• Title/Summary/Keyword: Functional characterization

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Characterization of a Glutamate Decarboxylase (GAD) from Enterococcus avium M5 Isolated from Jeotgal, a Korean Fermented Seafood

  • Lee, Kang Wook;Shim, Jae Min;Yao, Zhuang;Kim, Jeong A;Kim, Hyun-Jin;Kim, Jeong Hwan
    • Journal of Microbiology and Biotechnology
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    • v.27 no.7
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    • pp.1216-1222
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    • 2017
  • To develop starters for the production of functional foods or materials, lactic acid bacteria producing ${\gamma}-aminobutyric$ acid (GABA) were screened from jeotgals, Korean fermented seafoods. One isolate producing a high amount of GABA from monosodium $\text\tiny{L}$-glutamate (MSG) was identified as Enterococcus avium by 16S rRNA gene sequencing. E. avium M5 produced $18.47{\pm}1.26mg/ml$ GABA when incubated for 48 h at $37^{\circ}C$ in MRS broth with MSG (3% (w/v)). A gadB gene encoding glutamate decarboxylase (GAD) was cloned and overexpressed in E. coli BL21 (DE3) using the pET26b (+) expression vector. Recombinant GAD was purified through a Ni-NTA column and the size was estimated to be 53 kDa by SDS-PAGE. Maximum GAD activity was observed at pH 4.5 and $55^{\circ}C$and the activity was dependent on pyridoxal 5'-phosphate. The $K_m$ and $V_{max}$ values of GAD were $3.26{\pm}0.21mM$ and $0.0120{\pm}0.0001mM/min$, respectively, when MSG was used as a substrate. Enterococcus avium M5 secretes a lot of GABA when grown on MRS with MSG, and the strain is useful for the production of fermented foods containing a high amount of GABA.

Functional Characterization of the C-Terminus of YhaV in the Escherichia coli PrlF-YhaV Toxin-Antitoxin System

  • Choi, Wonho;Yoon, Min-Ho;Park, Jung-Ho
    • Journal of Microbiology and Biotechnology
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    • v.28 no.6
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    • pp.987-996
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    • 2018
  • Bacterial programmed cell death is regulated by the toxin-antitoxin (TA) system. YhaV (toxin) and Pr1F (antitoxin) have been recently identified as a type II TA system in Escherichia coli. YhaV homologs have conserved active residues within the C-terminus, and to characterize the function of this region, we purified native YhaV protein (without denaturing) and constructed YhaV proteins of varying lengths. Here, we report a new low-temperature method of purifying native YhaV, which is notable given the existing challenges of purifying this highly toxic protein. The secondary structures and thermostability of the purified native protein were characterized and no significant structural destruction was observed, suggesting that the observed inhibition of cell growth in vivo was not the result of structural protein damage. However, it has been reported that excessive levels of protein expression may result in protein misfolding and changes in cell growth and mRNA stability. To exclude this possibility, we used an [$^{35}S$]-methionine prokaryotic cell-free protein synthesis system in vitro in the presence of purified YhaV, and two C-terminal truncated forms of this protein (YhaV-L and YhaV-S). Our results suggest that the YhaV C-terminal region is essential for mRNA interferase activity, and the W143 or H154 residues may play an analogous role to Y87 of RelE.

Analysis of Physicochemical Properties of Nurungji Added with Various Materials (다양한 소재가 첨가된 누룽지의 이화학 특성 분석)

  • Kim, Myong-Ki;Cho, Seok-Cheol
    • Journal of Convergence for Information Technology
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    • v.10 no.2
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    • pp.102-108
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    • 2020
  • Increasing the rice consumption, seven kinds of Nurungji which served as a home meal replacer and snack were developed with some functional materials were added, and their physiochemical characteristics were analyzed. The water binding capacity was the highest in Nurungji with black sesame seed (264.13%), and the hardness was the lowest as 0.36 N in Nurungji with Tenebrio molitor. The reducing sugar content was the highest in Nurungji with sweet pumpkin (1.47%) when being soaked in water for 30 minutes. As a result of measuring 2,2-diphenyl-1-picryl-hydrazyl free radical scavenging activity of the tested Nurungji preparations, Nurungji with sweet pumpkin and beet showed the high scavenging activity against free radicals. As a result of sensory evaluation, overall acceptability was the highest in Nurungji with black sesame seeds (4.3). Physicochemical characterization and sensory evaluation of Nurongji added with various materials confirmed that they could be applied to various food products.

Physicochemical Characterization of Extrudate Solid Formulation of Angelica gigas Nakai Prepared by Hot Melt Extrusion Process

  • Azad, Md Obyedul Kalam;Cho, Hyun Jong;Koo, Ja Seong;Park, Cheol Ho;Kang, Wie Soo
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2018.04a
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    • pp.72-72
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    • 2018
  • The root of Angelica gigas Nakai (AGN) is used as a traditional herbal medicine in Korea for the treatment of many diseases. However, a major challenge associated with the usage of the active compounds from AGN is their poor water solubility. Therefore, this work aimed to enhance the solubility of active compounds by a chemical (viz. surfactant) and physical (hot melt extrusion) crosslinking method (CPC). Infrared Fourier transform spectroscopy (FT-IR) revealed multiple peaks in extrudate solids representing new functional groups including carboxylic acid, alkynes and benzene derivatives. Differential scanning calorimetry (DSC) analysis of the extrudate showed lower glass transition temperature (Tg) and lower enthalpy (${\Delta}H$) (Tg: $43^{\circ}C$; ${\Delta}H$: <6 (J/g)) compared to the non-extrudate (Tg $68.5^{\circ}C$; ${\Delta}H$: 123.2) formulations. X-ray powder diffraction (XRD) analysis revealed amorphization of crystal materials in extrudate solid. In addition, nanonization, enhanced solubility and higher extraction of phenolic compounds were achieved in the extrudate solid. Among the different extrudates, acetic acid- and Span 80-mediated formulations showed superior extractions. We conclude that the CPC method successfully enhanced the production of amorphous nano dispersions from extrudate solid formulations.

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Influence of the Pore Properties on Carbon Dioxide Adsorption of PAN-based Activated Carbon Nanofibers (폴리아크릴로니트릴계 활성나노탄소섬유의 기공특성이 이산화탄소 흡착에 미치는 영향)

  • Lee, Dayoung;Cho, Seho;Kim, Yesol;Lee, Young-Seak
    • Polymer(Korea)
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    • v.37 no.5
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    • pp.592-599
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    • 2013
  • In this study, polyacrylonitrile (PAN)-based porous carbon nanofibers were prepared from PAN polymer solution by electrospinning and KOH activation with various concentrations, and the characterization of pore structures and carbon dioxide adsorption was investigated. Manufactured PAN-based activated carbon nanofibers tend to decrease diameter and increase surface oxygen functional groups depending on the increasing concentration of KOH solution. In addition, according to the results of nitrogen adsorption for pore properties analysis, it indicated increase of the specific surface area in conformity with increasing concentration of KOH solution. Micropore volume of treated activated carbon nanofibers (ANCF) by 4 M KOH was the largest compared with other samples and mesopore volume of treated ANCF by 8 M KOH was the largest volume, respectively. The concentration of KOH effects textural and surface properties, as represented by BET and XPS, which enhance carbon dioxide adsorption capacity at 0 and $25^{\circ}C$.

Structural and Functional Characterization of CRAMP-18 Derived from a Cathelicidin-Related Antimicrobial Peptide CRAMP

  • Park, Kyong-Soo;Shin, Song-Yub;Hahm, Kyung-Soo;Kim, Yang-Mee
    • Bulletin of the Korean Chemical Society
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    • v.24 no.10
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    • pp.1478-1484
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    • 2003
  • CRAMP was identified from a cDNA clone derived from a mouse femoral marrow cells as a member of cathelicidin-derived antimicrobial peptide. Tertiary structure of CRAMP in TFE/$H_2O$ (1 : 1, v/v) solution has been determined by NMR spectroscopy previously and consists of two amphipathic $\alpha-helices$ from Leu4 to Lys10 and from Gly16 to Leu33. These two helices are connected by a flexible region from Gly11 to Gly16. Analysis of series of fragments composed of various portion of CRAMP revealed that an 18-residue fragment with the sequence from Gly16 to Leu33 (CRAMP-18) was found to retain antibacterial activity without cytotoxicity. The effects of two Phe residues at positions 14 and 15 of CRAMP-18 on structure, antibacterial activity, and interaction with lipid membranes were investigated by $Phe^{14,15}$ ${\rightarrow}$ Ala substitution (CRAMP-18-A) in the present study. Substitution of Phe with Ala in CRAMP-18 caused a significant reduction on antibacterial and membrane-disrupting activities. Tertiary structures of CRAMP-18 in 50% TFE/$H_2O$ (1 : 1, v : v) solution shows amphipathic ${\alpha}$-helix, from $Glu^2{\;}to{\;}Leu^{18}$, while CRAMP-18-A has relatively short amphipathic ${\alpha}$-helix from $Leu^4{\;}to{\;}Ala^{15}$. These results suggest that the hydrophobic property of $Phe^{14}{\;}and{\;}Phe^15$ in CRAMP-18 is essential for its antibacterial activity, ${\alpha}$-helical structure, and interactions with phospholipid membranes.

Functional Characterization and Regional Expression of Dopamine Transporter (도파민 수송체의 기능적 특성 및 발현에 관한 연구)

  • 이상훈;이송득;성기욱;이동섭;이용성;고재경
    • YAKHAK HOEJI
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    • v.39 no.2
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    • pp.161-168
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    • 1995
  • Brain dopamine systems play a central role in the control of movement, hormone release, and many complex behavior. The action of dopamine at its synapse is terminated predominately by high affinity reuptake into presynaptic terminals by dopamine transporter (DAT). The dopamine transporter(DAT) is membrane protein localized to dopamine-containing nerve terminals and closely related with cocaine abuse, Parkinsonism, and schizophrenia. In present study, the recombinant plasmid pRc/CMV-DAT, constructed by subcloning of a cDNA encoding a bovine DAT into eukaryotic expression vector pRc/CMV, was stably transfected into CV-1 cells(monkey kidney cell line). The DAT activities in the cell lines selected by Geneticin$^{R}$ were determined by measuring the uptake of $[^3H]$-dopamine. The transfected cell lines showed 30-50 fold higher activities than untransfected CV-1 cell line, and this result implies that DAT is well expressed and localized in transfected cells. The transfected cells accumulated $[^3H]$-dopamine in a dose-dependent manner with a $K_{m}$ of 991.6nM. Even though high doses of norepinephrine, epinephrine, serotonin, and choline neurotransmitters inhibited the uptake of $[^3H]$-dopamine, DAT in transfected cell line was proven to be much more specific to dopamine. The psychotropic drugs such as GBR12909, CFT, normifensine, clomipramine, desipramine, and imipramine inhibited significantly the dopamine uptake in tissue culture cells stably transfected with DAT cDNA. Radioactive in situ hybridization was done to map the cellular localization of DAT mRNA-containing cells in the adult rat central nervous system. The strong hybridization signals were detected only in the substantia nigra pars compacta and ventral tegmental area. The restricted anatomical localization of DAT mRNA-containing cells confirms the DAT as a presynaptic marker of dopamine-containing cells in the rat brain.

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Sonocatalytic Degradation of Rhodamine B in the Presence of TiO2 Nanoparticles by Loading WO3

  • Meng, Ze-Da;Sarkar, Sourav;Zhu, Lei;Ullah, Kefayat;Ye, Shu;Oh, Won-Chun
    • Korean Journal of Materials Research
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    • v.24 no.1
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    • pp.6-12
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    • 2014
  • In the present work, $WO_3$ and $WO_3-TiO_2$ were prepared by the chemical deposition method. Structural variations, surface state and elemental compositions were investigated for preparation of $WO_3-TiO_2$ sonocatalyst. X-ray diffraction (XRD), scanning electron microscopy (SEM), energy dispersive X-ray (EDX) and transmission electron microscopy (TEM) were employed for characterization of these new photocatalysts. A rhodamine B (Rh.B) solution under ultrasonic irradiation was used to determine the catalytic activity. Excellent catalytic degradation of an Rh.B solution was observed using the $WO_3-TiO_2$ composites under ultrasonic irradiation. Sonocatalytic degradation is a novel technology of treating wastewater. During the ultrasonic treatment of aqueous solutions sonoluminescence, cavitaties and "hot spot" occurred, leading to the dissociation of water molecules. In case of a $WO_3$ coupled system, a semiconductor coupled with two components has a beneficial role in improving charge separation and enhancing $TiO_2$ response to ultrasonic radiations. In case of the addition of $WO_3$ as new matter, the excited electrons from the $WO_3$ particles are quickly transferred to $TiO_2$ particle, as the conduction band of $WO_3$ is 0.74 eV which is -0.5 eV more than that of $TiO_2$. This transfer of charge should enhance the oxidation of the adsorbed organic substrate. The result shows that the photocatalytic performance of $TiO_2$ nanoparticles was improved by loading $WO_3$.

Improvement of Wettability and Removal of Skin Layer on Ar-Plasma-Treated Polypropylene Blend Surface (폴리프로필렌 복합소재의 아르곤 플라즈마 처리로 표면층 제거와 젖음성 향상)

  • Weon, Jong-Il;Lee, Sun-Yong
    • Polymer(Korea)
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    • v.36 no.4
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    • pp.461-469
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    • 2012
  • The surface modification and characterization of Ar-plasma treated polypropylene (PP) blend are investigated using x-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and contact angle measurement. An increase in Ar-plasma treatment time leads to an increase in wettability, oxygen containing polar functional groups, the amount of talc, and surface roughness on the PP blend surface. A careful observation using SEM indicates that there exists a skin layer consisting of only PP component. The difference in viscosity between PP and rubber particles facilities the formation of skin layer. However, it is found that an increase in Ar-plasma treatment time helps to decrease the thickness of skin layer. Additional methodologies for the elimination of skin layer during injection molding are also discussed. The surface modification and morphological alteration induced by Ar-plasma treatment provides a hydrophilic state, followed by the improvement in wettability, on the PP blend surface.

Quantitative Evaluation of Scratch Behavior for Polymeric Materials (고분자 소재의 스크래치 거동의 정량적 평가)

  • Baek, Ki-Wan;Lee, Sung-Goo;Lee, Jae-Heung;Choi, Kil-Yeong;Weon, Jong-Il
    • Polymer(Korea)
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    • v.33 no.4
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    • pp.273-283
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    • 2009
  • Recent research issues on the scratch behaviors of polymeric materials has been investigated. In this study, the scratch characterization of polymeric materials with respect of experimental parameters, such as nature of the material, temperature, applied load, test speed, surface treatment, scratch number of times, polymer structure/functional groups, degree of cross-linking, and crystallinity, are reviewed. In addition, the testing standards and methodologies which could quantify the scratch behaviors are introduced and the current international standards are compared and summarized. The latest technical approaches for evaluating the scratch behaviors and improving the scratch resistance of polymers are also discussed.