• Title/Summary/Keyword: Turn conformation

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Solution Structure of the D/E Helix Linker of Skeletal Troponin-C: As Studied by Circular Dichroism and Two-Dimensional NMR Spectroscopy

  • 이원태;G. M. Anatharamaiah;Herbert C. Cheung;N. Rama Krishna
    • Bulletin of the Korean Chemical Society
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    • v.19 no.1
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    • pp.57-62
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    • 1998
  • We have synthesized a 17-residue peptide with the amino acid sequence RQMKEDAKGKSEEELAD corresponding to residues 84-100 of chicken skeletal troponin C. This stretch of the protein sequence is in the middle one-third of the 32-residue 9-turn α-helix that connects the two globular domains of the dumbell-shaped molecule and includes the D/E linker helix. We describe here the solution conformation of the helix linker as studied by circular dichroism (CD) and two-dimensional nuclear magnetic resonance (2-D NMR) spectroscopy. The NOE connectivities together with the vicinal $^3J_{N{\alpha}}$ coupling constants suggest that the peptide exists in a fast conformational equilibrium among several secondary structure: a nascent helix near the N-terminus, a helix, and a substational population of extended and random coil forms. In addition, two interresidue α-α NOEs are observed suggesting a bent structure with a bend that includes the single glycine in position 92. These results are consistent with the ideas that in neutral solution the D/E linker region of the central helix in troponin C can adopt a helical conformation and the central helix may have a segmental flexibility around Gly 92.

Production of pediocin by Chemical Synthesis and Bactericidal Mode of Action

  • Koo, Min-Seon;Kim, Wang-June;Kwon, Dea-Young;Min, Kyung-Hee
    • Proceedings of the Korean Society for Applied Microbiology Conference
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    • 2001.06a
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    • pp.149-153
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    • 2001
  • To investigate the mode of bactericidal action for antimicrobial peptide, pediocin, synthetic and mutant pediocins were prepared by direct chemical synthesis. Native pediocin was purified from Pedio-coccus acidilactici M and its conformational structure and bactericidal functions were analyzed and compared to synthetic pediocin. Schematic mode of pediocin actions, how pediocin binds on the target cell membrane, penetrates and makes tunnel are proposed. For these purposes, primary and secondary structures of pediocin was analyzed and disulfide bond assignment was also done. The pediocin purified from P. acidilactici M had high effective bactericidal ability against gram positive bacteria, especially Listeria monocytogenes and was very stable at extreme pHs and even at high temperatures such as autoclaving temperature (121$^{\circ}C$). Pediocin was consisted of 44 amino acids with four cysteines. Novel synthetic peptides were achieved by solid phase peptide synthesis(SPPS) method. To explain the function of cysteine in C-terminal region, mutant pediocin, Ped[C24A+C44A], was synthesized and their structural and biological functions were analyzed. Second mutant pediocin, Ped[KllE], was prepared to explain the function of lysine at 11 of N-terminal part of pediocin, especially loop of $\beta$-sheet, and to predict the initial binding site of pediocin. The native and synthetic pediocins was showed random coil conformation by spectropolarimetry in moderate conditions. This conformation was observed in extreme conditions such as high temperature and low and high pHs, also. Circular dichroism(CD) data also showed the existence of $\beta$-turn structure in N-terminal part both native and synthetic pediocins. A structural model for pediocin predicts that 18 amino acids in the N-terminal part of the peptide assume a three-strand $\beta$-sheet conformation. This random coil in C-terminal part of pediocin was converted to folding structure, helix structure, in nonpolar solvents such as alcohol and TFE. The disulfide bond between $^{9}$ Cys and $^{14}$ Cys was concrete and inevitable, however, evidences of disulfide bond between $^{24}$ Cys and $^{44}$ Cys was not. Data of Ped[C24A+C44A], pediocin mutant showed that $^{44}$ Cys was required during killing the target cells but not inevitable, since Ped[C24A+C44A] still have bactericidal activity but much less than native pediocin. Another pediocin mutant, Ped[KllE], had still bactericidal activity, was controversial to propose that positive charge like as $^{11}$ Lys in loop or hinge in bacteriocin bound or helped to binding to microorganism with electrostatic interaction between cell membrane especially teichoic acid and positive amino acid nonspecifically. The conformation of pediocin among native, synthetic and mutant pediocins did not show big difference. The conformations between oxidized and reduced pediocin were almost similar regardless of native or synthetic.

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Solution Structure of pA2, the Mimotopic Peptide of Apolipoprotein A-I, by NMR Spectroscopy

  • Won, Ho-Shik
    • Bulletin of the Korean Chemical Society
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    • v.32 no.11
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    • pp.4016-4020
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    • 2011
  • A number of mimetic peptides of apolipoprotein A-I, a major component for high density lipoproteins (HDL), were screened from the phase-displayed random peptide library by utilizing monoclonal antibodies (A12). A mimetic peptide for A12 epitope against apolipoprotein A-I was selected as FVLVRDTFPSSVCCP(pA2) exhibiting 45% homology with Apo A-I in the BLAST search. Solution structure determination of this mimotope was made by using 2D-NMR data and NMR-based distance geometry (DG)/molecular dynamic calculations. The resulting DG structures had low penalty value of 0.4-0.6 ${\AA}^2$ and the total RMSD of 0.7-1.7 ${\AA}$. The mimotope pA2 exhibited a characteristic ${\beta}$-turn conformation from Val[2] to Phe[8] near Pro[9] residue.

NMR Studies of Metal-binding Luteinizing Hormone Releasing Hormone

  • Won, Ho-Shik
    • Bulletin of the Korean Chemical Society
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    • v.32 no.11
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    • pp.4021-4026
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    • 2011
  • Functions of the luteinizing hormone releasing hormone (LHRH) and its induced release by divalent metal ions have received great attention because this neurotransmitter subsequently regulates the secretion of luteinizing hormone (LH). Metal-LHRH complexes were synthesized by addition of various Cu(II),Ni(II),Zn(II) ions into LHRH in order to understand how the induced release of LHRH is possible. The degree of complexation was monitored by $^1H$, $^{13}C$-NMR chemical shifts, and final products were identified by Mass spectrometry. Solutionstate structure determination of Zn(II)-LHRH out of metal-complexes was accomplished by using NMR and NMR-based distance geometry (DG). Interproton distance information from nuclear Overhauser effect spectroscopy was utilized for structure determination. Structure obtained in this study has a cyclic conformation exhibiting a specific ${\alpha}$-helical turn with residue numbers His[2]-Leu[7] out of 10 amino acids. Comparison of chemical shifts and EPR studies of Ni(II),Cu(II)-LHRH complexes exhibit that these metal complexes have 4-coordination geometry.

Solution State Structure of pA1, the Mimotopic Peptide of Apolipoprotein A-I, by NMR Spectroscopy

  • Kim, Hyo-Joon;Won, Ho-Shik
    • Bulletin of the Korean Chemical Society
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    • v.32 no.9
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    • pp.3425-3428
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    • 2011
  • Apolipoprotein A-I (Apo A-I) is a major component for high density lipoproteins (HDL). A number of mimetic peptides of Apo A-I were screened from the phase-displayed random peptide library by utilizing monoclonal antibodies (A12). Mimetic peptide for A12 epitope against Apo A-I was selected as CPFARLPVEHHDVVGL (pA1). From the BLAST search, the mimetic peptide pA1 had 40% homology with Apo A-I. 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 0.3-0.7 ${\AA}^2$ and the total RMSD was 0.6-1.6 ${\AA}$. The mimotope pA1 exhibited characteristic conformation including a ${\beta}$-turn from Pro[7] to His[11].

Structure of CT16 in the C-terminal of Amyloid Precursor Protein Studied by NMR Spectroscopy

  • Lee, Kyoung-Ik;Baek, Dong-Ha;Shin, Song-Yub;Kim, Yang-Mee
    • Journal of the Korean Magnetic Resonance Society
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    • v.8 no.1
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    • pp.19-27
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    • 2004
  • C-terminal fragments of APP (APP-CTs), that contain complete Abeta sequence, are found in neuritic plaques, neurofibrillary tangles and the cytosol of lymphoblastoid cells obtained from AD patients. CT16, Lys649-Asp664 (KKQYTSIHHGVVEVD) has been known as the most toxic part in the C-terminal fragment of amyloid precursor protein (APP). The solution structure of CT16 was investigated using NMR spectroscopy in various membrane-mimicking environments. According to Circular Dichroim (CD) spectra, CT16 has a random structure in aqueous solution, while conformational change was induced by addition of TFE and SDS micelle. Tertiary structure as determined by NMR spectroscopy shows that CT16 has a ${\beta}$-turn conformation in trifluoroethanol-containing aqueous solution.

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Cyclic AMP Receptor Protein Adopts the Highly Stable Conformation at Millimolar cAMP Concentration (높은 cAMP 농도에서 cAMP 수용성 단백질의 열 안정화)

  • Kang, Jong-Baek;Choi, Young
    • Journal of Life Science
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    • v.13 no.5
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    • pp.751-755
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    • 2003
  • Cyclic AMP receptor proteins(CRP) activate many genes in Escherichia coli by binding of cAMP with not fully known mechanism. CRP existed as apo-CRP in the absence of cAMP, $CRP;(cAMP)_2$$_2$ at low(micromolar) cAMP concentration, or $CRP;(cAMP)_4$ at high(millimolar) concentration of cAMP. This study is designed to measure the thermal stability of S83G CRP, which substituted glycine for serine at amino acid 83 position, with CD spectrapolarimeter at 222nm by the constant elevation of temperature from $20^{\circ]C\; to\; 90^{\circ}C\; at\; 1^{\circ}C/min$. The non-linear regression analysis showed that melting temperatures were 68.4, 72.0, and $82.3^{\circ}C$ for no cAMP, 0.1mM cAMP, and 5mM cAMP, respectively. Result showed the strong thermal stability of CRP by binding of additional cAMP molecules to region between the hinge region and helix-turn-helix(HTH) motif at 5mM cAMP concentration.

Purification and Characterization of a Methanol Dehydrogenase Derived from Methylomicrobium sp. HG-1 Cultivated Using a Compulsory Circulation Diffusion System

  • Kim, Hee-Gon;Kim, Si-Wouk
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.11 no.2
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    • pp.134-139
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    • 2006
  • Methanotrophs are microorganisms that possess the unique ability to utilize methane as their sole source of carbon and energy. A novel culture system, known as the compulsory circulation diffusion system, was developed for rapid growth of methanotrophic bacteria. Methanol dehydrogenase (MDH, EC 1.1.99.8) from Methylomicrobium sp. HG-1, which belongs to the type I group of methanotrophic bacteria, can catalyze the oxidation of methanol directly into formaldehyde. This enzyme was purified 8-fold to electrophoretic homogeneity by means of a 4 step procedure and was found in the soluble fraction. The relative molecular weight of the native enzyme was estimated by gel filtration to be 120 kDa. The enzyme consisted of two identical dimers which, in turn, consisted of large and small subunits in an ${\alpha}_2{\beta}_2$ conformation. The isoelectric point was 5.4. The enzymatic activity of purified MDH was optimum at pH 9.0 and $60^{\circ}C$, and remained stable at that temperature for 20 min. MDH was able to oxidize primary alcohols from methanol to octanol and formaldehyde.

Emitting Properties of Poly(3-hexylthiophene) deposited by LB method (LB법에 의한 제막된 poly(3-hexylthiophene)의 발광특성)

  • Seo, Bu-Wan;Kim, Ju-Seung;Gu, Hal-Bon;Lee, Kyung-Sup;Park, Bok-Gi;Park, Gye-Chun
    • Proceedings of the KIEE Conference
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    • 1999.11d
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    • pp.962-964
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    • 1999
  • We studied emitting properties of devices fabricated using the spin-coating and Langmuir-Blodgett[LB] technique. The LB technique has the advantage of precise control of the thickness better than spin-coating method. Poly(3-hexylthiophene)[P3HT] LB films used as the emitting layer in light-emitting devices. LB monolayers were deposited 27 layers onto the indium-tin-oxide[ITO] as Y-type films by the vertical dipping method. The thickness is about 80nm. Absorption spectrum of LB films presented that P3HT is regiorandom conformation. Also, current-voltage-luminance characteristics and electroluminescence spectra of light-emitting devices fabricated by LB method is studied. In current-voltage-luminescence characteristics, turn-on voltage of P3HT LB film LEDs is higher than that of spin-coating LEDs. But electroluminescence spectrum is similar to the spin-coating LEDs. The orange-red light was clearly visible in a darkened room.

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A Study on Chemical Modification of Papermaking Fibers (I) - Improved Physical Characteristics from Partial Carboxymethylated Pulps - (제지용(製紙用) 섬유(纖維)의 화학적(化學的) 개질(改質)에 관한 연구(硏究) (I) - Partial Carboxymethylation 처리에 의한 물성(物性) 향상(向上) -)

  • Choi, Jeong-Heon;Jo, Byoung-Muk;Oh, Jung-Soo
    • Journal of the Korean Wood Science and Technology
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    • v.23 no.2
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    • pp.37-46
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    • 1995
  • The substitution of carboxymethylated hydroxyl group in pulp revealed more hydrophilic than hydroxyl group. And then fibers were more flexible, swell more which leads to better conformation between fibers in turn this raise paper strength. In this paper, we tried to chemical modifyings of recycled fiber, OCCs(old corrugated containers). Many researchers have examined chemical modification of papermaking fiber by partial carboxymethylation(PCM) using a organic solvent processes. We made modified PCM processes adapted waters m replace of the organic solvent. Our testings for the optimum conditions on the new method, conditions as reaction time, temperature, liquor ratios were designed likely plant system. Freenesses(SR$^{\circ}$) were increased following on carboxyl content of the samples. Handsheets of untreated samples and partial carboxymethylated OCCs were made by optimum conditions on different concentrations of the reagent. As results, maximum 25% strength increasing effects were obtained by the new method.

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