• Title/Summary/Keyword: Immobilized cell

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Cloning and functional expression of a cecropin-A gene from the Japanese oak silkworm, Antheraea yamamai (천잠 cecropin-A 유전자 클로닝 및 재조합 발현)

  • Kim, Seong-Ryul;Choi, Kwang-Ho;Kim, Sung-Wan;Goo, Tae-Won;Hwang, Jae-Sam
    • Journal of Sericultural and Entomological Science
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    • v.52 no.1
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    • pp.45-51
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    • 2014
  • A cecropin-A gene was isolated from the immunized larvae of the Japanese oak silkworm, Antheraea yamamai and designed Ay-CecA. The complete Ay-CecA cDNA consists of 419 nucleotides with 195 bp open reading frame encoding a 64 amino acid precursor that contains a putative 22-residue signal peptide, a 4-residue propetide and a 37-residue mature peptide with a theoretical mass of 4046.81. The deduced amino acid sequence of the peptide evidenced a significant degree of identity (62 ~ 78% identity) with other lepidopteran cecropins. Like many insect cecropin, Ay-CecA also harbored a glycine residue for C-terminal amidation at the C-end, which suggests potential amidation. To understand this peptide better, we successfully expressed bioactive recombinant Ay-CecA in Escherichia coli that are highly sensitive to the mature peptide. For this, we fused mature Ay-CecA gene with insoluble protein ketosteroid isomerase (KSI) gene to avoid the cell death during induction. The fusion KSI-CecA protein was expressed as inclusion body. The expressed fusion protein was purified by Ni-NTA immobilized metal affinity chromatography (IMAC), and cleaved by cyanogen bromide (CNBr) to release recombinant Ay-CecA. The purified recombinant Ay-CecA showed considerably antibacterial activity against Gram-negative bacteria, E. cori ML 35, Klebsiella pneumonia and Pseudomonas aeruginosa. Our results proved that this peptide with a potent antibacterial activity may play a role in the immune response of Japanese oak silkworm.

Preparation and Properties of Coimmobilized Glucose Oxidase-Catalase (Glucose Oxidase와 Catalase의 동시 고정화 제품과 성질)

  • Lee, Suk-Hee;Uhm, Tai-Boong;Cho, Sook-Ja;Byun, Si-Myung
    • Applied Biological Chemistry
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    • v.27 no.3
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    • pp.180-186
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    • 1984
  • For the study of glucose oxidase(GOD) and catalase(CAT) coimmobilization system, the enzymes were obtained from Penicillium spp., PS-8, and the strain itself was used as an immobilizing matrix. To separate glucose oxidase and catalase after the ammonium sulfate fractionation of the culture broth, DEAF-cellulose column was used and its activity yield was 54 and 34%, respectively. Both enzymes were immobilized on the cell matrix, followed crosslinking with 2.5% glutaraldehyde for 12hr. In the determination of efficiencies of GOD and CAT of dual, mixed and soluble enzyme systems, the dual immobilized one w-as superior to those of the soluble or mixed ones. In the comparison of pH profiles, the dual and mixed types showed broader maximum pH ranges than the soluble type. Varying CAT/GOD ratio of the dual system, the higher the ratio showed the broader activity profile. In the comparison of apparent $K_m$ of GOD only and CAT/GOD=10, they were $7.1{\times}10^{-2}$ and $5.1{\times}10^{-2}M$. Their activation energies showed 3.98kcal/mole/deg for GOD only and 2.98kcal/mole/deg for CAT/GOD=10.

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Genomic DNA Chip: Genome-wide profiling in Cancer

  • 이종호
    • Proceedings of the Korean Society for Bioinformatics Conference
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    • 2001.10a
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    • pp.61-86
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    • 2001
  • All cancers are caused by abnormalities in DNA sequence. Throughout life, the DNA in human cells is exposed to mutagens and suffers mistakes in replication, resulting in progressive, subtle changes in the DNA sequence in each cell. Since the development of conventional and molecular cytogenetic methods to the analysis of chromosomal aberrations in cancers, more than 1,800 recurring chromosomal breakpoints have been identified. These breakpoints and regions of nonrandom copy number changes typically point to the location of genes involved in cancer initiation and progression. With the introduction of molecular cytogenetic methodologies based on fluorescence in situ hybridization (FISH), namely, comparative genomic hybridization (CGH) and multicolor FISH (m-FISH) in carcinomas become susceptible to analysis. Conventional CGH has been widely applied for the detection of genomic imbalances in tumor cells, and used normal metaphase chromosomes as targets for the mapping of copy number changes. However, this limits the mapping of such imbalances to the resolution limit of metaphase chromosomes (usually 10 to 20 Mb). Efforts to increase this resolution have led to the "new"concept of genomic DNA chip (1 to 2 Mb), whereby the chromosomal target is replaced with cloned DNA immobilized on such as glass slides. The resulting resolution then depends on the size of the immobilized DNA fragments. We have completed the first draft of its Korean Genome Project. The project proceeded by end sequencing inserts from a library of 96,768 bacterial artificial chromosomes (BACs) containing genomic DNA fragments from Korean ethnicity. The sequenced BAC ends were then compared to the Human Genome Project′s publicly available sequence database and aligned according to known cancer gene sequences. These BAC clones were biotinylated by nick translation, hybridized to cytogenetic preparations of metaphase cells, and detected with fluorescein-conjugated avidin. Only locations of unique or low-copy Portions of the clone are identified, because high-copy interspersed repetitive sequences in the probe were suppressed by the addition of unlabelled Cotl DNA. Banding patterns were produced using DAPI. By this means, every BAC fragment has been matched to its appropriate chromosomal location. We have placed 86 (156 BAC clones) cytogenetically defined landmarks to help with the characterization of known cancer genes. Microarray techniques would be applied in CGH by replacement of metaphase chromosome to arrayed BAC confirming in oncogene and tumor suppressor gene: and an array BAC clones from the collection is used to perform a genome-wide scan for segmental aneuploidy by array-CGH. Therefore, the genomic DNA chip (arrayed BAC) will be undoubtedly provide accurate diagnosis of deletions, duplication, insertions and rearrangements of genomic material related to various human phenotypes, including neoplasias. And our tumor markers based on genetic abnormalities of cancer would be identified and contribute to the screening of the stage of cancers and/or hereditary diseases

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Immobilization of Bacillus sp. Strains, Catalase Producing Bacteria and Their Hydrogen Peroxide Removal Characteristics (카탈라제를 생산하는 고초균 (Bacillus sp.)의 고정화 및 과산화수소 분해 특성)

  • Han, Kyung-Ah;Jang, Yun-Hee;Rhee, Jong-Il
    • KSBB Journal
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    • v.25 no.6
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    • pp.520-526
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    • 2010
  • In this work we have investigated the production of catalase from Bacillus sp. strains, which were screened and identified from soil. These strains were cultivated in shaking flasks with tryptic soy broth (TSB) at $30^{\circ}C$ and 200 rpm. Effects of the temperature and pH on the stability of the native catalase and whole cell viability were studied in the temperature range of $25-60^{\circ}C$ and the pH range of 7-13. Korean natural zeolite was added to culture medium and mixed with microorganisms for 24 hours. The native catalase maintained its activity over $50^{\circ}C$. The enzyme acitiviy of the catalase from Bacillus flexus BKBChE-3 was highest among the Bacillus sp. strains studied. Bacillus flexus BKBChE-3 and immobilized Bacillus cells have survived under extreme conditions of over $50^{\circ}C$ and pH 12. 60 mL of 10.5 mM $H_2O_2$ solution were entirely removed within 1 hour with catalase produced from Bacillus sp. on the flask. When Bacillus cells were immobilized on Korean natural zeolite, colony forming unit of Bacillus flexus BKBChE-3 was increased and high efficiency of hydrogen peroxide removal was observed.

Early bone healing onto implant surface treated by fibronectin/oxysterol for cell adhesion/osteogenic differentiation: in vivo experimental study in dogs

  • Lee, Jung-Seok;Yang, Jin-Hyuk;Hong, Ji-Youn;Jung, Ui-Won;Yang, Hyeong-Cheol;Lee, In-Seop;Choi, Seong-Ho
    • Journal of Periodontal and Implant Science
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    • v.44 no.5
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    • pp.242-250
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    • 2014
  • Purpose: This study aimed to evaluate the effects of fibronectin and oxysterol immobilized on machined-surface dental implants for the enhancement of cell attachment and osteogenic differentiation, on peri-implant bone healing in the early healing phase using an experimental model in dogs. Methods: Five types of dental implants were installed at a healed alveolar ridge in five dogs: a machined-surface implant (MI), apatite-coated MI (AMI), fibronectin-loaded AMI (FAMI), oxysterol-loaded AMI (OAMI), and sand-blasted, large-grit, acid-etched surface implant (SLAI). A randomly selected unilateral ridge was observed for 2 weeks, and the contralateral ridge for a 4-week period. Histologic and histometric analyses were performed for the bone-to-implant contact proportion (BIC) and bone density around the dental implant surface. Results: Different bone healing patterns were observed according to the type of implant surface 2 weeks after installation; newly formed bone continuously lined the entire surfaces in specimens of the FAMI and SLAI groups, whereas bony trabecula from adjacent bone tissue appeared with minimal new bone lining onto the surface in the MI, AMI, and OAMI groups. Histometric results revealed a significant reduction in the BIC in MI, AMI, and OAMI compared to SLAI, but FAMI demonstrated a comparable BIC with SLAI. Although both the BIC and bone density increased from a 2- to 4-week healing period, bone density showed no significant difference among any of the experimental and control groups. Conclusions: A fibronectin-coated implant surface designed for cell adhesion could increase contact osteogenesis in the early bone healing phase, but an oxysterol-coated implant surface designed for osteoinductivity could not modify early bone healing around implants in normal bone physiology.

Current Generation from Microbial Fuel Cell Using Stainless Steel Wire as Anode Electrode (스텐철사를 전극으로 이용하는 미생물연료전지의 전류 발생)

  • Jang, Jae Kyung;Kim, Kyung Min;Byun, SungAh;Ryou, Young Sun;Chang, In Seop;Kang, Young Koo;Kim, Young Hwa
    • Journal of Korean Society of Environmental Engineers
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    • v.36 no.11
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    • pp.753-757
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    • 2014
  • Anode electrode in a microbial fuel cell (MFC) should transfer the receiving electron as well as provide large surface area that can be immobilized microorganisms. Microorganisms' population is one of important factors to improve the current generation and to treat the livestock wastewater by biological treatment. These studies were attempted to investigate if stainless-steel wire skein (SSWS) could be used as anode electrode replacement a graphite felt electrode in microbial fuel cell. For these studies, pretreated livestock wastewater was used diluted to 500 mg/L as COD before use. At this time, the current showed a little difference of about 5% when using each of a SSW and graphite felt (control). There was no significant difference in the current value. The organic removal rate in the microbial fuel cells used graphite felt and SSWS was 82.4% and 88.3%, respectively. The COD removal in the MFC used the SSWS was higher than that of graphite felt. Ammonium nitrogen was showed similar trend in two case all. These results about current generation and organic matter reduction seem possible that SSWS was used to anode electrode. When SSWS is used, the initial investment for system construction is expected to be able to reduce by approximately 1/50.

Characteristics of Heavy Metal Biosorption by Pseudomonas cepacia KH410 (Pseudomonas cepacia KH410의 중금속 흡착특성)

  • 박지원;김영희
    • Korean Journal of Microbiology
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    • v.37 no.3
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    • pp.197-203
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    • 2001
  • An ubiquitous bacterium, Pseudomonar cepacia KH410 was isolated from fresh water plant root and identified. Adsorption of heavy metals of lead, cadmium and copper by this strain was investigated. Optimal conditions foradsorption was 1.0 dry g-biomass, at pH 4.0 and temperature of $40^{\circ}C$. Adsorption equilibrium reached max-imum after 120 min in 1000 mg/l metal solutions. The adsorption capacity (K) of lead was 5.6 times higher thancadmium and 4.0 times higher than that of copper. Adsorption of lead was applicable for Langmuir modelwhereas Freundlich model for cadmium and copper, respectively. Adsorption strength (1/n) of heavy metal ionswere in the order of lead>copper>cadmium. Uptake capacity of lead, cadmium and copper by dried cell was83.2,42.0,65.2 mg/g-biomass, respectively. Effective desorption was induced 0.1 M HCI for lead and 0.1 $HNO_3$ for cadmium and copper. Pretreatment to increase ion strength was the most effective with 0.1 M KOH.Uptake by immobilized cell was 77.8,58.5,71.2 mg/g-biomass for lead, cadmium and copper, respectively. Theimmobilized cell was more effective than ion exchange resin on removal of heavy metals in solution containinglight metals.

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Enhancement of Electrochemical Performance of Cathode by Optimizing Laccase-Carbon Nanotubes Layers for Enzymatic Fuel Cells (Laccase-탄소나노튜브 적층을 통한 효소 연료전지의 cathode 성능 향상)

  • Wang, Xue;Kim, Chang-Joon
    • Korean Chemical Engineering Research
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    • v.60 no.4
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    • pp.550-556
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    • 2022
  • The performance of enzymatic fuel cells that convert chemical energy contained in various organic molecules such as sugar, alcohol, organic acids, and amino acids into electrical energy is greatly affected by the cathode as well as the anode. This study aimed to develop a laccase-based cathode with high performance. An enzyme composite composed of an laccase, redox mediator, and carbon nanotubes was immobilized on the surface of electrode in multiple layers, and the effect of the number of layers and the presence or absence of carbon nanotubes on electrode performance was investigated. As the number of layers of the enzyme-mediator (Lac-(PVI-Os-dCl)) on the electrode surface increased, the amount of reduction current generated at the electrode increased. The enzyme-carbon nanotube-mediator composite electrode (Lac-SWCNTs-(PVI-Os-dCl)) generated a current 1.7 times greater than that of the Lac-(PVI-Os-dCl). It was found that the largest amount of current (10.1±0.1 µA) was generated in the electrode composed of two layers of Lac-(PVI-Os-dCl) and two layers of Lac-SWCNTs-(PVI-Os-dCl) in the evaluation of electrodes with different ratio of Lac-SWCNTs-(PVI-Os-dCl) and Lac-(PVI-Os-dCl). The maximum power density of the cell using the cathode composed of a single layer of Lac-(PVI-Os-dCl) and the cell using the optimized cathode were 0.46±0.05 and 1.23±0.04 µW/cm2, respectively. In this study, it was demonstrated that the performance of cathode and the enzymatic fuel cell using the same can be improved by optimizing the layers of composites composed of laccase, redox mediator, and carbon nanotubes on the electrode surface.

The Effect of Alloderm on Prevention of Adhesions following Tenorrhaphy in the Rabbits (알로덤이 건 봉합술 후 발생되는 유착 방지에 미치는 효과)

  • Choi, Chang Yong;Song, Jin Woo;Kim, Jun Hyuk;Choi, Hwan Jun;Lee, Young Man
    • Archives of Plastic Surgery
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    • v.34 no.6
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    • pp.765-770
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    • 2007
  • Purpose: Peritendinous adhesion is one of the most notorious complication after the flexor tendon injury. In this study, $Alloderm^{(R)}$(LifeCell Corp., Branchburg, N.J.), which is the decellularized human dermal analogue with its intact native basement membrane components, was used for the prevention of peritendinous adhesions following flexor tendon repair. Methods: Thirty New Zealand white male rabbits were divided equally into 3 groups. In all groups, the flexor digitorum profundus of the third finger of the right back foot was cut totally and repaired by modified Kessler suture technique. Following tendon repair, $Alloderm^{(R)}$ was wrapt around the repaired tendon in the first group and sodium hyaluronate gel was sprayed to the operation field in the second group. In the control group, no external material was applied. The right back foot were immobilized for 6 weeks to optimize the formation of adhesion ingrowth. After death, the third finger that repaired tendons and sheaths was removed en bloc. We checked range of motion. and studied histologically for all groups. Results: The experimental groups had better range of motion than the control group. We checked that the range of motion was 73.5 degrees in $Alloderm^{(R)}$ group, 55.9 degrees in the hyaluronic acid group, and 38.3 degrees in the control group. in the histological study, the experimental group had less adhesions compared with the control group. Conclusion: This study concludes that $Alloderm^{(R)}$ can decrease peritendinous adhesions following flexor tendon repairs in rabbits. We think the method could be used in clinical cases.

Evaluating Carriers for Immobilizing Saccharomyces cerevisiae for Ethanol Production in a Continuous Column Reactor

  • Cha, Hye-Geun;Kim, Yi-Ok;Choi, Woon Yong;Kang, Do-Hyung;Lee, Hyeon-Yong;Jung, Kyung-Hwan
    • Mycobiology
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    • v.42 no.3
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    • pp.249-255
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    • 2014
  • We evaluated a more practical and cost-effective immobilization carriers for ethanol production using the yeast Saccharomyces cerevisiae. Three candidate materials-rice hull, rice straw, and sawdust-were tested for their cell-adsorption capacity and operational durability. Derivatizations of rice hull, rice straw, and sawdust with the optimal concentration of 0.5 M of 2-(diethylamino)ethyl chloride hydrochloride (DEAE HCl) resulted in > 95% adsorption of the initial yeast cells at 2 hr for DEAE-rice hull and DEAE-sawdust and in only approximately 80% adsorption for DEAE-rice straw. In addition, DEAE-sawdust was found to be a more practical carrier for immobilizing yeast cells in terms of operational durability in shaking flask cultures with two different speeds of 60 and 150 rpm. Furthermore, the biosorption isotherms of DEAE-rice hull, -rice straw, and -sawdust for yeast cells revealed that the $Q_{max}$ of DEAE-sawdust (82.6 mg/g) was greater than that of DEAE-rice hull and DEAE-rice straw. During the 404-hr of continuous column reactor operation using yeast cells immobilized on DEAE-sawdust, no serious detachment of the yeast cells from the DEAE-sawdust was recorded. Ethanol yield of approximately 3.04 g/L was produced steadily, and glucose was completely converted to ethanol at a yield of 0.375 g-ethanol/g-glucose (73.4% of the theoretical value). Thus, sawdust is a promising practical immobilization carrier for ethanol production, with significance in the production of bioethanol as a biofuel.