Laboratoire d'optique et biosciences
Laboratoire d'optique et biosciences
Laboratoire d'optique et biosciences

Articles

  • Conformation of the c552:aa3 electron transfer complex in Paracoccus denitrificans studied by EPR on oriented samples
    • Lipowski Gérard
    • Liebl Ursula
    • Guigliarelli Bruno
    • Nitschke Wolfgang
    • Schoepp-Cothenet Barbara
    FEBS Letters, Wiley , 2006, 580 (25), pp.5988 . The EPR spectral parameters of aa3 oxidase and cyt c552 from Paracoccus denitrificans were studied in purified oxidase and enriched cyt c552. The orientation of the g-tensors of hemes a and c552 were determined on partially ordered membranes, enriched cyt c552 and a c552:aa3 subcomplex. The known correlation of g-tensor to molecular axes in histidine/methionine ligated hemes permits us to position cyt c552 with respect to the parent membrane. Taken together with previous data on the interaction surface between aa3 oxidase and cyt c552, these results allow us to arrive at a single conformation for the c552:aa3 electron transfer complex. Cop. 2006 Federation of European Biochemical Societies. (10.1016/j.febslet.2006.09.038)
    DOI : 10.1016/j.febslet.2006.09.038
  • Single Lanthanide-doped Oxide Nanoparticles as Donors in Fluorescence Resonance Energy Transfer Experiments
    • Casanova Didier
    • Giaume D.
    • Gacoin Thierry
    • Boilot Jean-Pierre
    • Alexandrou Antigoni
    Journal of Physical Chemistry B, American Chemical Society , 2006, 110(39), pp.19264-19270 .
  • Hydrophobic distal pocket affects NO-heme geminate recombination dynamics in dehaloperoxidase and H64V myoglobin
    • Franzen S.
    • Jasaitis Audrius
    • Belyea J.
    • Brewer S.
    • Casey Romain
    • Macfarlane A.
    • Martin Jean-Louis
    • Stanley R.
    • Vos Marten H.
    Journal of Physical Chemistry B, American Chemical Society , 2006, 110 (29), pp.14483 . The recombination dynamics of NO with dehaloperoxidase ( DHP) from Amphitrite ornata following photolysis were measured by femtosecond time- resolved absorption spectroscopy. Singular value decomposition ( SVD) analysis reveals two important basis spectra. The first SVD basis spectrum reports on the population of photolyzed NO molecules and has the appearance of the equilibrium difference spectrum between the deoxy and NO forms of DHP. The first basis time course has two kinetic components with time constants of tau(11) approximate to 9 ps and tau(12) approximate to 50 ps that correspond to geminate recombination. The fast geminate process tau(11) arises from a contact pair with the heme iron in a bound state with S) 3/2 spin. The slow geminate process tau(12) corresponds to the recombination from a more remote docking site > 3 angstrom from the heme iron with the greater barrier corresponding to a S) 5/ 2 spin state. The second SVD basis spectrum represents a time- dependent Soret band shift indicative of heme photophysical processes and protein relaxation with time constants of tau(21) approximate to 3 ps and tau(22) approximate to 17 ps, respectively. A comparison between the more rapid rate constant of the slow geminate phase in DHP- NO and horse heart myoglobin ( HHMbNO) or sperm whale myoglobin ( SWMbNO) suggests that protein interactions with photolyzed NO are weaker in DHP than in the wild- type MbNOs, consistent with the hydrophobic distal pocket of DHP. The slower protein relaxation rate tau(22) in DHP- NO relative to HHMbNO implies less effective trapping in the docking site of the distal pocket and is consistent with a greater yield for the fast geminate process. The trends observed for DHP- NO also hold for the H64V mutant of SWMb ( H64V MbNO), consistent with a more hydrophobic distal pocket for that protein as well. We examine the influence of solution viscosity on NO recombination by varying the glycerol content in the range from 0% to 90% ( v/ v). The dominant effect of increasing viscosity is the increase of the rate of the slow geminate process, tau(12), coupled with a population decrease of the slow geminate component. Both phenomena are similar to the effect of viscosity on wild- type Mb due to slowing of protein relaxation resulting from an increased solution viscosity and protein surface dehydration. (10.1021/jp056790m)
    DOI : 10.1021/jp056790m
  • Coupling between surface plasmons in subwavelength hole arrays
    • Masson Jean-Baptiste
    • Gallot Guilhem
    Physical Review B: Condensed Matter and Materials Physics (1998-2015), American Physical Society , 2006, 73, pp.121401 .
  • Ionic contrast terahertz near field imaging of axonal water fluxes
    • Masson Jean-Baptiste
    • Sauviat Martin-Pierre
    • Martin Jean-Louis
    • Gallot Guilhem
    Proceedings of the National Academy of Sciences of the United States of America, National Academy of Sciences , 2006, 103 (13), pp.4808-4812 .
  • Parametric cascade downconverter for intense ultrafast mid-infrared generation beyond the Manley-Rowe limit
    • Fraser James M.
    • Ventalon Cathie
    Applied optics, Optical Society of America , 2006, 45 (17), pp.4109-4113 . We propose a scheme to generate intense, ultrafast mid-infrared pulses with conversion efficiencies exceeding the upper bound for single-stage difference-frequency mixing as predicted by the Manley-Rowe relations. Finite-element fast Fourier transform simulations of the mixing process show that the parametric cascade downconverter generates 1.7 times more photons (at 10 μm ) than in the initial pump pulse (center wavelength of 1.48 μm , duration of 130 fs , and pulse energy of 50 μJ ), with negligible pulse spatial and temporal distortion. © 2006 Optical Society of America (10.1364/AO.45.004109)
    DOI : 10.1364/AO.45.004109
  • Measuring the dynamics of circular dichroism in a pump-probe experiment with a Babinet-Soleil compensator
    • Niezborala Claire
    • Hache François
    Journal of the Optical Society of America B, Optical Society of America , 2006, 23 (11), pp.2418 . Circular dichroism contains rich information on the conformation of molecules and, in particular, of biomolecules, and measuring its variation in a pump-probe experiment is very promising but also very challenging. We propose a new technique to measure pump-induced variation of the circular dichroism, which is based on the measurement of the probe ellipticity and its variation with the pump. This technique has the advantage that it does not require modulation of the probe polarization, which allows modulation of the pump intensity. We show theoretically and demonstrate experimentally that this technique is very sensitive and user friendly. We also show that it can be used to measure pump-induced change in the optical rotation, allowing for a complete characterization of pump-induced optical activity. Cop. 2006 Optical Society of America. (10.1364/JOSAB.23.002418)
    DOI : 10.1364/JOSAB.23.002418
  • Nouvelle approche des fibroses par microscopie multiphotonique avec génération de second harmonique [New approach of fibrosis by multiphoton microscopy with second harmonic generation]
    • Hernest Monica
    • Pena Ana-Maria
    • Strupler Mathias
    • Beaurepaire Emmanuel
    • Martin Jean-Louis
    • Schanne-Klein Marie-Claire
    • Tharaux Pierre-Louis
    Médecine/Sciences, EDP Sciences , 2006, 22 (10), pp.820-821 . La fibrose est une réponse adaptative pathologique qui détruit non spécifiquement les tissus. Il s'agit d'un processus universel de réparation des tissus qui survient en réaction à de nombreux types d'agressions telles les contraintes mécaniques, les brûlures, les radiations ionisantes, l'ischémie, l'inflammation. Ces agressions concourent de manière intriquée à la physiopathologie des maladies infectieuses, tumorales ou auto-immunes, de l'hypertension artérielle et des maladies cardio-vasculaires. Le terme de fibrose décrit précisément l'accumulation nouvelle de protéines de la matrice extra-cellulaire selon un arrangement spatial fibrillaire caractéristique. Il s'agit essentiellement de molécules de collagènes de type I et III (voire de type II, V ou XI) synthétisées sous forme de triples hélices elles-mêmes assemblées en fibrilles par les cellules fibroblastiques. L'apparition des collagènes fibrillaires marque un changement qualitatif et quantitatif de composition des collagènes des tissus. Réciproquement, ces changements de la matrice extracellulaire influencent le phénotype des cellules qui y résident. Ainsi dans le rein normal, le collagène de type I n'existe que dans l'adventice artériel. Son apparition au sein des autres structures de cet organe marque une fibrose tubulo-interstitielle qui constitue le meilleur marqueur pronostic défavorable d'une évolution vers l'insuffisance rénale terminale, et ce quelle que soit la maladie causale. Ainsi, comme lors des fibroses compliquant les hépatopathies et les pneumopathies chroniques, les séquelles de brûlures ou d'abrasions cutanées-muqueuses ou encore le remodelage cardiaque et vasculaire, le réarrangement de la géométrie de la matrice extracellulaire altère l'organisation fonctionnelle du tissu considéré. De ce fait, ce processus de réparation a des effets fonctionnels délétères qui constituent un enjeu médical majeur. Les fibrilles de collagène ont des capacités d'auto-assemblage qui sont aussi catalysées et stabilisées ou au contraire empêchées par les enzymes de la matrice extracellulaire. Le développement de la fibrose ou sa régression dépend donc ainsi du bilan des équilibres biologiques de ces mécanismes. Il est donc crucial de caractériser les changements extracellulaires et cellulaires qui font du restutio ad integrum de l'architecture et de la fonction tissulaire un défi biomédical. (10.1051/medsci/20062210820)
    DOI : 10.1051/medsci/20062210820
  • Role of the middle residue in the triple tryptophan electron transfer chain of DNA photolyase: ultrafast spectroscopy of a Trp→Phe mutant
    • Lukacs Andras
    • Eker A.P.M.
    • Byrdin M.
    • Villette Sandrine
    • Pan J.
    • Brettel K.
    • Vos Marten H.
    Journal of Physical Chemistry B, American Chemical Society , 2006, 110 (32), pp.15654-15658 .
  • Endothelial nitric oxide synthase reduces nitrite anions to NO under anoxia
    • Gautier Clément
    • van Faassen E.
    • Mikula I.
    • Martasek P.
    • Slama-Schwok Anny
    Biochemical and Biophysical Research Communications, Elsevier , 2006, 341 (3), pp.816-821 . In this work, we demonstrate that endothelial nitric oxide synthase is capable of anoxic reduction of nitrite anions to nitric oxide at physiological pH by absorption and EPR spectroscopy and electrochemical measurements. The nitrite reduction is achieved at the oxygenase domain of the protein and proceeds even in the absence of the tetrahydrobiopterin cofactor. The nitrite pathway increases by sixfold the NO production with respect to the regular arginine pathway under hypoxia, which is largely blocked. Therefore, basal levels of NO release could be sustained by anoxic nitrite reduction. The reaction suggests a new pathway for fast NO delivery under hypoxia, precisely when the vasodilating properties of nitric oxide are most needed. (c) 2006 Elsevier Inc. All rights reserved. (10.1016/j.bbrc.2006.01.031)
    DOI : 10.1016/j.bbrc.2006.01.031
  • Photoinduced electron transfer from a novel nanotrigger addressed to the NADPH site within the endothelial NO-synthase to the flavin moieties of the protein
    • Beaumont Edward
    • Robin Anne-Claire
    • Berka Vladimir
    • Tsai Ah-Lim
    • Blanchard-Desce Mireille
    • Lambry Jean-Christophe
    • Slama-Schwok Anny
    Nitric Oxide: Biology and Chemistry, Elsevier , 2006, 14 (4), pp.14-15 . We developed a new selective molecular tool to trigger enzymatic activity in a synchronous manner and monitor the sequence of the kinetic events by ultra-fast transient spectroscopy. Our approach is based on a synthetic nanotrigger addressing a selected site within proteins, namely the conserved NADPH binding site common to many enzymes involved in bioreductive processes [1]. The nanotrigger combines a "docking" subunit responsible for the recognition of NADPH sites within proteins and a "chromophoric" subunit responsive to light excitation and able to transfer electrons to the flavin moieties of proteins. We present the first spectroscopic data on such a nanotrigger [1] in the presence of the reductase domain of the endothelial nitric oxide synthase eNOSred. (10.1016/j.niox.2006.04.050)
    DOI : 10.1016/j.niox.2006.04.050
  • Mechanisms involved in the swelling of erythrocytes caused by Pacific and Caribbean ciguatoxins
    • Sauviat Martin-Pierre
    • Boydron-Le Garrec Raphaële
    • Masson Jean-Baptiste
    • Lewis Richard L.
    • Vernoux Jean-Paul
    • Molgó Jordi
    • Laurent Dominique
    • Benoit Evelyne
    Blood Cells, Molecules and Diseases, Elsevier , 2006, 36(1), pp.1-9 . The mechanisms underlying the swelling of frog red blood cells (RBC), induced by Pacific (P-CTX-1) and Caribbean (C-CTX-1) ciguatoxins (CTXs), were investigated by measuring the length, width and surface of their elliptic shape. P-CTX-1 (0.5 to 5 nM) and C-CTX-1 (1 nM) induced RBC swelling within 60 min. The CTXs-induced RBC swelling was blocked by apamin (1 μM) and by Sr2+ (1 mM). P-CTX-1-induced RBC swelling was prevented and inhibited by H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (27 μM), an inhibitor of soluble guanylate cyclase (sGC), and NOS blockade by NG methyl-l-arginine (l-NMA; 10 μM). Cytochalasin D (cytD, 10 μM) increased RBC surface and mimicked CTX effect but did not prevent the P-CTX-1-induced l-NMA-sensitive extra increase. Calculations revealed that P-CTX-1 and cytD increase RBC total surface envelop and volume. These data strongly suggest that the molecular mechanisms underlying CTXs-induced RBC swelling involve the NO pathway by an activation of the inducible NOS, leading to sGC activation which modulates intracellular cGMP and regulates L-type Ca2+ channels. The resulting increase in intracellular Ca2+ content, in turn, disrupts the actin cytoskeleton, which causes a water influx and triggers a Ca2+-activated K+ current through SK2 isoform channels. (10.1016/j.bcmd.2005.10.007)
    DOI : 10.1016/j.bcmd.2005.10.007
  • True near field versus contrast near field imaging.
    • Masson Jean-Baptiste
    • Gallot Guilhem
    Optics Express, Optical Society of America - OSA Publishing , 2006, 14 (24), pp.11566-11574 . We demonstrate that in near field imaging, interaction between light and sample can be divided into two main areas: the true near field and the contrast near field domain. We performed extensive numerical simulations in order to identify the limits of these areas, and to investigate contrast near field imaging in which much easier propagation calculation can be achieved. Finally, we show an application with terahertz axonal imaging. © 2006 Optical Society of America (10.1364/OE.14.011566)
    DOI : 10.1364/OE.14.011566
  • Optical in situ size determination of single lanthanide-ion doped oxide nanoparticles
    • Casanova Didier
    • Giaume Domitile
    • Beaurepaire Emmanuel
    • Gacoin Thierry
    • Boilot Jean-Pierre
    • Alexandrou Antigoni
    Applied Physics Letters, American Institute of Physics , 2006, 89 (25), pp.253103 . We show that the size of a lanthanide-ion doped nanoparticle can be accurately determined from its luminosity. The optically determined size distribution is in very good agreement with the distribution obtained from transmission electron microscopy. These data confirm that single nanoparticles are visualized in microscopy experiments. Nanoparticles as small as 13 nm are detectable with integration times of 500 ms. (c) 2006 American Institute of Physics. (10.1063/1.2405871)
    DOI : 10.1063/1.2405871
  • A Hydrophobic Distal Pocket Affects NO-heme Geminate Recombination Dynamics in Dehaloperoxidase and H64V Myoglobin
    • Franzen Stefan
    • Jasaitis Audrius
    • Belyea Jennifer
    • Brewer Scott
    • Casey Romain
    • Macfarlane a W Iv
    • Stanley R.
    • Vos Marten H.
    • Martin Jean-Louis
    Journal of Physical Chemistry B, American Chemical Society , 2006, 110, pp.14483-14493 .
  • Interferometric Fourier transform coherent anti-stokes Raman scattering
    • Cui M.
    • Joffre Manuel
    • Skodack J.
    • Ogilvie Jennifer P.
    Optics Express, Optical Society of America - OSA Publishing , 2006, 14 (18), pp.8448-8458 . We present an interferometric time-domain Fourier transform implementation of coherent anti-Stokes Raman scattering (CARS). Based on a single femtosecond laser source, the method provides a straight-forward scheme for obtaining high resolution CARS spectra. We give a theoretical description of the method, and demonstrate good agreement between simulation and experimental CARS spectra. We also discuss the method's relation to other CARS approaches for microscopy and microspectroscopy applications. Cop. 2006 Optical Society of America. (10.1364/OE.14.008448)
    DOI : 10.1364/OE.14.008448
  • Dynamics of NO rebinding to the heme domain of NO synthase-like proteins from bacterial pathogens
    • Gautier Christian
    • Martasek P.
    • Mikula I.
    • Nioche P.
    • Raman C.S.
    • Slama-Schwok Anny
    Nitric Oxide , 2006, 15 (4), pp.312 . Some Gram-positive bacterial pathogens harbor a gene that encodes a protein (HNS, Heme domain of NO Synthase-like proteins) with striking sequence identity to the oxygenase domain of mammalian NO synthases (NOS). However, they lack the N-terminal and the Zn-cysteine motif participating to the stability of an active dimer in the mammalian isoforms. The unique properties of HNS make it an excellent model system for probing how the heme environment tunes NO dynamics and for comparing it to the endothelial NO synthase heme domain (eNOSHD) using ultrafast transient spectroscopy. NO rebinding in HNS from Staphylococcus aureus (SA-HNS) is faster than that measured for either Bacillus anthracis (BA-HNS) or for eNOSHD in both oxidized and reduced forms in the presence of arginine. To test whether these distinct rates arise from different energy barriers for NO recombination, we measured rebinding kinetics at several temperatures. Our data are consistent with different barriers for NO recombination in SA-HNS and BA-HNS and the presence of a second NO-binding site. The hypothesis that an additional NO-binding cavity is present in BA-HNS is also consistent with the effect of the NO concentration on its rebinding. The lack of the effect of NO concentration on the geminate rebinding in SA-HNS could be due to an isolated second site. We confirm the existence of a second NO site in the oxygenase domain of the reduced eNOS as previously hypothesized [A. Slama-Schwok, M. Négrerie, V. Berka, J.C. Lambry, A.L. Tsai, M.H. Vos, J.L. Martin, Nitric oxide (NO) traffic in endothelial NO synthase. Evidence for a new NO binding site dependent on tetrahydrobiopterin? J. Biol. Chem. 277 (2002) 7581-7586]. This site requires the presence of arginine and BH4; and we propose that NO dynamic and escape from eNOS is regulated by the active site H-bonding network connecting between the heme, the substrate, and cofactor. Cop. 2006 Elsevier Inc. All rights reserved. (10.1016/j.niox.2006.03.010)
    DOI : 10.1016/j.niox.2006.03.010
  • Time-resolved circular dichroism in carbonmonoxy-myoglobin: The central role of the proximal histidine
    • Dartigalongue Thibault
    • Hache François
    Chirality, Wiley , 2006, 18 (4), pp.273 . A calculation of the circular dichroism (CD) spectra of carbonmonoxy-and deoxy-myoglobin is carried out in relation to a time-resolved CD experiment. This calculation allows us to assign a dominant role to the proximal histidine in the definition of the electronic normal modes and to interpret the transient CD structure observed in a strain of the proximal histidine. This strain builds up in 10 ps and relaxes in 50 ps as the protein evolves towards its deoxy form. Cop. 2006 Wiley-Liss, Inc. (10.1002/chir.20254)
    DOI : 10.1002/chir.20254
  • Generation and complete characterization of intense mid-infrared ultrashort pulses
    • Ventalon Cathie
    • Fraser James M.
    • Likforman Jean-Pierre
    • Villeneuve D. M.
    • Corkum Paul B.
    • Joffre Manuel
    Journal of the Optical Society of America B, Optical Society of America , 2006, 23, pp.332 .
  • Role of heme iron coordination and protein structure in the dynamics and geminate rebinding of nitric oxide to the H93G myoglobin mutant: Implications for nitric oxide sensors
    • Négrerie Michel
    • Kruglik Sergei G.
    • Lambry Jean-Christophe
    • Vos Marten H.
    • Martin Jean-Louis
    • Franzen S.
    Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology , 2006, 281, pp.10389 . The influence of the heme iron coordination on nitric oxide binding dynamics was investigated for the myoglobin mutant H93G (H93G-Mb) by picosecond absorption and resonance Raman time-resolved spectroscopies. In the H93G-Mb, the glycine replacing the proximal histidine does not interact with the heme iron so that exogenous substituents like imidazole may coordinate to the iron at the proximal position. Nitrosylation of H93G-Mb leads to either 6- or 5-coordinate species depending on the imidazole concentration. At high concentrations, (imidazole)-(NO)-6-coordinate heme is formed, and the photoinduced rebinding kinetics reveal two exponential picosecond phases (~10 and ~100 ps) similar to those of wild type myoglobin. At low concentrations, imidazole is displaced by the trans effect leading to a (NO)-5-coordinate heme, becoming 4-coordinate immediately after photolysis as revealed from the transient Raman spectrum. In this case, NO rebinding kinetics remain bi-exponential with no change in time constant of the fast component whose amplitude increases with respect to the 6-coordinate species. Bi-exponential NO geminate rebinding in 5-coordinate H93G-Mb is in contrast with the single-exponential process reported for nitrosylated soluble guanylate cyclase (Negrerie, M., Bouzhir, L., Martin, J. L., and Liebl, U. (2001) J. Biol. Chem. 276, 46815-46821). Thus, our data show that the iron coordination state or the heme iron out-of-plane motion are not at the origin of the bi-exponential kinetics, which depends upon the protein structure, and that the 4-coordinate state favors the fast phase of NO geminate rebinding. Consequently, the heme coordination state together with the energy barriers provided by the protein structure control the dynamics and affinity for NO-binding enzymes. Cop. 2006 by The American Society for Biochemistry and Molecular Biology, Inc. (10.1074/jbc.M513375200)
    DOI : 10.1074/jbc.M513375200
  • Sensitivity of cardiac background inward rectifying K+ outward current (IK1) to the alkaloids lepadiformines A, B, and C
    • Sauviat Martin-Pierre
    • Vercauteren J.
    • Grimaud N.
    • Jugé M.
    • Nabil M.
    • Petit J.-Y.
    • Biard J.-F.
    Journal of Natural Products, American Chemical Society , 2006, 69 (4), pp.558 . Three marine alkaloids, purified from Clavelina moluccensis, were structurally identified as lepadiformines A, B, and C and studied on frog atrial myocytes IK1, using the patch-clamp technique. Lepadiformine A (0.4 to 3.3 μM) blocked IK1 dose-dependently with an apparent dissociation constant (KD) equal to 1.42 μM and a stoichiometry of 0.77. The block is voltage-dependent, suggesting that lepadiformine A occupies a receptor site located at about two-thirds of the membrane depth. The shortening of the aliphatic chain at position C13 of lepadiformine B decreased the potency of the molecule to block IK1 but not the affinity (KD = 1.56 μM) and stoichiometry (0.72). Additional deletion of the oxygenated side chain at C2 in lepadiformine C markedly decreased the inhibitory effect of the molecule. In conclusion, lepadiformine modulates IK1 response in cardiac muscle. The oxygenated side chain in C2 is implicated in the affinity of lepadiformine, which behaved as an amine, for a receptor located near or inside the IK1 pore, and the aliphatic chain length at position C13 is involved in the degree of IK1 blockage. (10.1021/np050215s)
    DOI : 10.1021/np050215s
  • Imaging lipid bodies in cells and tissues using third-harmonic generation microscopy.
    • Débarre Delphine
    • Supatto Willy
    • Pena Ana-Maria
    • Fabre Aurelie J
    • Tordjmann Thierry
    • Combettes Laurent
    • Schanne-Klein Marie-Claire
    • Beaurepaire Emmanuel
    Nature Methods, Nature Publishing Group , 2006, 3 (1), pp.47-53 . Lipid bodies have an important role in energy storage and lipid regulation. Here we show that lipid bodies are a major source of contrast in third-harmonic generation (THG) microscopy of cells and tissues. In hepatocytes, micrometer-sized lipid bodies produce a THG signal 1-2 orders of magnitude larger than other structures, which allows one to image them with high specificity. THG microscopy with approximately 1,200 nm excitation can be used to follow the distribution of lipid bodies in a variety of unstained samples including insect embryos, plant seeds and intact mammalian tissue (liver, lung). We found that epi-THG imaging is possible in weakly absorbing tissues because bulk scattering redirects a substantial fraction of the forward-generated harmonic light toward the objective. Finally, we show that the combination of THG microscopy with two-photon and second-harmonic imaging provides a new tool for exploring the interactions between lipid bodies, extracellular matrix and fluorescent compounds (vitamin A, NADH and others) in tissues. (10.1038/nmeth813)
    DOI : 10.1038/nmeth813
  • Glu-Q-tRNAAsp synthetase coded by the yadB gene, a new paralog of aminoacyl-tRNA synthetase that glutamylates tRNAAsp anticodon
    • Blaise Mickaël
    • Becker Hubert F.
    • Lapointe Jacques
    • Cambillau Christian
    • Giegé Richard
    • Kern Daniel
    Biochimie, Elsevier , 2005, 87 (9-10), pp.847-861 . Analysis of the completed genome sequences revealed presence in various bacteria of an open reading frame (ORF) encoding a polypeptide chain presenting important similarities with the catalytic domain of glutamyl-tRNA synthetases but deprived of the C-terminal anticodon-binding domain. This paralog of glutamyl-tRNA synthetases, the YadB protein, activates glutamate in the absence of tRNA and transfers the activated glutamate not on tRNA(Glu) but instead on tRNA(Asp). It has been shown that tRNA(Asp) is able to accept two amino acids: aspartate charged by aspartyl-tRNA synthetase and glutamate charged by YadB. The functional properties of YadB contrast with those of the canonical glutamyl-tRNA synthetases, which activate Glu only in presence of the cognate tRNA before aminoacylation of the 3'-end of tRNA. Biochemical approaches and mass spectrometry investigations revealed that YadB transfers the activated glutamate on the cyclopenthene-diol ring of the modified nucleoside queuosine posttranscriptionally inserted at the wobble position of the anticodon-loop to form glutamyl-queuosine. Unstability of the ester bond between the glutamate residue and the cyclopenthene-diol (half-life 7.5 min) explains why until now this modification escaped detection. Among Escherichia coli tRNAs containing queuosine in the wobble position, only tRNA(Asp) is substrate of YadB. Sequence comparison reveals a structural mimicry between the anticodon-stem and loop of tRNA(Asp) and the amino acid acceptor-stem of tRNA(Glu). YadB, renamed glutamyl-Q-tRNA(Asp) synthetase, constitutes the first enzyme structurally related to aminoacyl-tRNA synthetases which catalyzes a hypermodification in tRNA, and whose function seems to be conserved among prokaryotes. The discovery of glutamyl-Q-tRNA(Asp) synthetase breaks down the current paradigm according to which the catalytic domain of aminoacyl-tRNA synthetases recognizes the amino acid acceptor-stem of tRNA and aminoacylates the 3'-terminal ribose. The evolutionary significance of the existence of an aminoacyl-tRNA synthetase paralog dedicated to the hypermodification of a tRNA anticodon will be discussed. (10.1016/j.biochi.2005.03.007)
    DOI : 10.1016/j.biochi.2005.03.007
  • Structure sensitivity in third-harmonic generation microscopy
    • Débarre Delphine
    • Supatto Willy
    • Beaurepaire Emmanuel
    Optics Letters, Optical Society of America - OSA Publishing , 2005, 30, pp.2134-2136 . We characterize experimentally the influence of sample structure and beam focusing on signal level in third-harmonic generation (THG) microscopy. In the case of a homogeneous spherical sample, the dependence of the signal on the size of the sphere can be controlled by modifying the Rayleigh length of the excitation beam. More generally, the influence of excitation focusing on the signal depends on sample geometry, allowing one to highlight certain structures within a complex system. We illustrate this point by focusing-based contrast modulation in THG images of Drosophila embryos. (10.1364/OL.30.002134)
    DOI : 10.1364/OL.30.002134
  • Role of arginine 220 in the oxygen sensor FixL from Bradyrhizobium japonicum
    • Balland Véronique
    • Bouzhir-Sima Latifa
    • Kiger Laurent
    • Marden Michael C.
    • Vos Marten H.
    • Liebl Ursula
    • Mattioli Tony A.
    Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology , 2005, 280, pp.15279-15288 . In the heme-based oxygen sensor protein FixL, conformational changes induced by oxygen binding to the heme sensor domain regulate the activity of a neighboring histidine kinase, eventually restricting expression of specific genes to hypoxic conditions. The conserved arginine 220 residue is suggested to play a key role in the signal transduction mechanism. To obtain detailed insights into the role of this residue, we replaced Arg220 by histidine (R220H), glutamine (R220Q), glutamate (R220E), and isoleucine (R220I) in the heme domain FixLH from Bradyrhizobium japonicum. These mutations resulted in dramatic changes in the O2 affinity with Kd values in the order R220I < R220Q < wild type < R220H. For the R220H and R220Q mutants, residue 220 interacts with the bound O2 or CO ligands, as seen by resonance Raman spectroscopy. For the oxy-adducts, this H-bond modifies the π acidity of the O2 ligand, and its strength is correlated with the back-bonding-sensitive ν4 frequency, the koff value for O2 dissociation, and heme core-size conformational changes. This effect is especially strong for the wild-type protein where Arg220 is, in addition, positively charged. These observations strongly suggest that neither strong ligand fixation nor the displacement of residue 220 into the heme distal pocket are solely responsible for the reported heme conformational changes associated with kinase activity regulation, but that a significant decrease of the heme π* electron density because of strong back-bonding toward the oxygen ligand also plays a key role. (10.1074/jbc.M413928200)
    DOI : 10.1074/jbc.M413928200