• Title/Summary/Keyword: 23 kDa protein

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Cross-reacting and specific antigenic components in cystic fluid from metacestodes of Echinococcus grannlosus and Taenia solium (포충 및 유구낭미충 낭액에 있어서 공통항원 및 특리이원 분획)

  • Yoon Kong;Shin-Yong Kang;Seung-Yull Cho;Duk-Young Min
    • Parasites, Hosts and Diseases
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    • v.27 no.2
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    • pp.131-140
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    • 1989
  • Sera from confirmed patients of 5 hydatidosis, 67 neurocysticercosis and 89 other parasitic diseases were tested for specific antibody (IgG) levels by ELISA to cystic quid antigens from metacestodes of Echinococcus granuzosus (HF) and Taenia sodium (CF). All hydatidosis sera reacted positively to both HF and CF while neuro- cysticercosis sera did in 49.3% to HF and 85.1% to CF, The frequencies of cross- reactions were lower in other parasitic diseases to both antigens, By SDS-PAGE, protein bands of 64, 35, 22 and 7 kilodaltons (kDa) were found common in HF and CF. SDS-PAGE/immunoblot exhibited that hl·datidosis sera reacted crossly to CF at 135, 110, 100, 86, 64, 45, 39, 35 and 24 kDa bands while neurocysticercosis sera did to HF at 135, 100, 86, 64, 52, 39, 35, 29 and 24 kDa bands. These results indicated that protein bands of 135, 100, 86, 64, 39, 35 and 24 kDa were major common components in HF and CF. Protein bands of 7 kDa in HF and 15, 10 and 7 kDa in CF did not react crossly and were specific components in respective antigens.

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Mapping of Human Cytomegalovirus IE1 Responsive Elements in the c-jun Promoter (사람세포거대바이러스 (Human Cytomegalovirus)의 극초기항원-1 (Immediate Early-1, IE-1)에 반응하는 c-jun Promoter의 유전자 지도 분석)

  • Park, Chung-Gyu;Han, Tae-Hee;Kim, Dae-Joong;Kim, Jin-Hee;Hwang, Eung-Soo;Choi, Sung-Bae;Cha, Chang-Yong
    • The Journal of Korean Society of Virology
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    • v.28 no.3
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    • pp.267-274
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    • 1998
  • Human cytomegalovirus (HCMV) has the ability to activate the expression of many viral and cellular genes. Among various viral proteins, the immediate early proteins (IE1-72kDa, IE2-86kDa) have been known to be potent transactivators. The product of c-jun proto-oncogene is important in cell activation and differentiation. Here, we tried to find out if the IE could activate the c-jun promoter and also tried to identify the responsible sequence elements in the c-jun activation by IE1-72kDa. We found HCMV IE expression transactivated the c-jun promoter in human embryonal lung fibroblasts (HEL). The activation fold by IE1-72kDa, IE2-86kDa and IE2-55kDa was 23, 35, and 5, respectively. When the expression of each IE was combined, it showed synergism. Expression of (IE1-72kDa + IE2-86kDa) and (IE1-72kDa + IE2-86kDa + IE2-55kDa) resulted in 131 and 162 fold increase, respectively. The c-jun promoter region between -117 and -59 contains binding sites for the transcription factors Spl, CAAT, AP-l like (ATF/CREB), and MEF2. Transient expression assays were performed using various reporter plasmids containing the c-jun promoter-regulatory region linked to the luciferase gene and a plasmid expressing HCMV IE1 gene. Deletional and point mutational analysis showed that the sequence between -225 to -160 and the CTF binding site were involved in the up-regulation of c-jun promoter.

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Isolation and Purification of Protein-bound Polysaccharides from Mycelia of Flammulina velutipes Grown on Sawdust Medium (팽나무버섯 수확후의 톱밥배지로부터 단백다당류의 분리 및 정제)

  • Ha, Hyo-Cheol;Park, Shin;Park, Kyung-Sook;Lee, Shun-Woo;Jung, In-Chang;Kim, Seon-Hee;Kwon, Yong-Il;Lee, Jae-Sung
    • KSBB Journal
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    • v.10 no.5
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    • pp.589-597
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    • 1995
  • Protein-bound polysaccahrides(PBP) were isolated, purified, and characterized from the sawdust media after harvesting the fruit body of Flammulina velutipes. The yield of the crude PBP(Fr.CA) extracted from the sawdust media, was 0.367% relative to the original sawdust media. The total sugar and protein contents of Fr.CA were 19.8% and 23.8% respectively. Using the membrane filtration, the fraction of which the molecular weight is over 300 kDa(Fr.A) was isolated from the Fr.CA and the yield was 44.6% relative to the Fr.CA. This result indicates that high molecular PBP is the dominant components of the Fr.CA. The Fr.A was separated into three fractions (Fr.A-1, Fr.A-2 and Fr.A-3) whose yields are 5.8%, 8.5% and 13.2% respectively. These fractions were further purified using gel filtration, obtaining a single peak in each fraction that considered as pure PBP Among them, the yield of Fr.A-1-${\alpha}$ was 20.9% relative to the Fr.A-1, and the molecular weight was 800 kDa. Monosaccharide components such as glucose, galactose, mannose, xylose and fucose could be detected all fractions by a HPLC analysis. Especially in the Fr.A-1-${\alpha}$ fraction, the content of glucose and galactose appeared to be high.

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Serological characteristics and antigenicities of Vibrio harveyi isolated from marine cultured fish (해산어류에서 분리된 Vibrio harveyi의 혈청학적 특성과 항원성)

  • Oh, Yun-Kyeong;Kim, Myoung-Sug;Park, Myoung-Ae;Kim, Jin-Woo;Cho, Ji-Young;Jeong, Hyun-Do
    • Journal of fish pathology
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    • v.22 no.1
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    • pp.23-33
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    • 2009
  • Vibrio harveyi was a significant pathogenic agent and cause a high mortality of cultured fish and shrimp in the aquaculture industry. In this study, we have investigated biochemical, physiological, serological and immunological characteristics of V. harveyi isolated from marine cultured fish. The phenotypes of V. harveyi were differentiated with their own biochemical characteristics and colors of colony on the TCBS agar. V. harveyi were classified into more than four serogroups by agglutination test. Most isolates were classified into a group A which is categorized with the same band pattern generated by western blotting. Group A was characterized by a major protein, which is ranged from 26 and 34 kDa in size, and has a virulence to oliver flounder more than reference strain KCCM40866. Oliver flounders vaccined with FKC of V. harveyi C05011 were highly resistant to infection by other strains of group A.

Decolorization of Acid Green 25 by Surface Display of CotA laccase on Bacillus subtilis Spores

  • Park, Jong-Hwa;Kim, Wooil;Lee, Yong-Suk;Kim, June-Hyung
    • Journal of Microbiology and Biotechnology
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    • v.29 no.9
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    • pp.1383-1390
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    • 2019
  • In this study, we expressed cotA laccase from Bacillus subtilis on the surface of B. subtilis spores for efficient decolorization of synthetic dyes. The cotE, cotG, and cotY genes were used as anchoring motifs for efficient spore surface display of cotA laccase. Moreover, a $His_6$ tag was inserted at the C-terminal end of cotA for the immunological detection of the expressed fusion protein. Appropriate expression of the CotE-CotA (74 kDa), CotG-CotA (76 kDa), and CotY-CotA (73 kDa) fusion proteins was confirmed by western blot. We verified the surface expression of each fusion protein on B. subtilis spore by flow cytometry. The decoloration rates of Acid Green 25 (anthraquinone dye) for the recombinant DB104 (pSDJH-EA), DB104 (pSDJH-GA), DB104 (pSDJH-YA), and the control DB104 spores were 48.75%, 16.12%, 21.10%, and 9.96%, respectively. DB104 (pSDJH-EA) showed the highest decolorization of Acid Green 25 and was subsequently tested on other synthetic dyes with different structures. The decolorization rates of the DB104 (pSDJH-EA) spore for Acid Red 18 (azo dye) and indigo carmine (indigo dye) were 18.58% and 43.20%, respectively. The optimum temperature for the decolorization of Acid Green 25 by the DB104 (pSDJH-EA) spore was found to be $50^{\circ}C$. Upon treatment with known laccase inhibitors, including EDTA, SDS, and $NaN_3$, the decolorization rate of Acid Green 25 by the DB104 (pSDJH-EA) spore decreased by 23%, 80%, and 36%, respectively.

Partial Purification of Lectin from Mycoparasitic Species of Trichoderma

  • Singh, Tanuja;Saikia, Ratul;Arora, Dilip K.
    • The Plant Pathology Journal
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    • v.21 no.4
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    • pp.301-309
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    • 2005
  • Trichoderma species/isolates exhibited varied degree of agglutination on sclerotial (Sc) and hyphal (Hy) surface of Macrophomina phaseolina. The agglutination efficiencies on Sc and Hy ranged from $11\;to\;57\%$. Isolates of T. harzianum (Th) and T. viride (Tv) showed greater agglutination on Sc ($23-57\%$) and Hy ($16-47\%$). Different enzymes (trypsin, pepsin, proteinase k, a-chymotrypsin, lyticase and glucosidase) and inhibitors (tunicamycin, cycloheximide, brefeldin A, sodium azide, dithiothreitol and SDS) reduced the agglutination potential of conidia of Th-23/98 and Tv-25/98; however, the extent of response varied greatly in different treatments. Different fractions of Th-23/98 and Tv-25/98 exhibited haemagglutinating reaction with human blood group A, B, AB and O. Haemagglutinating activity was inhibited by different sugars and glycoproteins tested. Crude haemagglutinating protein from outer cell wall protein fraction of Th-23/98 and Tv-25/98 were eluted on Sephadex G-100 column. Initially Th-23/98 and Tv-25/98 exhibited two peaks showing no agglutination activity; however, lectin activity was detected in the third peak. Similar to crude lectin, the purified lectin also exhibited haemagglutinating activity with different erythrocyte source. SDS-PAGE analysis of partially purified lectin revealed single band with an estimated molecular mass of 55 and 52 kDa in Th-23/98 and Tv-25/98, respectively. Trypsin, chymotrypsin and b-1,3-glucanase totally inhibited lectin activity. Similarly, various pH also affected the haemagglutinating activity of Th-23/98 and Tv-25/98. From the present observations, it can be concluded that the recognition/attachment of mycoparasite (T. harzianum and T. viride) to the host surface (M. phaseolina) may be most likely due to lectin-carbohydrate interaction.

Analysis of Acid Stress Response in Streptococcus mutans KCTC 3065 (산에 대한 Streptococcus mutans KCTC 3065의 스트레스 반응에 관한 연구)

  • Kang, Kyung-Hee;Kim, Ji-Young
    • Journal of dental hygiene science
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    • v.7 no.1
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    • pp.21-24
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    • 2007
  • Dental caries is initiated by the acid accumulated in dental plaque. Streptococcus mutans, one of a major causal agents of dental caries, is component of the dental plaque and produces various organic acids such as lactic acid as the end-product of glycolysis. As a consequence, we investigated the acid stress response of S. mutans KCTC 3065 in this study. The addition of lactic acid to the growth media had a concentration-dependent effect on the growth of S. mutans. S. mutans exhibited higher maximum culture OD compared with the more acidic growth pH values. At treatment of centration of 20 mmol/L lactic acid in the mid-log phage, cell growth was reduced to 40% relative to the control. The following results were obtained with the treatment of cells with a concentration of 20 mmol/L lactic acid in the mid-log phage for 2hrs: Analysis of fatty acids extracted from cells showed that growth at a concentration of 20 mmol/L lactic acid resulted in changes in $C_{14:0}$, $C_{16:0}$, $C_{18:0}$ and $C_{18:1}$ fatty acids. Protein profiles investigated by SDS-PAGE showed that approximately 70, 60, 45, 40 and 23 kDa proteins were highly expressed in S. mutans KCTC 3065.

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Clonorchis sinensis: Analysis Characterization of Somatic and Metabolic Antigen (II) Profile of the Worm, Excretory-secretory and Billis Antigen in C. sinensis Infected Rabbit (간흡충 : 충체 및 대사성 항원의 특성분석 (II) 간흡충 감염 가토에서 간흡충, 분비배설액 및 담즙 항원의 분획 양상)

  • Yong-Suk Ryang;Yoon-Kyung Cho;Ji-Sook Lee
    • Biomedical Science Letters
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    • v.3 no.2
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    • pp.89-94
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    • 1997
  • The authors characterized the proteins of the crude antigen obtained from Clonorchis sinensis worm and excretory-secretory and billis from rabbits, experimentally infected for 3 months. Protein composition was observed after adding a cysteine proteinase inhibitor E-64 and a serine proteinase inhibitor PMSF, respectively. SDS-PAGE of the crude antigen from C. sinensis recovered from the infected rabbits, the crude antigen from the adult worm excretory-secretory, and the crude antigen from billis of the rabbits resolved 26, 27 and 19 profiles between 200-9 kDa, respectively. When E-64 supplemented 29, and 22 bands, respectively. More study should be carried out in the future on the immunological characteristics and the monoclonal antibody of the each antigen.

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Overexpression and Characterization of Lycopene Cyclase (CrtY) from Marine Bacterium Paracoccus haeundaensis

  • Jeong, Tae Hyug;Ji, Keunho;Kim, Young Tae
    • Journal of Microbiology and Biotechnology
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    • v.23 no.2
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    • pp.144-148
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    • 2013
  • Lycopene cyclase converts lycopene to ${\beta}$-carotene by catalyzing the formation of two beta-rings at each end of the linear carotene structure. This reaction takes place as a two-step reaction in which both sides of of the lycopene molecule are cyclized into ${\beta}$-carotene rings via the monocyclic ${\gamma}$-carotene as an intermediate. The crtY gene coding for lycopene cyclase from Paracoccus haeundaensis consists of 1,158 base pairs encoding 386 amino acids residues. An expression plasmid containing the crtY gene (pET44a-CrtY) was constructed and expressed in Escherichia coli, and produced a recombinant protein of approximately 43 kDa, corresponding to the molecular mass of lycopene cyclase. The expressed protein was purified to homogeneity by His-tag affinity chromatography and showed enzymatic activity corresponding to lycopene cyclase. We also determined the lycopene substrate specificity and NADPH cofactor requirements of the purified protein. The $K_m$ values for lycopene and NADPH were 3.5 ${\mu}M$ and 2 mM, respectively. The results obtained from this study will provide a wider base of knowledge on the enzyme characterization of lycopene cyclase at the molecular level.

Purification and Characterization of an Alkaliphilic Alginate Lyase AlgMytC from Saccharophagus sp. Myt-1

  • Sakatoku, Akihiro;Tanaka, Daisuke;Nakamura, Shogo
    • Journal of Microbiology and Biotechnology
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    • v.23 no.6
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    • pp.872-877
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    • 2013
  • In a previous study, we isolated and reported a second species of the Saccharophagus genus, Saccharophagus sp. strain Myt-1. In the present study, an alginate lyase gene (algMytC) from the genomic DNA of Myt-1 was cloned and characterized. The DNA sequence fragment obtained contained an open reading frame of 1,032 bp that encoded a protein of 343 amino acids with an estimated molecular mass of 37.6 kDa and a pI of 6.60. The deduced protein, AlgMytC, had the conserved amino acid sequences (RTELREM, QIH, YFKAGVYNQ) of the polysaccharide lyase family 7. A BLAST homology search indicated that AlgMytC shared an amino acid sequence identity of 95.9% with alg7A of S. degradans 2-40. The cloned and purified AlgMytC protein showed optimal activity at $40^{\circ}C$, and retained more than 90% of its total activity even after treatment at $25^{\circ}C$ for 24 h. AlgMytC was very alkaliphilic with an optimal pH of 9.0, and more than 90% of its activity was retained in the pH range 8.5-10.0. Moreover, AlgMytC was stable over a wide pH range. The activity of AlgMytC was also stable in the presence of various detergents.