• Proceedings of the Korean Society of Applied Pharmacology
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• 1995.04a
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• pp.76-76
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• 1995

Virus-encoded HIV-1 reverse transcriptase (RTase) is one of the major targets for the development of drugs for HIV-1 since it is an essential enzyme-for the replication cycle of HIV-1. We cloned the entire reverse trancriptase gene into an inducible expression vector with tac promotor= RTase was stably overexpressed and induced by IPTG and the highly-expressed RTase was purified partially by use of DEAE cellulose and Mono Q column. The partially purified enzyme (663kDa, 51kDa) as exhibited by SDS-PAGE showed the high specific activity (16,570U/mg) when the assay for the RTase activity was carried out using $^3$H-dTTP and poly(rA): oligo(dT)12-18 as the substrate.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.150.3-151
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• 2003

To analyze the genome of Cucumber mosaic virus(CMV) in pepper, we developed a new extraction method for double-stranded RNA(dsRNA). To isolate the dsRNA, 0.1g of pepper leaves homogenized with 1ml of 5${\times}$EXB extraction buffer[0.5M glycin, 0.5M NaCl, 5mM EDTA(pH9.0/NaOH), 10％ Sodium N-lauryl salcosinate(NLS), 10％ Sodium dodecylsulfate(SDS)] and purified with the 1/4 volume of phenol： chloroform： isoamylalcohol(25：24：1). dsRNAs from the aqueous phase was precipitated with isopropanol. This procedure was able to detect a minimal amount of dsRNA from CMV infected plant tissue and to distinguish different CMV satellite RNAs by polyacrylamide gel electrophoresis(PAGE). Moreover, this method can be applied CMV infected in pepper or Rice dwarf virus (RDV) infected rice.

• Advances in environmental research
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• v.11 no.1
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• pp.1-16
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• 2022

Amongst 27 isolates from deteriorated leather samples, Arthrobacter creatinolyticus KP015744 zzx28 was found to be an efficient collagenase producer. Collagenase production of 13.33 µmoles/min was shown at an optimum temperature at 37℃ after 72h and at pH 7.5 by using 2 ml/dL inoculum in 10 mg/ml collagen peptide type I as a substrate. In presence of Hg²⁺, EDTA and 𝛽-mercaptoethanol the collagenase production by the isolate was strongly inhibited however Fe²⁺, Ca²⁺and DMSO enhanced production of the enzyme. Specific activity was found to be 19.46×10³ U/mg and molecular weight 66 kD by SDS PAGE. Isolate also has potential to hydrolyze keratin which is another important protein found in leather. Experimental results propose that collagenase can be effectively used as a tool for collagen and keratin rich solid waste treatment.

• Applied Biological Chemistry
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• v.36 no.3
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• pp.178-183
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• 1993

Xylose reductase (alditol: $NADP^+$ 1-oxidoreductase, EC 1.1.1.21) from the xylose-fermenting yeast, Candida sp. BT001, was purified via salt fractionation, ion-exchange, gel filtration and affinity chromatography, and its properties were characterized. The enzyme from the yeast was active with both NADPH and NADH as coenzyme. The xylose reductase activity with NADH was approximately 51% of that with NADPH and the specific activities of purified enzyme with NADPH and NADH were 11.78 U/mg and 6.01 U/mg, respectively. Molecular weight of the purified enzyme was 31,000 on SDS-PAGE and 61,000 on gel filtration. The Km for D-xylose, NADPH, and NADH was $94.2{\times}10^{-3}M,\;0.011{\times}10^{-3}M\;and \;0.032{\times}10^{-3}M$, respectively. The purified xylose reductase had relatively higher substrate affinity for L-arabinose than other aldoses tested. The optimal pH was 6.2 and the optimal reaction temperature was $45^{\circ}C$. The thermal stability of the enzyme was for 20 minutes at $30^{\circ}C$.

• Food Science of Animal Resources
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• v.34 no.4
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• pp.543-551
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• 2014

The aim of this study was to evaluate the effect of proteolytic (Serratia liquefaciens, match %: 99.39) or lipolytic (Acinetobacter genomospecies 10, match %: 99.90) psychrotrophic bacteria (bacterial counts, analysis of free fatty acids (FFA) and analysis of free amino acids) on the microbial and chemical properties (yogurt composition), and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of yogurt during storage. Yogurts were prepared with raw milk preinoculated with each psychrotrophic bacteria. The total solid, fat, and protein content were not affected by preinoculation, but the pH of yogurt preinoculated with psychrotrophic bacteria was higher than in control. There was a dramatic increase in short chain free fatty acids among FFA in yogurt with Acinetobacter genomospecies 10. For 14 d of cold storage condition, SCFFA was 25.3 mg/kg to 34.4 mg/kg (1.36 times increased), MCFFA was 20.4 mg/kg to 25.7 mg/kg (1.26 times increased), and LCFFA was 240.2 mg/kg to 322.8 mg/kg (1.34 times increased). Serratia liquefaciens (match %: 99.39) in yogurt caused a greater accumulation of free amino acids (FAA), especially bitter peptides such as leucine, valine, arginine, and tyrosine, but SDS-PAGE showed that the inoculation of Serratia liquefaciens did not affect the degree of casein degradation during storage. Taken together, the excessive peptides and FFA in yogurt generated from psychrotrophic bacteria could develop off-flavors that degrade the quality of commercial yogurt products.

• Korean Journal Plant Pathology
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• v.12 no.1
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• pp.99-108
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• 1996

3종의 빙핵세균 Peudomonas syringae 8401, Pseudomonas fuorescens 8701, Erwinia herbicola 8701의 세포 외막으로부터 아무런 변성제도 사용치 않고 sucrose density gradient centrifugation, Sephacryl gel filtration chromatography, DEAE-cellulose ion exchange chromatography, non-denaturing buffer를 이용한 PAGE, electroelution, SDS-PAGE를 통해 빙핵활성 단백질을 고도로 정제할 수 있었다. P. suringae와 P. fluorescens에서는 각각 3종류(155 kD, 75 kD, 50 kD)의 빙핵활성 단백질이, E. herbicola에서는 155 kD를 제외한 2종류(75 kD, 50 kD)의 빙핵활성 단백질은 이 연구를 통해 처음 밝혀진 것으로 , 지금까지 보고된 빙핵활성 단백질(150 KD 이상)보다는 훨씬 작은 것이다. 이는 빙핵활성을 나타내는 단백질의 기본단위는 이 실험의 결과만에 의하면 최대 50 kD임을 시사한다. 이들 단백질은 그 유래된 세균의 종류나 또는 단백질 분자량의 크기에 관계없이 모두 -5.5~7.5$^{\circ}C$에서 물을 동결시키는 높은 빙핵활성을 갖고 있었다. 이는 지금까지 보고된 어느 정제단백질의 빙핵활성보다 높은 것이다. 정제된 단백질의 빙핵활성은 trypsin 처리에 의해 상실되었고, pH6~8범위에서는 안정하였으며, pH5이하, pH9이상에서는 활성을 상실하였다. 보존온도에 대한 영향은 3$0^{\circ}C$이상이 되면 점차 활성이 감소하는 경향을 보이다 37$^{\circ}C$이상에서는 활성이 완전히 상실되었다. 금속이온으로서 Hg\ulcorner이온과 SDS에 의해 활성이 상실되었으나 phosphatidylinositol의 첨가에 의해서는 활성이 약간 증가(-1$^{\circ}C$)하였다.

• Journal of Microbiology and Biotechnology
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• v.8 no.3
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• pp.270-276
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• 1998

Extracellular ${\alpha}$-glucosidase was purified to homogeneity from moderately thermophilic Bacillus sp. DG0303. The thermostable ${\alpha}$-glucosidase was purified by ammonium sulfate fractionation, ion-exchange chromatography, preparative polyacrylamide gel electrophoresis (PAGE), and electroelution. The molecular weight of the enzyme was estimated to be 60 kDa by SDS-PAGE. The optimum temperature for the action of the enzyme was at $60^{\circ}C$. It had a half-life of 35 min at $60^{\circ}C$. The enzyme was stable at the pH range of 4.5~7.0 and had an optimum pH at 5.0. The enzyme preparation did not require any metal ion for activity. The thermostable ${\alpha}$-glucosidase hydrolyzed the ${\alpha}$-1,6-linkages in isomaltose, isomaltotriose, and panose, and had little or no activity with maltooligosaccharides and other polysaccharides. The $K_m$ (mM) for p-nitrophenyl-${\alpha}$-D-glucopyranoside (pNPG), panose, isomaltose, and isomaltotriose were 4.6, 4.7, 40.8, and 3.7 and the $V_{max}$(${\mu}mol{\cdot}min^-1$$mg^-1$) for those substrates were 5629, 1669, 3410, and 1827, respectively. The N-terminal amino acid sequence of the enzyme was MERVWWKKAV. Based on its substrate specificity and catalytic properties, the enzyme has been assigned to be an oligo-1,6-glucosidase.

• Journal of Aquaculture
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• v.15 no.1
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• pp.31-37
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• 2002

Effect of 2,4-Dichlorophenoxy acetic acid (2,4-D) on vitellogenin (VTG) production and estrogen ($E_2$)-estrogen receptor (ER) binding affinity were examined in primary hepatocyte cultures of rainbow trout, Oncorhynchus mykiss. Hepatocytes were pre-cultured for 2 days; subsequently, $E_2$( 2$\times$$10^{-6}$/ M) and 2,4-D ($10^{-9}~10^{-6}/M$) were simultaneously added to the incubation medium. They were cultured for more than 5 days. VTG and $E_2$-ER binding affinities were analyzed by SDS-PAGE and ELISA, respectively. 2,4-D concentration used had no appreciable effect on the morphology, viability, and DNA content of hepatocytes in culture. It had also no effect on VTG production. However, it interfered with $E_2$-ER binding affinity, which was reduced with increasing concentration of 2,4-D. The affinity was inhibited by 25 and 30% at $10^{-7}$ M and $10^{-6}$ M of 2,4-D, respectively. This result suggested that although 2,4-D had no effect on VTG production, it acted as reno-estrogenic contaminant in ER.

• KSBB Journal
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• v.11 no.3
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• pp.295-302
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• 1996

The measurements of rubisco parameters are important in photosynthetic studies. In this experiment, we used photometric assay method to detect these major parameters, such as activity, carbamylation and amount of rubisco. The main advantages of this method are very simple and as sensitive as conventional methods which usually produce radioactive waste. In this study, with kidney bean (Phaseolus vulgatis L.) leaves grown at normal $CO_2$ (350ppm) and high $CO_2$ (650 ppm), we investigated the effect of $CO_2$ concentration on activation and carbamylation of rubisco by measuring the rubisco activity, carbamylation rate and amount of rubisco using a dual beam (334nm and 405nm) spectrophotometer, and analyzed the polypeptide profiles of rubisco by SDS-PAGE. When $CO_2$ concentration was raised from 350ppm to 650ppm, all parameters of rubisco were decreased : $41.2{\mu}M/m^2/s and 52.2{\mu}M/m^2/s$ to $27.4{\mu}M/m^2/s and 46.1{\mu}M/m^2/s$ for initial and total rubisco activity, respectively ; from 79% to 58.9% for carbamylation rate ; from $1.94 {\mu}M/m^2$ to 1.58{\mu}M/m^2$ for amount of rubisco. These results suggests that the decrease in rubisco activity at high $CO_2$ was caused by carbamylation. The analysis of the preparation by SDS-PAGE showed two major polypeptides at 50 and 14.5 kD which were identified as the large and the small subunits of rubisco. There were no differences in the intensity compared high $CO_2$ to normal $CO_2$ in both 50 kD and 14.5 kD bands. We also found that these inhibitory effects of $CO_2$ were reversible. When high $CO_2$ was switched to normal $CO_2$, the parameters of rubisco changed were almost the same as normal rubisco parameters. These data provide an evidence that activity of rubisco was recovered by $CO_2$ concentration of 350 ppm.

• Parasites, Hosts and Diseases
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• v.26 no.4
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• pp.245-254
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• 1988

We studied the serological reaction between various antigenic components from Cysticercus cellulosae and IgG antibodies in sera of cysticercosis, sparganosis, hydatidosis patients and normal humans by ELISA and EITB. In serological tests by ELISA, we recognized cross reaction of Cysticercus antigenic components with IgG antibodies in heterologous sera such as sparganosis and hydatidosis patients or normal humans. The crude antigenic components of Cysticercus showed lower ELISA sensitivity in homologous sera from cysticercosis patients than heterologous sera from hydatidosis patients. A total of 31 polypeptide bands with 260 KDa~22 KDa molecular weights were detected by SDS-PAGE, and 11 of them showed strong intensity. Total 22 components of them were recognized by IgG antibodies in cysticercosis patients sera. However, 12 of them were recognized also by normal human sera, 11 were by sparganosis sera, and-21 were by hydatidosis patients sera. The crude antigenic components of 104 KDa, 82 KDa, 72 KDa, 59 KDa and 34 KDa molecular weights were nonspecific ones, which cross-reacted with sera of either cysticerco, =is, sparganosis, hydatidosis patients or normal humans.