increased the cAMP concentration in sperm (p < 0.05). Flow cytometry showed that 0.1 ㎍/mL OPN-treated sperm had better acrosome reaction ability. In vitro fertilization (IVF) showed that 0.1 ㎍/mL OPN significantly increased the rate of embryo division. In conclusion, this study found that 0.1 ㎍/mL porcine OPN protein can significantly improve porcine capacitated sperm motility, cAMP concentration after capacitation sperm, acrosome reaction ability, and embryo division during IVF and provides new clues to explore the mechanism of OPN's function on sperm.

• 한국식품과학회지
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• 제46권3호
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• pp.315-322
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• 2014

본 연구는 미생물로부터 유래된 5종(AH, AK, AY, PS와 R)의 lipase들을 Amberlite XAD 7에 흡착법으로 고정화 시킨 후 각 immobilized lipase들의 특성을 알아보았다. 고정화 전과 후의 단백질 함량 및 각 free lipase들과 immobilized lipase들을 이용한 interesterification 반응물의 지방산과 TAG 조성을 분석하였다. 또한, immobilized lipase에 있어 중요한 요인인 reusability를 확인하였다. Free lipase의 단백질 함량은 2.22-11.41%로 AH가 가장 높았던 반면, immobilized lipase에서는 AH, PS와 AK가 mg protein/g support이 높았다. 한편, 반응 특성을 알아보기 위해 카놀라유, PEE와 StEE를 기질로 하여 batch type interesterification을 진행하였을 때, free lipase의 경우 free lipase R을 제외한 다른 free lipase들은 반응시간이 1시간에서 72시간으로 증가함에 따라 총 포화지방산 함량이 증가하였으며 그 중 free lipase AH가 반응성이 가장 높았다. 또한, RP-HPLC를 통해 free lipase AK 반응물을 분석한 결과, 반응시간이 길어질수록 카놀라유(0시간)에서 볼 수 있었던 57.49 area%의 ${\bigcirc}{\bigcirc}{\bigcirc}$가 6.53 area%로 감소하였다. 이는 각 free lipase들이 PEE와 StEE를 효소적 반응에 이용했기 때문이라고 판단된다. 한편, immobilized lipase AY와 R의 경우 반응시간이 1시간에서 48시간으로 증가하여도 카놀라유(0시간)의 총 포화지방산 함량과 큰 차이가 없었으나 immobilized AK의 경우 48시간에서 38.3 area%의 포화지방산 함량으로 가장 높은 활성을 보였다. 또한, 이를 사용하여 continuous type으로 반응하였을 때 유속이 느릴수록 효소와 기질 사이의 접촉 시간이 길어져 반응물의 총 포화지방산 함량이 증가함을 알 수 있었다. Reusability는 immobilized AK, AH와 PS 모두에서 두 번째 반복 반응을 하였을 때, 첫 번째 반응보다 총 포화지방산이 120-196.5% 증가하였다. 그러나 bounding protein 함량이 가장 높았던 immobilized AK는 support에 흡착되었던 free lipase AK의 탈착이 일어나 2번째 반응 후부터 활성이 감소한 반면, immobilized AH와 PS는 활성이 비교적 유지되었다.