• Title/Summary/Keyword: zymogram

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Esterase Isozyme Patterns of Rice-off Ope Groups Classified by Grain Characteristics (종실특성에 따라 분류한 벼 이형주군의 Esterase 동위효소 형태변이 및 분리양상)

  • 김동관;진일두;정병관
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.47 no.3
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    • pp.206-210
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    • 2002
  • This study was carried out to investigate the genetic segregations and characteristics of off-type rice plants collected in Korea which were classified into seven groups based on grain characteristics. In the analysis of esterase electrophoresis, the long-grain red group was classified as 1 and 3 esterase isozyme zymogram(EIZ), the long-grain normal group was classified as 1, 3 and 7 EIZ. The extremely late sterility group was segregated variously as 1, 2, 1+2, 5, 6, 5+6, 7,8 ,7+8 and 12 EIZ. The long-grain red rice lines with 1 EIZ had a longer culm length and a lower length/width ratio to brown rice than the long-grain red rice lines with 3 EIZ. The long-grain normal rice lines with 3 EIZ had a longer culm length, shorter panicle length, greater number of tillers, lower length/width ratio of brown rice, and fewer number of grains per panicle than did the long-grain red rice lines with 1 or 7 EIZ.

Purification and Characterization of Carboxymethyl Cellulase IV from Penicillium verruculosum (Penicillium verruculosum 으로부터 Carboxymethyl Cellulase IV 의 정제(精製) 및 특성(特性))

  • Kim, Jeong-Ho;Lee, Jae-Chang;Lee, Yong-Kyu;Kim, Kang-Hwa;Chun, Soon-Bai;Chung, Ki-Chul
    • The Korean Journal of Mycology
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    • v.21 no.1
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    • pp.28-37
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    • 1993
  • An endo-type cellulase, carboxymethyl cellulase(CMCase) IV, was purified from culture filtrate of cellulolytic fungus Penicillium verruculosum. The CMCase IV was acidic glycoprotein having carbohydrate of 13% as glucose and pI value of 4.0. The CMCase IV was 52 KDa of molecular weight in SDS-polyacrylamide gel electrophoresis and have optimum temperature and pH of $50^{\circ}C$ and 5.0 for enzyme activity. The CMCase IV liberated glucose and cellobiose as major products of the enzyme against carboxymethyl cellulose(CMC) and seemed to has transglycosylation activity simultaneously. Cellulase activity staining(zymogram) showed that the cellulase components of P. verruculosum were not aggregated in the medium. P. vrttuculosum mRNA was translated in vitro and translation product by the mRNA coding for CMCase IV was identified by immunoprecipitation.

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Purification and Characterization of Cyclodextrin Glycosyltransferase from Bacillus brevis CD162 (Bacillus brevis CD162 Cyclodextrin Glycosyltransferase의 정제 및 특성)

  • Kim, Myung-Hee;Lim, Young-Hee;Oh, Tae-Kwang;Sohn, Cheon-Bae
    • Applied Biological Chemistry
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    • v.40 no.6
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    • pp.465-471
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    • 1997
  • The cyclodextrin glycosyltransferase (CGTase, EC 3.2.1.19) from Bacillus brevis CD162 was purified by precipitating with ammonium sulfate, DEAE-Sepharose CL-6B column chromatography and Sephadex G-150 column chromatography. The molecular mass and pI of the purified enzyme were estimated to be 74,000 and 6.3 by SDS-PAGE and isoelectric focusing, respectively. The purified enzyme was clearly identified as the CGTase by zymogram after SDS-PAGE. The optimum pH and temperature for the enzyme activity were 8.0 and $55^{\circ}C$, respectively. The enzyme was stable at the range of pH $5.5{\sim}9.0$, and up to $50^{\circ}C$. The amino acid sequence from the $NH_2-terminal$ of the purified CGTase was Ser-Val-Thr-Asn-Lys-Val-Asn-Tyr-Ser-Lys-Asp-Val-Ile-Tyr-Gln. The yields of the products from starch as the substrate were 1.3% for ${\alpha}-$, 33.9% for ${\beta}-$, and 9.7% for ${\gamma}-cyclodextrin$.

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Optimization of \beta-mammanase Production from Bacillus subtilis JS-1. (\beta-Mannanase를 생산하는 Bacillus subtilis JS-1의 분리 및 효소 생산성)

  • 임지수;정진우;이종수;강대경;강하근
    • Microbiology and Biotechnology Letters
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    • v.31 no.1
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    • pp.57-62
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    • 2003
  • A bacteria strain producing extracellular $\beta$-mannanase was isolated from soil and was identified as Bacillus subtilis by 16S rRNA sequence comparison and biochemical determinations. The optimum pH and temperature for the $\beta$-mannanase activity were 5.0 and 5.5$^{\circ}C$, respectively. The zymogram technique revealed a single protein band exhibiting $\beta$-mannanase activity from the culture supernatant. The molecular mass of the enzyme was estimated at approximately 130 kDa. The addition of 0.5% lactose or 0.5% locust bean gum to the LB medium caused to Increase significantly the $\beta$-mannanase productivity from Bacillus subtilis JS-1. The cells grown on LB medium supplemented with lactose produced maximal enzyme activity at the stationary phase. In contrast to this, the $\beta$-mannanase was induced at the logarithmic phase from the cells grown on LB medium supplemented with locust bean gum. The discrepancy in induction times suggests that $\beta$-mannanase was induced by different induction mechanisms depending on the carbon sources in Bacillus subtilis JS-1 .

Molecular Cloning and Expression of an Endo-xylanase Gene from Bacillus stearothermuphilus into Escherichia coli (Bacillus stearothermophilus로 부터 Endo-xylanase 유전자의 클로닝 및 Escherichia coli에서의 발현)

  • 조상구;박성수;박영인;최용진
    • Microbiology and Biotechnology Letters
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    • v.20 no.3
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    • pp.271-279
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    • 1992
  • Genomic DNA of Bacillus stearothemzophilus, which expressed alkalophilic and thermophilic xylanases, was partially digested with HindIII, cloned into pBR322, and subsequently transferred into the Escherichra coli HB101 cells. Three among 5, 000 transformants screened formed clear zones around their colonies. From the functional clones, three recombinant plasmids (pMG11, pMG12 and pMG13) had been isolated, and they were identified to carry the same 4 kb HindIII fragment originated from B. stearothemzophilus which was responsible for the xylanase activity. pMGl3, however, had the foreign DNA of opposite orientation compared to the other two recombinant plasmids. This recombinant plasmid gave much lower xylanase activity. B. stearothermophilus was observed to produce at least three xylanase activities as evidenced by the PAGE-xylan zymogram. The xylanase from E. coli HB101/pMG12 was judged to correspond to the largest among the three B. stearothermophilus xylanases observed in the zymogrom. The enzyme hydrolyzed xylooligosaccharides larger than xylotriose and degraded xylan to produce xylobiose and xylotriose as major products. The xylanase was considered to have trans-xylosidase activity, too.

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Effects of (-)-Epigallocatechin-3-gallate on Brain Infarction and the Activity Change of Matrix Metalloproteinase-9 Induced by Middle Cerebral Artery Occlusion in Mice

  • Qian, Yong-Ri;Kook, Ji-Hyun;Hwang, Shin-Ae;Kim, Do-Kyung;Kim, Jong-Keun
    • The Korean Journal of Physiology and Pharmacology
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    • v.11 no.3
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    • pp.85-88
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    • 2007
  • Matrix metalloproteinases (MMPs) can degrade a wide range of extracellular matrix components. It has been reported that MMP-9 are activated after focal ischemia in experimental animals. (-)-Epigallocatechin-3-gallate (EGCG), a major constituent of green tea polyphenols, is a potent free radical scavenger and reduces the neuronal damage caused by oxygen free radicals. And it has been known that EGCG could reduce the infarction volume in focal brain ischemia and inhibit MMP-9 activity. To delineate the relationship between the anti-ischemic action and the MMP-9-inhibiting action of EGCG, we investigated the effect of EGCG on brain infarction and the activity of matrix metalloproteinase-9 induced by permanent middle cerebral artery occlusion (pMCAO) in ICR mice. EGCG (40 mg/kg, i.p. $15{\sim}30min$ prior to MCAO) significantly decreased infarction volume at 24 hr after MCAO. GM 6001 (50 mg/kg, i.p. $15{\sim}30min$ prior to MCAO), a MMP inhibitor, also significantly reduced infarction volume. In zymogram, MMP-9 activities began to increase at ipsilateral cortex at 2 hr after MCAO, and the increments of MMP-9 activities were attenuated by EGCG treatment. Western blot for MMP-9 also showed patterns similar to that of zymogram. These findings demonstrate that the anti-ischemic action of EGCG ire mouse focal cerebral ischemia involves its inhibitory effect on MMP-9.

Isolation and Characterization of Endocellulase-Free Multienzyme Complex from Newly Isolated Thermoanaerobacterium thermosaccharolyticum Strain NOI-1

  • Chimtong, Suphavadee;Tachaapaikoon, Chakrit;Pason, Patthra;Kyu, Khin Lay;Kosugi, Akihiko;Mori, Yutaka;Ratanakhanokchai, Khanok
    • Journal of Microbiology and Biotechnology
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    • v.21 no.3
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    • pp.284-292
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    • 2011
  • An endocellulase-free multienzyme complex was produced by a thermophilic anaerobic bacterium, Thermoanaerobacterium thermosaccharolyticum strain NOI-1, when grown on xylan. The temperature and pH optima for growth were $60^{\circ}C$ and 6.0, respectively. The bacterial cells were found to adhere to insoluble xylan and Avicel. A scanning electron microscopy analysis showed the adhesion of xylan to the cells. An endocellulase-free multienzyme complex was isolated from the crude enzyme of strain NOI-1 by affinity purification on cellulose and Sephacryl S-300 gel filtration. The molecular mass of the multienzyme complex was estimated to be about 1,200 kDa. The multienzyme complex showed one protein on native PAGE, one xylanase on a native zymogram, 21 proteins on SDS-PAGE, and 5 xylanases on a SDS zymogram. The multienzyme complex consisted of xylanase, ${\beta}$-xylosidase, ${\alpha}$-L-arabinofuranosidase, ${\beta}$-glucosidase, and cellobiohydrolase. The multienzyme complex was effective in hydrolyzing xylan and corn hulls. This is the first report of an endocellulase-free multienzyme complex produced by a thermophilic anaerobic bacterium, T. thermosaccharolyticum strain NOI-1.

A Systematic Study on the Selected Amentiferous Plant Taxa -On the Genus Quercus- (유이화서 식물군의 계통분류학적 연구 -참나무과를 중심으로-)

  • Lee, Yoo Sung
    • Journal of Plant Biology
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    • v.27 no.2
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    • pp.61-72
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    • 1984
  • The concept of natural grouping of plant designated as the "Amentiferous" is no longer given serious credence, and many of the families included in this grouping have been dispersed in diverse order. Because a review of taxonomic treatments of amentiferous taxa reveals diverse classifications, it has become necessary to investigate new characteristics and attempt to determine the significance of these characteristics in terms of amentiferous taxonomy. Protein analyses by isoelectrofocusing(IEF) and rocket immunoelectrophoresis(RIE) have proved to be useful in the delicitation of Quercus taxa using pollen extracts from selected taxa. When Quercus pollen extracts were separated by electrophoresis based on their isoelectric points in a stable pH gradient and substrates for estrase activity were stained, ten bands were revealed between pH 5-14. Within Lepidobalanus grouping, a great diversity in the pollen protein zymograms was observed with some segregation corresponding to the designated taxonomic sections. Two taxa of Cyclobalanopsis produced a zymogram that is somewhat similar to taxa included within the section Prinus of Lepidobalanus, and less similar to taxa within the section Cerris of the same subgenus. Three tested taxa of the Cerris are in the similar zymogram each other, being segregable from the taxa of Prinus. Quantitative and qualitative analyses for serological relationships within and among th Quercus were also employed. To calculate the degree of protein similarity, total rocket heights obtained from RIE provided an index of serological correspondence(SC). It is reconfirmed that the Quercus is distantly separated from the Fagus according to SC. Comparative data from rocket number and SC in the tested taxa of Quercus also indicate that Lepidobalanus is separable from Cyclobalanopsis. Within the Lepidobalanus Q. acutissima and Q. acutissima x variabilis are almost homogeneors and distinguishable from the other tested taxa of same subgenus. Although the number of taxa tested has been limited, the overall serological evidence best reflects the classification proposed by Redher(1940) and Melchior (1964), having the genus Quercus subdivided into three subgunera: Erythrobalanus, Lepidobalanus, and Cyclobalanopsis.alanopsis.

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Purification and Characterization of Carboxymethyl Cellulase from Bacillus stearothermophilus No. 236

  • Kim, Sohng-Hwan;Cho, Ssang-Goo;Choi, Yong-Jin
    • Journal of Microbiology and Biotechnology
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    • v.7 no.5
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    • pp.305-309
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    • 1997
  • Bacillus stearothermophilus No. 236, an effective xylanolytic bacterium, produced an extracellular carboxymethyl cellulase when the strain was grown on xylan. The carboxymethyl cellulase was purified to homogeneity as judged by SDS-PAGE and zymogram, The carboxymethyl cellulase had a pI of 4.0, and a molecular mass of 95 kDa. The highest level of enzyme activity was observed at pH 6.5 and $60^{\circ}C$. The $K_m$, and $V_{max}$ values of the enzyme to carboxymethyl cellulose were 20.8 mg/ml and $0.63 {\mu}mole$/min/mg protein, respectively, The enzyme was found to act also on filter paper and xylan as well as carboxymethyl cellulose. Therefore, it is expected that this xylanolytic strain isolated from soil could be efficiently used for xylan biodegradation.

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알로에 성분 Ny932의 혈관생성 촉진작용과 그 작용기전

  • 김규원
    • Proceedings of the Korean Society of Toxicology Conference
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    • 1997.05a
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    • pp.57-60
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    • 1997
  • Aloe의 추출 시료들 중 혈관생성을 촉진하는 물질이 G1M1D1M1 분획에 있음을 chorioallantoic-membrane(CAM) assay, in vitro angiogenesis assay, u-PA, PAI, u-PAR, 및 matrix metalloprotease(MMP)의 유전자 발현조사, gelatin zymogram assay, modified CAM assay를 하여 확인 하였다. 그런 다음 G1M1D1M1 분획을 다시 분리하여 단일 성분인 NY932와 3종의 분획들을(U>3,000, 3,000>U>1,000, U<1,000) 얻었다. 이들 각각에 대한 혈관 생성 촉진 작용을 CAM assay 방법으로 조사한 결과 순수단일 성분인NY932가 혈관 생성 촉진 활성을 가장 높게 나타났다. 즉 NY932의 농도를 100ng, 2, 10, 35, 60 $\mu$g으로 투여하였을 때, 각각 2/18($11\%$), 4/10($49\%$), 16/22($73\%$), 15/17($88\%$), 9/9($100\%$)의 혈관생성 촉진 효과를 보였다. 따라서 순수 단일 성분인 NY932의 혈관 생성 촉진 작용기전을 규명하기 위하여 혈관 생성에 관련된 효소들[matrix metalloprotease(MMP)s, urokinase-plasminogen activator(uPA) 등]과 그 저해제들(TIMPs, PAI)의 유전자 발현을 조사하였으며, wounding migration assay등을 수행하였다.

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