• Title/Summary/Keyword: In cell NMR

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Characterization of the Effects of Silver Nanoparticles on Liver Cell Using HR-MAS NMR Spectroscopy

  • Kim, Si-Won;Kim, So-Sun;Lee, Sang-Mi;Kwon, Bo-Bae;Choi, Jin-Hee;Hyun, Jin-Won;Kim, Suhk-Mann
    • Bulletin of the Korean Chemical Society
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    • v.32 no.6
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    • pp.2021-2026
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    • 2011
  • AgNPs (silver nanoparticles) has been widely used for the commercial products, which have antimicrobial agent, medical devices, food industry and cosmetics. Despite, AgNPs have been reported as toxic to the mammalian cell, lung, liver, brain and other organs and many researchers have investigated the toxicity of AgNPs. In this study, we investigated toxicity of the AgNPs to the liver cell using metabolomics based on HRMAS NMR (High Resolution Magic Angle Spinning Nuclear Magnetic Resonance) technics, which could apply to the intact tissues or cells, to avoid the sample destruction. Target profiling and multivariative statistical analysis were performed to analyze the 1D $^1H$ spectrum. The results show that the concentrations of many metabolites were affected by the AgNPs in the liver cell. The concentrations of glutathione (GSH), lactate, taurine, and glycine were decreased and most of amino acids, choline analogues, and pyruvate were increased by the AgNPs. Moreover, the levels of the metabolites were recovered upto similar level of metabolites in the normal cell by the pre-treatment of NAC, external antioxidant. The results suggest that the depletion of the GSH by the AgNPs might induce the conversion of lactate and taurine to the pyruvate.

Cadmium Detoxification Mechanism in Klebsiella aerogenes ATCC 10031 (Klebsiella aerogenes ATCC 10031의 카드뮴 해독기작)

  • 이기성;유순애;곽인영;박영식;최영길
    • Korean Journal of Microbiology
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    • v.28 no.2
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    • pp.134-144
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    • 1990
  • In order to examine that what kind of system correlated with cadmium detoxification mechanism in Klebsiella aerogenes ATCC 10031, we tried to investigate the effect of phosphate upon the detoxification and also elucidate whether the cadmium phosphate and/or polymeric Cd-Pi complex is formed actually in cell or not. As the results, it was shown that growing pattern had long lag adaptive phase of 12 hr to 24 hr, at the concentrations of 0.02 mM and 0.08 mM cadmium, respectively. Cadmium was accumulated more highly in the fraction of cell wall and membrane than in those of cytoplasm. In case of phosphate starving cells added cadmium, inorganic polyphosphate system was primarily correlated with Cd-detoxification during the lag phase for the accommodation to cadmium, on the other hand, Cd:Sulfide complex system secondarily correlated it during the stationary phase. These results implied that polyphosphate system and Cd:sulfide complex system, these two systems were operated compensatively each other. Considering the results obsdrved with EM and examined tha changes of sulfide and polyphosphate amount, it was reflected that Cd:S complex was located at the cell surface. In the results of $in-vivo^{31}$P NMR spectra in the cells with cadmium pressure, several phosphate signals arose newly from the polyphosphate region with moving chemical shift of it. This phinomenon strongly implied the actual existence of Dd:Pi comples and /or Cd:poly-P complex in the cell and also the cellular compartmentalization of cadmium detoxifying mechanism.

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Identification of Antioxidative Component from Stem Bark of Rhus verniciflua (옻나무 껍질에서 분리한 항산화물질의 성분)

  • Kim, Jung-Bae
    • The Korean Journal of Food And Nutrition
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    • v.16 no.1
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    • pp.60-65
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    • 2003
  • An antioxidant compound was obtained from the water extract of the stem bark from Rhus verniciflua, which has been used in traditional folk remedies. The compound was purified by HPLC, using DEAE, CN and ODS columns. The chemical structure of the compound was identified as gallic acid (3,4,5-hydroxylbenzoic acid) by spectral data including UV, IR, EI (HR)-MS, $^1$H-NMR, $\^$13/C-NMR and elemental analyzer. This compound was found show cytotoxicity against HeLa cell ( IC$\_$50/ : 8.5$\mu\textrm{g}$/$m\ell$).

PFG NMR Study of Intra-cellular Drug Uptake in Xenopus laevis Oocyte

  • Kwan, soo-Hong;Yeom Gyo-Seon;Kim, Eun-Hee;Lee, Chul-hyun;Lee, Sang-Do;Cheong, chae-joon
    • Journal of the Korean Magnetic Resonance Society
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    • v.9 no.1
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    • pp.29-37
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    • 2005
  • Intra-cellular drug uptake in Xenopus laevis oocyte has been elucidated using localized MR spectroscopy (MRS) and PFG NMR techniques at a 600 $MH_z$(Bruker, 14.1 T) NMR spectrometer. The localized MRS has been done with a homemade probe, and shows the intra-cellular uptake of nicotinamide. The self-diffusion of the molecule in Xenopus oocyte was obtained by PFG NMR technique. The measured data are well fitted with a linear combination of two exponential functions, which shows that there are two types of drug molecules, intra-and extra-cellular molecules. Diffusion coefficients of intra- and extra-cellular drug molecules are 3.7 $\times$ $10^{-11}$ $\m^{2}/s$and 6.4 $\times$ $10^{-10}$ $\m^{2}/s$, respectively. In the weighting factors there is shown that about 5% of drug molecule is inside the cells. These techniques can be used for drug screening in molecule-, cell-, and tissue-based preclinical test.

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Structure-Activity Relationship of the N-terminal Helix Analog of Papiliocin, PapN

  • Jeon, Dasom;Jeong, Min-Cheol;Kim, Jin-Kyoung;Jeong, Ki-Woong;Ko, Yoon-Joo;Kim, Yangmee
    • Journal of the Korean Magnetic Resonance Society
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    • v.19 no.2
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    • pp.54-60
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    • 2015
  • Papiliocin, from the swallowtail butterfly, Papilio xuthus, shows high bacterial cell selectivity against Gram-negative bacteria. Recently, we designed a 22mer analog with N-terminal helix from $Lys^3$ to $Ala^{22}$, PapN. It shows outstanding antimicrobial activity against Gram-negative bacteria with low toxicity against mammalian cells. In this study, we determined the 3-D structure of PapN in 300 mM DPC micelle using NMR spectroscopy and investigated the interactions between PapN and DPC micelles. The results showed that PapN has an amphipathic ${\alpha}$-helical structure from $Lys^3$ to $Lys^{21}$. STD-NMR and DOSY experiment showed that this helix is important in binding to the bacterial cell membrane. Furthermore, we tested antibacterial activities of PapN in the presence of salt for therapeutic application. PapN was calcium- and magnesium-resistant in a physiological condition, especially against Gram-negative bacteria, implying that it can be a potent candidate as peptide antibiotics.

Paratope Mapping of Anti-Ex-A IgG as Studied by NMR (NMR에 의한 anti-Ex-A IgG의 항원결합부위 해석)

  • Kim, Ha-Hyeong;Lee, Gwang-Pyo
    • YAKHAK HOEJI
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    • v.40 no.4
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    • pp.422-427
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    • 1996
  • The anti-Ex-A IgG was specifically labeled with stable isotopes, DL-His-2,4-$d_2$, L-Phe-$d_5$, L-Trp-$d_5$, L-Tyr-2,6-$d_2$ and L-[1-$^{13}C$]Trp, by growing hybridoma cell in serum-free medium. By use of NMR spectroscopy with selectively labeled Fab fragment, we applied a paratope mapping on antigen-antibody complex. Assignments of the observed carbonyl carbon resonances have been determined by using $^{13}C$-$^{15}N$ double labeling method in order to assign the Trp resonances. Photo CIDNP was also applied to investigate the antigen-binding site(s) on the surface residues of antibody. We found that Trp 36, which is located at the $V_H$ domain, is an important residue to bind to Ex-A, however, two Tyr on the surface of anti-Ex-A IgG plays no crucial role to bind to antigen. On the basis of these results, we demonstrate that stable isotope-aided NMR strategy can be extended to molecular structural analyses of the complex of an Fab fragment and a protein antigen.

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Optimization of Expression, Purification, and NMR Measurement for Structural Studies of Syndecan-4 Transmembrane Region

  • Park, Tae-Joon;Lee, Min-Hye;Choi, Sung-Sub;Kim, Yong-Ae
    • Journal of the Korean Magnetic Resonance Society
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    • v.15 no.1
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    • pp.25-39
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    • 2011
  • Syndecan-4 is a transmembrane heparan sulfate proteoglycan, which is a coreceptor with integrins in cell adhesion. To get better understand the mechanism and function of Syndecan-4, it is critical to elucidate the three-dimensional structure of a single transmembrane spanning region of them. Unfortunately, it is hard to prepare the peptide because syndecan-4 is membrane-bound protein that transverse the lipid bilayer of the cell membrane. Generally, the preparation of transmembrane peptide sample is seriously difficult and time-consuming. In fact, high yield production of transmembrane peptides has been limited by experimental adversities of insufficient yields and low solubility of peptide. Here, we demonstrate experimental processes and results to optimize expression, purification, and NMR measurement condition of Syndecan-4 transmembrane peptide.

Quantification of Methanol Concentration in the Polymer Electrolyte Membrane of Direct Methanol Fuel Cell by Solid-state NMR

  • Kim, Seong-Soo;Paik, Youn-Kee;Kim, Sun-Ha;Han, Oc-Hee
    • Journal of the Korean Magnetic Resonance Society
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    • v.12 no.2
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    • pp.96-102
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    • 2008
  • Direct quantification of methanol in polymer electrolyte membrane (PEM) by solid-state nuclear magnetic resonance (NMR) spectroscopy was studied and the methanol concentrations in PEM produced by crossover and diffusion were compared. The error range of the quantification was not smaller than ${\pm}15%$ and the amount of the methanol crossed over in our direct methanol fuel cells (DMFCs) was less than the methanol diffused to PEM. The methanol concentration in the PEM of the DMFC operated at different current densities were equivalent.

Purification and Characterization of the Regulatory Substance of Furfural Biodegradation in Pseudomonas fluorescens (Pseudomonas fluorescens에 의한 Furfural의 분해대사 조절물질에 관하여)

  • 이병웅;유병설;이계준;하영칠
    • Korean Journal of Microbiology
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    • v.23 no.4
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    • pp.241-247
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    • 1985
  • The objectives of this study were to isolate and identify ninhydrin positive substande(s) produced in the culture broth of Pseudomonas fluorescens. It was found that the NPS could stimulate bioconversion of furfural into furoic acid. In order to isolate the NPS from the culture broth, cell free filtrate was subjected to ion-exchange chromatography, gel-permeation and finally to cellulose column chromatography. The purified NPS was white amorphous power and very soluble in water, slightly soluble in methanol and very insoluble in organic solvents. UV, and IR absorption spectra. $^H$ and $^{13}C-NMR$ were measured in order to identify the chemical structure of the NPS.

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