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Accueil > Départements > Biochimie, Biophysique et Biologie Structurale > Sun UN : RPE Haut Champ des systèmes biologiques

Publications de l’équipe

2019


  • S. Leach, N. C. Jones, S. V. Hoffmann, et S. Un, « Isoquinoline gas-phase absorption spectrum in the vacuum ultraviolet between 3.7 and 10.7 eV. New valence and Rydberg electronic states », Rsc Advances, vol. 9, nᵒ 9, p. 5121-5141, févr. 2019.
    Résumé : VUV photons from a synchrotron source were used to record the gas-phase absorption spectrum of isoquinoline over the range 3.5 to 10.7 eV. The rich spectrum exhibits both broad and sharp features, of varying intensities, that are analyzed into eight valence and eight Rydberg transitions. Previous data on the valence transitions of isoquinoline were essentially limited to solution spectra up to 5.4 eV. Our study increases their number considerably. The features in the 3.96 eV region are discussed in terms of vibronic coupling between the n pi* 1(1)A '' and pi pi*2(1)A' valence electronic states. The intensities of some spectral features are augmented by collective p-electron modes considered to be of plasmon-type. Assignments of the valence transitions were facilitated by our DFT calculations and by earlier Pariser-Parr-Pople MO calculations. The calculation results are compared and their relative value is discussed. The DFT calculations reproduce very well a number of experimentally determined properties of the ground state of isoquinoline, in particular its bond distances and angles, rotational constants, vibrational frequencies and dipole moment. No Rydberg series of isoquinoline have previously been observed. Three of the newly observed Rydberg series converge to the D-0 electronic ground state of the ion, while two converge to the D-1 and three to the D-3 excited electronic states of the cation. Astrophysical applications of the VUV absorption spectrum of isoquinoline, in particular the measured absorption cross-sections, are briefly discussed. A comparison between the absorption spectra of isoquinoline and quinoline highlights their similarities and differences, related to their respective molecular orbitals.
    Mots-clés : B3S, BHFMR, density-functional theory, excitation-energies, infrared-spectroscopy, ionization-potentials, molecules, photoelectron-spectroscopy, polycyclic aromatic-hydrocarbons, quinoline, radiationless transitions, vibrational-spectra.

  • M. Nagel, C. E. Seal, L. Colville, A. Rodenstein, S. Un, J. Richter, H. W. Pritchard, A. Börner, et I. Kranner, « Wheat seed ageing viewed through the cellular redox environment and changes in pH », Free Radical Research, vol. 53, nᵒ 6, p. 641-654, juin 2019.
    Résumé : To elucidate biochemical mechanisms leading to seed deterioration, we studied 23 wheat genotypes after exposure to seed bank storage for 6-16 years compared to controlled deterioration (CD) at 45 °C and 14 (CD14) and 18% (CD18) moisture content (MC) for up to 32 days. Under two seed bank storage conditions, seed viability was maintained in cold storage (CS) at 0 °C and 9% seed MC, but significantly decreased in ambient storage (AS) at 20 °C and 9% MC. Under AS and CS, organic free radicals, most likely semiquinones, accumulated, detected by electron paramagnetic resonance, while the antioxidant glutathione (GSH) was partly lost and partly converted to glutathione disulphide (GSSG), detected by HPLC. Under AS the glutathione half-cell reduction potential (EGSSG/2GSH) shifted towards more oxidising conditions, from -186 to -141 mV. In seeds exposed to CD14 or CD18, no accumulation of organic free radicals was observed, GSH and seed viability declined within 32 and 7 days, respectively, GSSG hardly changed (CD14) or decreased (CD18) and EGSSG/2GSH shifted to -116 mV. The pH of extracts prepared from seeds subjected to CS, AS and CD14 decreased with viability, and remained high under CD18. Across all treatments, EGSSG/2GSH correlated significantly with seed viability (r = 0.8, p<.001). Data are discussed with a view that the cytoplasm is in a glassy state in CS and AS, but during the CD treatments, underwent transition to a liquid state. We suggest that enzymes can be active during CD but not under the seed bank conditions tested. However, upon CD, enzyme-based repair processes were apparently outweighed by deteriorative reactions. We conclude that seed ageing by CD and under seed bank conditions are accompanied by different biochemical reactions.
    Mots-clés : Antioxidants, B3S, BHFMR, EPR, ESR, gene bank, germination, organic radicals, seed longevity.

  • M. Strianese, G. J. Palm, D. Kohlhause, L. A. Ndamba, L. C. Tabares, et C. Pellecchia, « Azurin and HS-: Towards Implementation of a Sensor for HS- Detection », European Journal of Inorganic Chemistry, nᵒ 6, p. 885-891, févr. 2019.
    Résumé : Hydrogen sulfide (H2S) is an important biological messenger, which recently joined nitric oxide (NO) and carbon monoxide (CO) as an endogenously produced gasotransmitter. This discovery resulted in a renewed interest towards H2S and, in particular, in its reactivity with biological and bioinorganic targets. However, in the scientific literature studies exploring the reactivity of H2S/HS- with non-heme proteins are rare. In this work, we studied the interaction of HS- with the natural metalloprotein Cu-azurin and its cobalt variant via UV-Vis, fluorescence, and EPR spectroscopy, as well as X-ray diffraction. Interaction of HS- significantly enhances the fluorescence emission of fluorescently labelled copper azurin in buffered solutions and at physiological pH. In contrast, fluorescently labelled cobalt azurin undergoes a fluorescence quenching in the presence of HS- in line with the changes observed in the absorption spectrum of the construct. The results highlight the potential of azurin for implementation in a fast and simple HS- sensor.
    Mots-clés : Azurin, B3S, BHFMR, binding, carbon-monoxide, Cobalt, complexes, Copper, cytochrome-oxidase, Fluorescence, fluorescent-probes, fluorogenic probes, Hydrogen sulfide, hydrogen-sulfide, mechanism, nitric-oxide reactivity, spectroscopic characterization, Structure elucidation.

2018


  • A. Barwinska-Sendra, A. Baslé, K. J. Waldron, et S. Un, « A charge polarization model for the metal-specific activity of superoxide dismutases », Physical chemistry chemical physics: PCCP, janv. 2018.
    Résumé : The pathogenicity of Staphylococcus aureus is enhanced by having two superoxide dismutases (SODs): a Mn-specific SOD and another that can use either Mn or Fe. Using 94 GHz electron-nuclear double resonance (ENDOR) and electron double resonance detected (ELDOR)-NMR we show that, despite their different metal-specificities, their structural and electronic similarities extend down to their active-site 1H- and 14N-Mn(ii) hyperfine interactions. However these interactions, and hence the positions of these nuclei, are different in the inactive Mn-reconstituted Escherichia coli Fe-specific SOD. Density functional theory modelling attributes this to a different angular position of the E. coli H171 ligand. This likely disrupts the Mn-H171-E170' triad causing a shift in charge and in metal redox potential, leading to the loss of activity. This is supported by the correlated differences in the Mn(ii) zero-field interactions of the three SOD types and suggests that the triad is important for determining metal specific activity.
    Mots-clés : B3S, BHFMR.

  • S. Leach, N. C. Jones, S. V. Hoffmann, et S. Un, « VUV Absorption Spectra of Gas-Phase Quinoline in the 3.5-10.7 eV Photon Energy Range », The Journal of Physical Chemistry. A, juill. 2018.
    Résumé : The absorption spectrum of quinoline was measured in the gas phase between 3.5 and 10.7 eV using a synchrotron photon source. A large number of sharp and broad spectral features were observed, some of which have plasmon-type collective π-electron modes contributing to their intensities. Eight valence electronic transitions were assigned, considerably extending the number of π-π* transitions previously observed mainly in solution. The principal factor in solution red-shifts is found to be the Lorentz-Lorenz polarizability parameter. Rydberg bands, observed for the first time, are analyzed into eight different series, converging to the D0 ground and two excited electronic states, namely, D3 and D4, of the quinoline cation. The R1 series limit is 8.628 eV for the first ionization energy of quinoline, a value more precise than previously published. This value, combined with cation electronic transition data, provides precise energies, respectively, 10.623 and 11.355 eV, for the D3 and D4 states. The valence transition assignments are based on density functional theory (DFT) calculations as well as on earlier Pariser-Parr-Pople (P-P-P) self-consistent field linear combination of atomic orbitals molecular orbital results. The relative quality of the P-P-P and DFT data is discussed. Both are far from spectroscopic accuracy concerning electronic excited states but were nevertheless useful for our assignments. Our time-dependent DFT calculations of quinoline are excellent for its ground-state properties such as geometry, rotational constants, dipole moment, and vibrational frequencies, which agree well with experimental observations. Vibrational components of the valence and Rydberg transitions mainly involve C-H bend and C═C and C═N stretch modes. Astrophysical applications of the vacuum UV absorption of quinoline are briefly discussed.
    Mots-clés : B3S, BHFMR.

  • A. Litvinov, A. Feintuch, S. Un, et D. Goldfarb, « Triple resonance EPR spectroscopy determines the Mn2+ coordination to ATP », Journal of Magnetic Resonance, vol. 294, p. 143-152, sept. 2018.
    Résumé : Mn2+ often serves as a paramagnetic substitute to Mg2+, providing means for exploring the close environment of Mg2+ in many biological systems where it serves as an essential co-factor. This applies to proteins with ATPase activity, where the ATP hydrolysis requires the binding of Mg2+-ATP to the ATPase active site. In this context, it is important to distinguish between the Mn2+ coordination mode with free ATP in solution as compared to the protein bound case. In this work, we explore the Mn2+ complexes with ATP, the non-hydrolysable ATP analog, AMPPNP, and ADP free in solution. Using W-band P-31 electron-nuclear double resonance (ENDOR) we obtained information about the coordination to the phosphates, whereas from electron-electron double resonance (ELDOR) - detected NMR (EDNMR) we determined the coordination to an adenosine nitrogen. The coordination to these ligands has been reported earlier, but whether the nitrogen and phosphate coordination is within the same nucleotide molecules or different ones is still under debate. By applying the correlation technique, THYCOS (triple hyperfine correlation spectroscopy), and measuring N-15-P-31 correlations we establish that in Mn-ATP in solution both phosphates and a nitrogen are coordinated to the Mn2+ ion. We also carried out DFT calculations to substantiate this finding. In addition, we expanded the understanding of the THYCOS experiment by comparing it to 2D-EDNMR for Mn-55-P-31 correlation experiments and through simulations of THYCOS and 2D-EDNMR spectra with N-15-P-31 correlations. (C) 2018 Elsevier Inc. All rights reserved.
    Mots-clés : adenosine 5-triphosphate, atp, B3S, BHFMR, crystal-structure, divalent metal, ELDOR detected NMR, eldor-detected nmr, endor, gaussian-type basis, High field EPR, metal-ion-binding, Mn2+ ion, Mn2+-ATP, molecular-orbital methods, nuclear-magnetic-resonance, organic-molecules, ternary complexes, thycos.

2016


  • D. Akhmetzyanov, H. Y. V. Ching, V. Denysenkov, P. Demay-Drouhard, H. C. Bertrand, L. C. Tabares, C. Policar, T. F. Prisner, et S. Un, « RIDME spectroscopy on high-spin Mn(2+) centers », Physical chemistry chemical physics: PCCP, vol. 18, nᵒ 44, p. 30857-30866, nov. 2016.
    Résumé : Pulsed EPR dipolar spectroscopy is a powerful tool for determining the structure and conformational dynamics of biological macromolecules, as it allows precise measurements of distances in the range of 1.5-10 nm. Utilization of high-spin Mn(2+) species as spin probes for distance measurements is of significant interest, because they are biologically compatible and endogenous in numerous biological systems. However, to date dipolar spectroscopy experiments with this kind of species have been underexplored. Here we present pulsed electron electron double resonance (PELDOR also called DEER) and relaxation-induced dipolar modulation enhancement (RIDME) experiments, which have been performed at W-band (94 GHz) and J-band frequencies (263 GHz) on a bis-MnDOTA (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetate) model system. The distances obtained from these experiments are in good agreement with predictions. RIDME experiments reveal a significantly higher modulation depth compared to PELDOR, which is an important consideration for biological samples. These experiments also feature higher harmonics of the dipolar coupling frequency due to effective multiple-quantum relaxation of high-spin Mn(2+) as well as the multiple-component background function. Harmonics of the dipolar coupling frequency were taken into account by including additional terms in the kernel function of Tikhonov regularization analysis.
    Mots-clés : B3S, BHFMR.


  • H. Y. V. Ching, F. C. Mascali, H. C. Bertrand, E. M. Bruch, P. Demay-Drouhard, R. M. Rasia, C. Policar, L. C. Tabares, et S. Un, « The Use of Mn(II) Bound to His-tags as Genetically Encodable Spin-Label for Nanometric Distance Determination in Proteins », The Journal of Physical Chemistry Letters, vol. 7, nᵒ 6, p. 1072-1076, mars 2016.
    Mots-clés : B3S, BHFMR, Electron Spin Resonance Spectroscopy, Histidine, Manganese, Molecular Dynamics Simulation, Peptides, Protein Structure, Tertiary, Proteins, Spin Labels.

  • P. Demay-Drouhard, H. Y. V. Ching, D. Akhmetzyanov, R. Guillot, L. C. Tabares, H. C. Bertrand, et C. Policar, « A Bis-Manganese(II)-DOTA Complex for Pulsed Dipolar Spectroscopy », Chemphyschem: A European Journal of Chemical Physics and Physical Chemistry, vol. 17, nᵒ 13, p. 2066-2078, juill. 2016.
    Résumé : High-spin gadolinium(III) and manganese(II) complexes have emerged as alternatives to standard nitroxide radical spin labels for measuring nanometric distances by using pulsed electron-electron double resonance (PELDOR or DEER) at high fields/frequencies. For certain complexes, particularly those with relatively small zero-field splitting (ZFS) and short distances between the two metal centers, the pseudosecular term of the dipolar coupling Hamiltonian is non-negligible. However, in general, the contribution from this term during conventional data analysis is masked by the flexibility of the molecule of interest and/or the long tethers connecting them to the spin labels. The efficient synthesis of a model system consisting of two [Mn(dota)](2-) (MnDOTA; DOTA(4-) =1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetate) directly connected to the ends of a central rodlike oligo(phenylene-ethynylene) (OPE) spacer is reported. The rigidity of the OPE is confirmed by Q-band PELDOR measurements on a bis-nitroxide analogue. The Mn(II) -Mn(II) distance distribution profile determined by W-band PELDOR is in reasonable agreement with one simulated by using a simple rotamer analysis. The small degree of flexibility arising from the linking MnDOTA arm appears to outweigh the contribution from the pseudosecular term at this interspin distance. This study illustrates the potential of MnDOTA-based spin labels for measuring fairly short nanometer distances, and also presents an interesting candidate for in-depth studies of pulsed dipolar spectroscopy methods on Mn(II) -Mn(II) systems.
    Mots-clés : B3S, BHFMR, EPR spectroscopy, gadolinium, Manganese, pulsed electron-electron double resonance, Spin Labels.


  • F. C. Mascali, H. Y. V. Ching, R. M. Rasia, S. Un, et L. C. Tabares, « Using Genetically Encodable Self-Assembling Gd(III) Spin Labels To Make In-Cell Nanometric Distance Measurements. », Angewandte Chemie International Edition, vol. 55, nᵒ 37, p. 11041-11043, sept. 2016.
    Mots-clés : B3S, BHFMR, EPR spectroscopy, gadolinium, in cell spectroscopy, protein structures, Spin Labels.

2015


  • E. M. Bruch, A. de Groot, S. Un, et L. C. Tabares, « The effect of gamma-ray irradiation on the Mn(II) speciation in Deinococcus radiodurans and the potential role of Mn(II)-orthophosphates », Metallomics: Integrated Biometal Science, vol. 7, nᵒ 5, p. 908-916, mai 2015.
    Résumé : D. radiodurans accumulates large quantities of Mn(II), which is believed to form low molecular weight complexes with phosphate and metabolites that protect D. radiodurans from radiation damage. The concentration of Mn(II) species in D. radiodurans during the exponential and stationary phase was determined using high-field EPR and biochemical techniques. In the exponential growth phase cells a large fraction of the manganese was in the form of Mn(II)-orthophosphate complexes. By contrast, the intracellular concentration of these compounds in stationary phase cells was less than 16 μM, while that of Mn superoxide dismutase was 320 μM and that of another, yet unidentified, Mn(II) protein was 250 μM. Stationary cells were found to be equally resistant to irradiation as the exponential cells in spite of having significant lower Mn(II)-orthophosphate concentrations. Gamma irradiation induced no changes in the Mn(II) speciation. During stationary growth phase D. radiodurans favours the production of the two Mn-proteins over low molecular weight complexes suggesting that the latter were not essential for radio-resistance at this stage of growth.
    Mots-clés : B3S, BHFMR, Deinococcus, Electron Spin Resonance Spectroscopy, Gamma Rays, Manganese, Phosphates.


  • E. M. Bruch, S. Thomine, L. C. Tabares, et S. Un, « Variations in Mn(II) speciation among organisms: what makes D. radiodurans different. », Metallomics, vol. 7, nᵒ 1, p. 136-144, 2015.
    Mots-clés : B3S, BHFMR, BIOCELL, Deinococcus, Escherichia coli, Manganese, MINION, Nuclear Magnetic Resonance, Biomolecular, Nucleic Acids, Phosphates, Saccharomyces cerevisiae, Water.


  • E. M. Bruch, M. T. Warner, S. Thomine, L. C. Tabares, et S. Un, « Pulse Electron Double Resonance Detected Multinuclear NMR Spectra of Distant and Low Sensitivity Nuclei and Its Application to the Structure of Mn(II) Centers in Organisms », The Journal of Physical Chemistry B, vol. 119, nᵒ 43, p. 13515-13523, oct. 2015.
    Mots-clés : B3S, BHFMR, BIOCELL, Deinococcus, Electron Spin Resonance Spectroscopy, Escherichia coli, Manganese, MINION, Models, Molecular, Molecular Structure, Nuclear Magnetic Resonance, Biomolecular, Organometallic Compounds, Quantum Theory.

  • S. Un et E. M. Bruch, « How Bonding in Manganous Phosphates Affects their Mn(II)-(31)P Hyperfine Interactions », Inorganic Chemistry, vol. 54, nᵒ 21, p. 10422-10428, nov. 2015.
    Résumé : Manganous phosphates have been postulated to play an important role in cells as antioxidants. In situ Mn(II) electron-nuclear double resonance (ENDOR) spectroscopy has been used to measure their speciation in cells. The analyses of such ENDOR spectra and the quantification of cellular Mn(II) phosphates has been based on comparisons to in vitro model complexes and heuristic modeling. In order to put such analyses on a more physical and theoretical footing, the Mn(II)-(31)P hyperfine interactions of various Mn(II) phosphate complexes have been measured by 95 GHz ENDOR spectroscopy. The dipolar components of these interactions remained relatively constant as a function of pH, esterification, and phosphate chain length, while the isotropic contributions were significantly affected. Counterintuitively, although the manganese-phosphate bonds are weakened by protonation and esterification, they lead to larger isotropic values, indicating higher unpaired-electron spin densities at the phosphorus nuclei. By comparison, extending the phosphate chain with additional phosphate groups lowers the spin density. Density functional theory calculations of model complexes quantitatively reproduced the measured hyperfine couplings and provided detailed insights into how bonding in Mn(II) phosphate complexes modulates the electron-spin polarization and consequently their isotropic hyperfine couplings. These results show that various classes of phosphates can be identified by their ENDOR spectra and provide a theoretical framework for understanding the in situ (31)P ENDOR spectra of cellular Mn(II) complexes.
    Mots-clés : B3S, BHFMR, Electron Spin Resonance Spectroscopy, Models, Chemical, Organometallic Compounds.


  • H. Y. Vincent Ching, P. Demay-Drouhard, H. C. Bertrand, C. Policar, L. C. Tabares, et S. Un, « Nanometric distance measurements between Mn(ii)DOTA centers. », Phys. Chem. Chem. Phys., vol. 17, nᵒ 36, p. 23368-23377, 2015.
    Mots-clés : B3S, BHFMR, Coordination Complexes, Cysteine, Electron Spin Resonance Spectroscopy, Heterocyclic Compounds, 1-Ring, Maleimides, Manganese, Models, Molecular, Peptides, Protein Structure, Secondary, Spin Labels.
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1. Tabares, L. C. and Un, S. (2013) In situ Determination of Manganese (II) speciation in Deinococcus radiodurans by High Magnetic-Field EPR : detection of high levels of Mn(II) bound to proteins, J. Biol. Chem. 288, 5050–5.

2. Tabares, L. C., Gatjens, J., and Un, S. (2010) Understanding the influence of the protein environment on the Mn(II) centers in Superoxide Dismutases using High-Field Electron Paramagnetic Resonance, Biochim Biophys Acta 1804, 308-317.

3. Sjödin, M., Gatjens, J., Tabares, L. C., Thuery, P., Pecoraro, V. L., and Un, S. (2008) Tuning the redox properties of manganese(II) and its implications to the electrochemistry of manganese and iron superoxide dismutases, Inorg Chem 47, 2897-2908.

4. Gätjens, J., Sjödin, M., Pecoraro, V. L., and Un, S. (2007) The relationship between the manganese(II) zero-field interaction and Mn(II)/Mn(III) redox potential of Mn(4’-X-terpy)2 complexes, J Am Chem Soc 129, 13825-13827.

5- Tabares, L. C., Cortez, N., Agalidis, I., and Un, S. (2005) Temperature-dependent coordination in E. coli manganese superoxide dismutase. J. Am. Chem. Soc. 127, 6039–47

6. Faller, P., Goussias, C., Rutherford, A.W., and Un, S. (2003) Resolving intermediates in biological proton-coupled electron transfer : A tyrosyl radical prior to proton movement, Proc. Natl. Acad. Sci. USA 100, 8732–8735.

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