• Title/Summary/Keyword: Protein engineering techniques

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Preparation and Characterization of Natural Material Extracted from Germinated Brown Rice

  • Lim, Ki-Taek;Choi, Jeong Moon;Lim, Won-Chul;Kim, Jangho;Cho, Hong-Yon;Chung, Jong Hoon
    • Journal of Biosystems Engineering
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    • v.39 no.3
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    • pp.235-243
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    • 2014
  • Purpose: The aim of this study was to prepare and evaluate a natural material extracted from germinated brown rice (GBR). Herein, we evaluated whether the natural material could positively activate the biological effects seen during bone formation, including enhancement of metabolic activity, osteogenesis, and the expression of vascular endothelial growth factor (VEGF), one of the growth factors in human osteoblast-like cells. Methods: The natural material was created by a hot water extraction process after being soaked for 2~3 days in tap water and dried at $50^{\circ}C$. The material was characterized using field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), and Fourier transformed infrared (FTIR) spectroscopy. The biological behaviors of the material were also investigated; we performed tests to assess cell cytotoxicity, metabolic activity, osteogenic markers related to bone formation, and VEGF. Results: The EDX, XRD, and FTIR results for the natural material indicated the presence of organic compounds. The natural material caused positive increases in cell metabolic activity and mineralized bone formation without cytotoxicity. The protein levels in the extract for the $6.25{\mu}g/mL$, $12.25{\mu}g/mL$, $25{\mu}g/mL$, $50{\mu}g/mL$, and $100{\mu}g/mL$ groups were significantly different from that for the control. Conclusions: The GBR-based natural material was easy to prepare and had characteristics of a potential biomaterial. The biocompatibility of this natural material was evaluated using in vitro techniques; our findings indicate that this novel material is promising for agricultural and biological applications.

Field gradient calculation of HTS double-pancake coils considering the slanted turns and the splice

  • Baek, Geonwoo;Kim, Jinsub;Lee, Woo Seung;Song, Seunghyun;Lee, Onyou;Kang, Hyoungku;Ko, Tae Kuk
    • Progress in Superconductivity and Cryogenics
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    • v.19 no.1
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    • pp.51-55
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    • 2017
  • To obtain Nuclear Magnetic Resonance (NMR) measurement of membrane protein, an NMR magnet is required to generate high intensity, homogeneity, and stability of field. A High-Temperature Superconducting (HTS) magnet is a promising alternative to a conventional Low-Temperature Superconducting (LTS) NMR magnet for high field, current density, and stability margin. Conventionally, an HTS coil has been wound by several winding techniques such as Single-Pancake (SP), Double-Pancake (DP), and layer-wound. The DP winding technique has been frequently used for a large magnet because long HTS wire is generally difficult to manufacture, and maintenance of magnet is convenient. However, magnetic field generated by the slanted turns and the splice leads to field inhomogeneity in Diameter of Spherical Volume (DSV). The field inhomogeneity degrades performance of NMR spectrometer and thus effect of the slanted turns and the splice should be analyzed. In this paper, field gradient of HTS double-pancake coils considering the slanted turns and the splice was calculated using Biot-Savart law and numerical integration. The calculation results showed that magnetic field produced by the slanted turns and the splice caused significant inhomogeneity of field.

Proteomics in Rheumatoid Arthritis Research

  • Park, Yune-Jung;Chung, Min Kyung;Hwang, Daehee;Kim, Wan-Uk
    • IMMUNE NETWORK
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    • v.15 no.4
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    • pp.177-185
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    • 2015
  • Although rheumatoid arthritis (RA) is the most common chronic inflammatory autoimmune disease, diagnosis of RA is currently based on clinical manifestations, and there is no simple, practical assessment tool in the clinical field to assess disease activity and severity. Recently, there has been increasing interest in the discovery of new diagnostic RA biomarkers that can assist in evaluating disease activity, severity, and treatment response. Proteomics, the large-scale study of the proteome, has emerged as a powerful technique for protein identification and characterization. For the past 10 years, proteomic techniques have been applied to different biological samples (synovial tissue/fluid, blood, and urine) from RA patients and experimental animal models. In this review, we summarize the current state of the application of proteomics in RA and its importance in identifying biomarkers and treatment targets.

Solution State Structure of pB1, the Mimotopic Peptide of Apolipoprotein B-100, by NMR

  • Lee, Sung-Ran;Kim, Dae-Sung;Kim, Hyo-Joon;Lee, Yong-Woo;Won, Ho-Shik
    • Bulletin of the Korean Chemical Society
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    • v.25 no.12
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    • pp.1845-1849
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    • 2004
  • Apolipoprotein B-100 (Apo-B100) is a major protein component for low density lipoproteins (LDL). A number of mimetic peptides of Apo-B100 were screened from the phase-displayed random peptide library by utilizing monoclonal antibody (B9). Mimetic peptide for B9 epitope against apo B-100 was CRNVPPIFNDVYWIAF (pB1). From the BLAST search, the mimetic peptide pB1 had 40% homology with apo B-100. As a result of the structural determination of this mimotope using homo/hetero nuclear 2D-NMR techniques and NMR-based distance geometry (DG)/molecular dynamic (MD) computations, DG structure had low penalty value of 0.3-0.6 ${\AA}^2$ and the total RMSD was 0.5-1.5 ${\AA}. Moreover, pB1 structure included a weak $3_{10}$-helix from $Ile^7$,/TEX> to $Trp^{13}$.

Targeted Immunotherapy for Autoimmune Disease

  • Seung Min Jung;Wan-Uk Kim
    • IMMUNE NETWORK
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    • v.22 no.1
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    • pp.9.1-9.23
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    • 2022
  • In the past few decades, biological drugs and small molecule inhibitors targeting inflammatory cytokines, immune cells, and intracellular kinases have become the standard-of-care to treat autoimmune diseases. Inhibition of TNF, IL-6, IL-17, and IL-23 has revolutionized the treatment of autoimmune diseases, such as rheumatoid arthritis, ankylosing spondylitis, and psoriasis. B cell depletion therapy using anti-CD20 mAbs has shown promising results in patients with neuroinflammatory diseases, and inhibition of B cell survival factors is approved for treatment of systemic lupus erythematosus. Targeting co-stimulatory molecules expressed on Ag-presenting cells and T cells is also expected to have therapeutic potential in autoimmune diseases by modulating T cell function. Recently, small molecule kinase inhibitors targeting the JAK family, which is responsible for signal transduction from multiple receptors, have garnered great interest in the field of autoimmune and hematologic diseases. However, there are still unmet medical needs in terms of therapeutic efficacy and safety profiles. Emerging therapies aim to induce immune tolerance without compromising immune function, using advanced molecular engineering techniques.

Surface Modification of Polypropylene Membrane by ${\gamma}$ Irradiation Methods and their Solutes Permeation Behaviors

  • Shim, J. K.;Lee, S. H.;Kwon, O. H.;Lee, Y. M.;Nho, Y. C.
    • Proceedings of the Membrane Society of Korea Conference
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    • 1998.04a
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    • pp.99-101
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    • 1998
  • 1. Introduction : The conventional grafting polymerization technique requires chemically reactive groups on the surface as well as on the polymer chains. For this reason, a series of prefunctionalization steps are necessary for covalent grafting. The surface prefunctionalizational technique for grafting can be used to ionization radiation, UV, plasma, ion beam or chemical initiators. Of these techniques, radiation method is one of the useful methods because of uniform and rapid creation of active radical sites without catalytic contamination in grafted samples. If the diffusion of monomer into polymer is large enough to come to the inside of polymer substrate, a homogeneous and uniform grafting reaction can be carried out throughout the whole polymer substrate. Radiation-induced grafting method may attach specific functional moieties to a polymeric substrate, such as preirradiation and simultaneous irradiation. The former is irradiated at backbone polymer in vacuum or nitrogen gas and air, and then subsequent monomer grafting by trapped or peroxy radicals, while the latter is irradiated at backbone polymer in the presence of the monomer. Therefore, radiation-induced polymerization can be used to modification of the chemical and physical properties of the polymeric materials and has attracted considerable interest because it imparts desirable properties such as blood compatibility. membrane quality, ion excahnge, dyeability, protein adsorption, and immobilization of bioactive materials. Synthesizing biocompatible materials by radiation method such as preirradiation or simultaneous irradiation has often used $\gamma$-rays to graft hydrophilic monomers onto hydrophobic polymer substrates. In this work, in attempt to produce surfaces that show low levels of anti-fouling of bovine serum albumin(BSA) solutions, hydroxyethyl methacrylate(HEMA) was grafted polypropylene membrane surfaces by preirradiation technique. The anti-fouling effect of the polypropylene membrane after grafting was examined by permeation BSA solution.

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Anti-inflammatory Effect of the Processed Gardeniae Fructus in LPS-induced Macrophages (LPS로 유도된 대식세포에서 수치 치자의 항염효과)

  • An, Iseul;Kim, Sang Chan;Byun, Sung Hui;Lee, Jong Rok;Park, Sook Jahr
    • Herbal Formula Science
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    • v.27 no.4
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    • pp.245-255
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    • 2019
  • Objective : Herbal processing is one of the traditional techniques used in Korean medicine to increase the effectiveness of herbs or reduce their toxicity. In this study, Gardeniae Fructus processed with ginger juice and alcohol was prepared to evaluate the anti-inflammatory effect on lipopolysaccharide (LPS)-induced macrophages. Methods : The processing of Gardeniae Fructus was performed by adding 40 % ginger juice or 10% alcohol to the total weight of Gardeniae Fructus and then roasting at 150℃ for 5 minutes. Cell viability was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. To detect nitric oxide (NO) production, culture media were mixed with Griess reagent and measured the absorbance at 540 nm. Prostaglandin E2 (PGE2) and pro-inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA). Western blot was applied to monitor protein expression levels. Results : LPS-induced NO, PGE2 and inflammatory cytokines were decreased by the treatment of normal or processed Gardeniae Fructus ethanol extracts (GFE). Compared to normal GFE, the processed GFE showed a stronger inhibitory effect on the production of NO and PGE2. These inhibitory effect of GFE was due to the suppression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mediated from the inhibition of nuclear factor kappa B (NF-κB). Furthermore, processed GFE showed more suppressive effects on the expression of iNOS, COX-2 and IκBα proteins than normal GFE. Conclusion : From these results, it was concluded that GFE had an improved anti-inflammatory effect compared to normal GFE. These results provide an objective evidences for the use of herbal processing in Korean medicine.

Growth of Endothelial Cells on Microfabricated Silicon Nitride Membranes for an In Vitro Model of the Blood-brain Barrier

  • Harris, Sarina G.;Shuler, Michael L.
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.8 no.4
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    • pp.246-251
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    • 2003
  • The blood-brain barrier (BBB) is composed of the brain capillaries, which are lined by endothelial cells displaying extremely tight intercellular junctions. Several attempts at creating an in vitro model of the BBB have been met with moderate success as brain capillary endothelial cells lose their barrier properties when isolated in cell culture. This may be due to a lack of recreation of the in vivo endothelial cellular environment in these models, including nearly constant contact with astrocyte foot processes. This work is motivated by the hypothesis that growing endothelial cells on one side of an ultra-thin, highly porous membrane and differentiating astrocyte or astrogliomal cells on the opposite side will lead to a higher degree of interaction between the two cell types and therefore to an improved model. Here we describe our initial efforts towards testing this hypothesis including a procedure for membrane fabrication and methods for culturing endothelial cells on these membranes. We have fabricated a 1 $\mu\textrm{m}$ thick, 2.0 $\mu\textrm{m}$ pore size, and 55% porous membrane with a very narrow pore size distribution from low-stress silicon nitride (SiN) utilizing techniques from the microelectronics industry. We have developed a base, acid, autoclave routine that prepares the membranes for cell culture both by cleaning residual fabrication chemicals from the surface and by increasing the hydrophilicity of the membranes (confirmed by contact angle measurements). Gelatin, fibronectin, and a 50/50 mixture of the two proteins were evaluated as potential basement membrane protein treatments prior to membrane cell seeding. All three treatments support adequate attachment and growth on the membranes compared to the control.

Targeting Analysis of Lumenal Proteins of Chloroplast of Wheat using Proteomic Techniques

  • Kamal, Abu Hena Mostafa;Kim, Da-Eun;Oh, Myoung-Won;Chung, Keun-Yook;Cho, Yong-Gu;Kim, Hong-Sig;Song, Beom-Heon;Lee, Chul-Won;Uozumi, Nobuyuki;Choi, Jong-Soon;Cho, Kun;Woo, Sun-Hee
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2010.05a
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    • pp.14-14
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    • 2010
  • Plastid proteomics are essential organelles present in virtually all cells in plants and green algae. Plastids are responsible for the synthesis and storage of key molecules required for the basic architecture and functions of plant cells. The proteome of plastid, and in particular of chloroplast, have received significant amounts of attention in recent years. Various fractionation and mass spectrometry (MS) techniques have been applied to catalogue the chloroplast proteome and its sub-organelles compartments. To better understanding the function of the lumenal sub-organelles within the thylakoid network, we have carried out a systematical analysis and identification of the lumenal proteins in the thylakoid of wheat by using Tricine-SDS-PAGE, and LTQ-ESI-FTICR mass spectrometry followed by SWISS-PROT database searching. We isolation and fractionation these membrane from fully developed wheat leaves using a combination of differential and gradient centrifugation couple to high speed ultra-centrifuge. After collecting all proteins to eliminate possible same proteins, we estimated that there are 407 different proteins including chloroplast, chloroplast stroma, lumenal, and thylakoid membrane proteins excluding 20 proteins, which were identified in nucleus, cytoplasm and mitochondria. A combination of these three programs (PSORT, TargetP, TMHMM, and TOPPRED) was found to provide a useful tool for evaluating chloroplast localization, transit peptide, transmembranes, and also could reveal possible alternative processing sites and dual targeting. Finally, we report also sub-cellular location specific protein interaction network using Cytoscape software, which provides further insight into the biochemical pathways of photosynthesis. The present work helps understanding photosynthesis process in wheat at the molecular level and provides a new overview of the biochemical machinery of the thylakoid in wheat.

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Life Cycle Assessment of the Domestic Dairy Cow System (국내 낙농우(젖소)로부터의 우유생산에 대한 전과정평가)

  • Park, Yoo Sung;Lee, Kun Mo;Yang, Seung Hak
    • Journal of Korean Society of Environmental Engineers
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    • v.37 no.1
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    • pp.52-59
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    • 2015
  • Recently the dairy cow industry have faced environmental issues such as eutrophication, global warming, etc. An LCA was used to quantify the environmental impact of a dairy cow system and to identify key issues contributing to the impact. The system boundary crop cultivation for feeding dairy cow, feed production, rearing and manure management (cradle-to-gate). The functional unit was 1 kg of milk (fat protein corrected milk, FPCM) produced. Rearing and cultivation of feed crops stages in system boundary to the environmental impact of the domestic dairy cow system were dominant issues. Techniques such as suppression of enteric fermentation, improvement of the energy efficiency of farm equipment and apparatuses, management of leachate generated during the crop cultivation, and development of controling the loss of fertilizer during crop production would be necessary for the improvement of the environmental key issues of the dairy cow system.