• Title/Summary/Keyword: covalent

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Fusion Protein Cleavage by Urokinase Covalentley Immobilized to Activated Sepharose Gels (활성화된 Sepharose Gels에 공유결합으로 고정화된 Urokinase를 이용한 융합단백질 절단반응)

  • 서창우;강관엽;이효실;안상점;이은규
    • KSBB Journal
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    • v.15 no.1
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    • pp.42-48
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    • 2000
  • Urokinase (UK), a thrombolytic enzyme used to clear catheters obstructed by blood clots, can be also used industrially in the recombinant protein purification system to cleave a fusion protein linked with a certain fragment of GST. We have immobilized UK by covalent attachment to activated Sepharose 6B-Cl gels and evaluated its performance to cleave a fusion protein of hGH and GST. The Sepharose gels were activated by etherification with glycidol (2,3-epoxypropanol) and further oxidized with periodate resulting in glyceryl-Sepharose gels. After the activation treatment, surface density of the aldehyde groups was 7-30 $\mu$mol-aldehde/mL-gel. Immobilization yield was higher than 99% at high pH (10.5), and the immobilized UK maintained ca. 80% specific activity of the soluble UK. In a column reaction the cleavage yield heavily depended on the feed rate, and it was nearly 86% of that from soluble UK. And the immobilized UK was successfully regenerated by unfolding and refolding with 6M GuHCl. After cleavaging reaction, the monomeric hGH was purified by using expanded bed adsorption chromatography.

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Covalent Interactions of Toluenediisocyanate with DNA and Proteins

  • Jeong, Yo-Chan;Park, Misun;Kim, Dong-Hyun
    • Toxicological Research
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    • v.14 no.4
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    • pp.525-533
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    • 1998
  • The covalent interactions of toluenediisocyanate (TDI) with macromolecules were investigated both in vitro and in vivo. In vitro incubations of 2,4- and 2,6-TDI with DNA or proteins resulted in dose-dependent formation of TDI-protein and TDI-DNA adducts. TDI-treated DNA was highly resistant to enzymatic digestion and thermal hydrolysis, but was readily hydrolyzed under acidic conditions by releasing its corresponding toluenediamine (TDA), suggesting that TDI caused the crosslinking of DNA. Reaction of TDI with albumin and globin resulted in the formation of several adducts, and some adducts were formed in blood of TDI-treated rats in a dose-dependent fashion. Administration of TDI to rats resulted also in a dose-dependent binding of TDI to hepatic tissue. Levels of TDI-albumin adducts were 10 times higher than those of TDI-globin adducts; the biological half lives of TDI-albumin and TDI-globin adducts were 1.2 and 12.5 days, respectively. Globin adducts were detected up to 28 days after the treatment. Hepatic TDI protein adducts were persistent for a substantial period whereas the levels of hepatic TDI-DNA adduct were decreased rapidly. These results indicate that the isocyanato group of TDI is not readily hydrolyzed under physiological conditions, is transported to other organs, and is bound to DNA and/or proteins without further metabolic activation. As the adducted products degrade in the body, TDA is released and introduced to the liver. TDA may additionally bind to hepatic tissue after metabolic activation. Thus, the toxic effect of TDI exposure is considered to persist during the lifetime of the adducted biological macromolecules.

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Are Bound Residues a Solution for Soil Decontamination\ulcorner

  • Bollag, Jean-Marc
    • Proceedings of the Korean Society of Soil and Groundwater Environment Conference
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    • 2003.10a
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    • pp.111-124
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    • 2003
  • Processes that cause immobilization of contaminants in soil are of great environmental importance because they may lead to a considerable reduction in the bioavailability of contaminants and they may restrict their leaching into groundwater. Previous investigations demonstrated that pollutants can be bound to soil constituents by either chemical or physical interactions. From an environmental point of view, chemical interactions are preferred, because they frequently lead to the formation of strong covalent bonds that are difficult to disrupt by microbial activity or chemical treatments. Humic substances resulting from lignin decomposition appear to be the major binding ligands involved in the incorporation of contaminants into the soil matrix through stable chemical linkages. Chemical bonds may be formed through oxidative coupling reactions catalyzed either biologically by polyphenol oxidases and peroxidases, or abiotically by certain clays and metal oxides. These naturally occurring processes are believed to result in the detoxification of contaminants. While indigenous enzymes are usually not likely to provide satisfactory decontamination of polluted sites, amending soil with enzymes derived from specific microbial cultures or plant materials may enhance incorporation processes. The catalytic effect of enzymes was evaluated by determining the extent of contaminants binding to humic material, and - whenever possible - by structural analyses of the resulting complexes. Previous research on xenobiotic immobilization was mostly based on the application of $^{14}$ C-labeled contaminants and radiocounting. Several recent studies demonstrated, however, that the evaluation of binding can be better achieved by applying $^{13}$ C-, $^{15}$ N- or $^{19}$ F-labeled xenobiotics in combination with $^{13}$ C-, $^{15}$ N- or $^{19}$ F-NMR spectroscopy. The rationale behind the NMR approach was that any binding-related modification in the initial arrangement of the labeled atoms automatically induced changes in the position of the corresponding signals in the NMR spectra. The delocalization of the signals exhibited a high degree of specificity, indicating whether or not covalent binding had occurred and, if so, what type of covalent bond had been formed. The results obtained confirmed the view that binding of contaminants to soil organic matter has important environmental consequences. In particular, now it is more evident than ever that as a result of binding, (a) the amount of contaminants available to interact with the biota is reduced; (b) the complexed products are less toxic than their parent compounds; and (c) groundwater pollution is reduced because of restricted contaminant mobility.

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Sensitivity dependence on the effective surface area for cholesterol biosensor fabricated by silanization process (실란화 공정으로 제작된 콜레스테롤 센서의 전극 표면적에 따른 감도 특성)

  • Song, M.J.;Yoon, D.H.;Jin, J.H.;Min, N.K.;Hong, S.I.
    • Proceedings of the KIEE Conference
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    • 2004.07c
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    • pp.2114-2116
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    • 2004
  • 센서의 소형화 되는 추세에 전극 면적은 sensitivity의 중요한 요인이다. 본 연구에서는 콜레스테롤을 측정하기 위해 각각 planar 전극과 porous 전극에 효소 고정화 방법으로 covalent binding인 silanization 공정을 이용하여 전극 면적에 따른 전기화학적 감도를 비교하였다. Handles-Sevcik equation을 이용하여 전극 면적을 구한 결과 planar 전극의 경우 0.1608 $cm^2$, porous 전극의 경우 0.5054 $cm^2$로 porous 전극 면적이 planar전극에 비해 약 3.1배 증가하였다. 또, planar 센서의 sensitivity는 0.08567 ${\mu}A/mM{\cdot}cm^2$이고 porous 센서의 sensitivity는 planar sensor에 비해 약 3.1배 증가한 0.2656 ${\mu}A/mM{\cdot}cm^2$였다. 따라서 porous sensor의 sensitivity의 증가는 전극 면적의 증가에 따른 것이라고 할 수 있다.

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Superconducting Tunnel Junction Detectors for Mass Spectrometry

  • Ohkubo, M.;Zen, N.;Kitazume, T.;Ukibe, M.;Shiki, S.;Koike, M.
    • Progress in Superconductivity
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    • v.14 no.2
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    • pp.77-81
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    • 2012
  • With conventional mass spectrometry (MS), ions are separated according to mass/charge (m/z) ratios. We must speculate the z values to obtain the m values. Superconducting tunnel junction (STJ) detectors can solve this problem, and true mass spectrometry becomes possible instead of m/z spectrometry. The STJ detectors were installed in MS instruments with a variety of ion sources. As an example, we report fragmentation analysis of a non-covalent protein complex of hemoglobin.

Effect of Valence Electron Concentration on Elastic Properties of 4d Transition Metal Carbides MC (M = Y, Zr, Nb, and Rh)

  • Kang, Dae-Bok
    • Bulletin of the Korean Chemical Society
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    • v.34 no.7
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    • pp.2171-2175
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    • 2013
  • The electronic structure and elastic properties of the 4d transition metal carbides MC (M = Y, Zr, Nb, Rh) were studied by means of extended H$\ddot{u}$ckel tight-binding band electronic structure calculations. As the valence electron population of M increases, the bulk modulus of the MC compounds in the rocksalt structure does not increase monotonically. The dominant covalent bonding in these compounds is found to be M-C bonding, which mainly arises from the interaction between M 4d and C 2p orbitals. The bonding characteristics between M and C atoms affecting the variation of the bulk modulus can be understood on the basis of their electronic structure. The increasing bulk modulus from YC to NbC is associated with stronger interactions between M 4d and C 2p orbitals and the successive filling of M 4d-C 2p bonding states. The decreased bulk modulus for RhC is related to the partial occupation of Rh-C antibonding states.

A Strong Dependence of the P-P Bond Length on the Transition Metal Component in ThCr2Si2-Type Phosphides CaM2P2 (M = Fe, Ni): The Influence of d Band Position and σp* Mixing

  • Kang, Dae-Bok
    • Bulletin of the Korean Chemical Society
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    • v.24 no.8
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    • pp.1215-1218
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    • 2003
  • An analysis of the bonding situation in CaM₂P₂ (M=Fe, Ni) with ThCr₂Si₂ structure is made in terms of DOS and COOP plots. The main contributions to covalent bonding are due to M-P and P-P interactions in both compounds. Particularly, the interlayer P-P bonding by variation in the transition metal is examined in more detail. It turns out that the shorter P-P bonds in CaNi₂P₂ form as a result of the decreasing electron delocalization into ${{\sigma}_p}^*$ of P₂ due to the weaker bonding interaction between the metal d and ${{\sigma}_p}^*$ as the metal d band is falling from Fe to Ni.

Purification and Immobilization of Cyclodextrin glucanotransferase from recombinant Bacillus subtilis

  • Seo, Hyo-Jin;Kim, Yeong-Hwa;Kim, Seong-Gu
    • 한국생물공학회:학술대회논문집
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    • 2001.11a
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    • pp.671-674
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    • 2001
  • Cyclodextrin glucanotransferase(CGTase) derived from recombinant Bacillus subtilis was partial purified and concentrated by ultrafiltration. The prepared CGTase were immobilized on various matrices by ionic interaction or covalent bond. CGTase covalently bound on CNBr-activated sepharose 4B were identified to be the highest immobilization activity among various immobilization methods. The optimum conditions for CGTase immobilization were determined; $30^{\circ}C$, 6Orpm, using O.2g CNBr-activated sepharose 4B in pH 6.0 phosphate buffer and 9hr immobilization.

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Silica Sulfuric Acid/Wet $SIO_2$as a Novel System for the Deprotection of Acetals by Using Microwave Irradiation under Solvent Free Conditions (무용매 조건하에서 황산/젖은 $SIO_2$와 마이크로웨이브를 이용한 아세탈의 새로운 탈보호기 방)

  • BiBi Fathemeh, Mirjalili; Mohammad Ali, Zolfigol;Abdolhamid, Bamoniri
    • Journal of the Korean Chemical Society
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    • v.45 no.6
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    • pp.546-548
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    • 2001
  • Neat chlorosulfonic acid reacts with silica gel to give silica sulfuric acid in which sulfuric acid is immobilized on the surface of silica gel via covalent bond. A combination of silica sulfuric acid and wet SiO$_2$ was used as an effective deacetalizating agent for the conversion of acetals to their corresponding carbonyl derivatives by using microwave irradiation under solvent free conditions.

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The Hydrogen Binding Property Study by Density Functional Theory for Zr, V, Fe and Al (밀도 함수를 이용한 지르코니움, 바나듐, 철과 수소와의 반응성 연구)

  • Park, Taesung;Lee, Taeckhong
    • Transactions of the Korean hydrogen and new energy society
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    • v.25 no.6
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    • pp.602-608
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    • 2014
  • The sequence of bond overlap population of metal hydrogen binding is in Al-H > Fe-H > Zr-H > V-H. This results shows the binding energy of Al-H is the biggest in this metals (Al, Fe, Zr, and V) and hydrogen interaction. The Vanadium-hydrogen binding shows the weakest binding energy compared to other metals and it causes easy hydrogen desorption from the corresponding metals. The net charge of Al-H show the biggest value of 0.2248 and the severe localizations of electrons around aluminum and imply strongest covalent binding nature in these metals. This study is applicable to the purification of hydrogen in other bulk gas.