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

Articles

  • Fast ligand and electron transfer dynamics in oxidases and cytochrome c
    • Vos Marten H.
    Biochimica et Biophysica Acta (BBA) - Reviews on Bioenergetics, Elsevier , 2008, 1777 (Supplement), pp.S66 . The active site of heme-copper oxidases contains two cofactors, heme a3 and CuB, which can both bind external ligands as the substrate O2 and signalling molecules NO and CO, and which are both involved in electron transfer processes. Over the last few years we have exploited the fact that the heme-ligand bond can be dissociated by a short light pulse to explore the dynamics of CO and NO in the active site and the interaction between the two cofactors using ultrafast spectroscopic techniques. For example, we have time-resolved the CO transfer from heme a3 to CuB and shown that it occurs in a ballistic way in ∼ 500 fs, which presumably reflects rigidity of the active site. Heme a is located close to heme a3 (∼ 7 Å edge-to-edge) and acts as electron donor for the active site. Using mixed valence oxidases we have extended the 'reverse electron flow' technique to the ultrafast regime and demonstrated that this electron transfer process occurs in only 1.2 ns. The process is activationless and associated with a very low reorganization energy (< 200 meV), in contrast to common assumptions but in general agreement with the hydrophobic environment of the reactants. Finally, ligand dynamics in native and modified cytochrome c reflects the rigidity required for optimal electron transfer properties. (10.1016/j.bbabio.2008.05.259)
    DOI : 10.1016/j.bbabio.2008.05.259
  • Processus ultra-rapides dans les hémoprotéines [Ultrafast processes in heme proteins]
    • Vos Marten H.
    L'Actualité Chimique, Société chimique de France (SCF) , 2008, 317 (52), pp.55 . Les hémoprotéines sont impliquées dans une grande diversité de fonctions biologiques qui incluent transport et stockage d'oxygène, catalyse, transfert d'électron et signalisation. Le fer de l'hème peut être lié aux acides aminés et également aux molécules diatomiques (O2, NO, CO). Ces liaisons peuvent être rompues par une impulsion lumineuse avec un rendement élevé. En utilisant des techniques de spectroscopie optique ultrarapide, cette propriété donne la possibilité unique d'étudier la dynamique structurelle et électronique de l'hème et de son environnement protéique à l'échelle femtoseconde-picoseconde, c'est-à-dire à l'échelle même des vibrations internes de la macromolécule. Cet article décrit des développements récents dans ce domaine, en particulier l'implication de mouvements concertés hème-protéine dans des réactions balistiques de transfert de ligands et la possibilité de suivre des étapes intermédiaires de propagation d'un " signal " au sein de protéines de signalisation. Heme proteins contribute to a large variety of biological functions including transport and storage of oxygen, catalysis, electron transfer and signalling. The iron atom in the heme can bind amino acids and also diatomic molecules (O-2, NO, CO). These bonds can efficiently be broken by a light pulse. Using ultrafast optical spectroscopy techniques, this property allows the study of structural and electronic dynamics in the heme and its proteic environment at femtosecond-picosecond time scales, i.e. at the time scale of internal vibrations of the macromolecular systems. This review describes recent progress in this domain, specifically the role of concerted movements of the heme-protein system in ballistic ligand transfer reactions as well as the real-time monitoring of signal propagation within signalling proteins.
  • Two photon-induced electron injection from a nanotrigger in native endothelial NO-synthase
    • Beaumont Edward
    • Lambry Jean-Christophe
    • Robin A.-C.
    • Martasek P.
    • Blanchard-Desce M.
    • Slama-Schwok Anny
    ChemPhysChem, Wiley-VCH Verlag , 2008, 9 (16), pp.2325 . We have recently designed a nanotrigger (NT), a photoactive molecule addressing the NADPH sites of proteins. This nanotrigger has a 103 times larger two-photon cross-section compared to the ubiquitous NADPH cofactor. In this work, we tested whether two-photon excitation of the bound NT to NADPH sites may be used to initiate enzymatic catalysis by appropriate electron injection. To establish proof of principle, we monitored the ultrafast absorption of NT bound to the fully active endothelial NO-Synthase (eNOS) following excitation by one and two-photons at 405 and 810 nm, respectively. Electron injection from NT* to FAD in eNOS initiated the catalytic cycle in 15±3 ps at both exciting wavelengths. The data proved for the first time that electron transfer can be promoted by two-photon excitation. We also show that the nanotrigger decays faster in homogeneous solvents than in the NADPH site of proteins, suggesting that hindered environments modified the natural decay of NT. The nanotrigger provides a convenient way of synchronizing an ensemble of proteins in solution with a femtosecond laser pulse. The ability of NT to initiate NOS catalysis by two-photon excitation may be exploited for controlled and localized release of free NO in cells with enhanced spatial and temporal resolution. Cop. 2008 Wiley-VCH Verlag GmbH & Co. KGaA. (10.1002/cphc.200800411)
    DOI : 10.1002/cphc.200800411
  • Mechanical factors activate beta-catenin-dependent oncogene expression in APC1638N/+ mouse colon
    • Whitehead J.
    • Vignjevic Danijela
    • Fütterer C.
    • Beaurepaire Emmanuel
    • Robine Sylvie
    • Farge Emmanuel
    HFSP Journal , 2008, 2 (5), pp.286 . catenin acts as a critical regulator of gastrointestinal homeostasis through its control of the Wnt signaling pathway, and genetic or epigenetic lesions which activate Wnt signaling are the primary feature of colon cancer. -catenin is also a key element of mechanotranscription pathways, leading to upregulation of master developmental gene expression during Drosophila gastrulation, or regulating mammalian bone development and maintenance. Here we investigate the impact of mechanical stimulation on the initiation of colon cancer. Myc and Twist1, two oncogenes regulated through -catenin, are expressed in response to transient compression in APC deficient "APC1638N/+... colon tissue explants, but not in wild-type colon explants. Mechanical stimulation of APC1638N/+ tissue leads to the phosphorylation of -catenin at tyrosine 654, the site of interaction with E-cadherin, as well as to increased nuclear localization of -catenin. The mechanical activation of Myc and Twist1 expression in APC1638N/+ colon can be prevented by blocking -catenin phosphorylation using Src kinase inhibitors. Microenvironmental signals are known to cooperate with genetic lesions to promote the nuclear -catenin accumulation which drives colon cancer. Here we demonstrate that when APC is limiting, mechanical strain, such as that associated with intestinal transit or tumor growth, can be interpreted by cells of preneoplastic colon tissue as a signal to initiate a -catenin dependent transcriptional program characteristic of cancer (10.2976/1.2955566)
    DOI : 10.2976/1.2955566
  • Ultrafast dynamics of ligands within heme proteins
    • Vos Marten H.
    Biochimica et Biophysica Acta (BBA) - Reviews on Bioenergetics, Elsevier , 2008, 1777, pp.15-31 . Physiological bond formation and bond breaking events between proteins and ligands and their immediate consequences are difficult to synchronize and study in general. However, diatomic ligands can be photodissociated from heme, and thus in heme proteins ligand release and rebinding dynamics and trajectories have been studied on timescales of the internal vibrations of the protein that drive many biochemical reactions, and longer. The rapidly expanding number of characterized heme proteins involved in a large variety of functions allows comparative dynamics-structure-function studies. In this review, an overview is given of recent progress in this field, and in particular on initial sensing processes in signaling proteins, and on ligand and electron transfer dynamics in oxidases and cytochromes. (10.1016/j.bbabio.2007.10.004)
    DOI : 10.1016/j.bbabio.2007.10.004
  • Conformational changes in photoexcited (R)-(+)-1,1'-bi-2-naphthol studied by time-resolved circular dichroism
    • Niezborala Claire
    • Hache François
    Journal of the American Chemical Society, American Chemical Society , 2008, 130 (38), pp.12783-12786 . Conformational changes following photoexcitation of (R)-(+)-1,1′-bi-2-naphthol are studied with a time-resolved circular dichroism (CD) experiment. Two wavelengths are investigated. For λ = 237 nm, we observe a bleaching of the ground-state absorption and a transient CD structure. Thanks to a coupled-oscillator calculation, we can attribute this effect to a decrease of the dihedral angle. For λ = 245 nm, excited-state absorption and CD are observed. All these effects are solvent-dependent. In particular, it is shown that dynamics is slower in a protic solvent, which is attributed to hydrogen-bonding of the hydroxy groups with the solvent. (10.1021/ja8039844)
    DOI : 10.1021/ja8039844
  • Flavin-dependent thymidylate synthase X limits chromosomal DNA replication
    • Escartin Frédéric
    • Skouloubris Stéphane
    • Liebl Ursula
    • Myllykallio Hannu
    Proceedings of the National Academy of Sciences of the United States of America, National Academy of Sciences , 2008, 105 (29), pp.9948-9952 .
  • Electron hopping through the 15 Å triple tryptophan molecular wire in DNA photolyase occurs within 30 ps
    • Lukacs Andras
    • Eker André P.M.
    • Byrdin Martin
    • Brettel Klaus
    • Vos Marten H.
    Journal of the American Chemical Society, American Chemical Society , 2008, 130 (44), pp.14394 . DNA photolyase is a photoactive flavoprotein that contains three tryptophan residues between the FAD cofactor and the protein surface, the solvent-exposed Trp being located 14.8 Å from the flavin. Photoreduction of the neutral radical FADH* form to the catalytically active FADH- form occurs via electron transfer through this chain. The first step in this chain takes 30 ps, the second less than 4 ps. Using a combination of site-directed mutagenesis and femtosecond polarization spectroscopy to discriminate the spectroscopically indistinguishable Trp residues, we show that the third step occurs in less than 30 ps. This implies that the first photoreduction step is rate limiting and that the Trp chain effectively acts as molecular "wire" ensuring rapid and directed long-range charge translocation across the protein. This finding is important for the functioning of the large class of cryptochrome blue-light receptors, where the Trp chain is conserved. In DNA photolyase we make use of the natural photoactivation of the process, but more generally chains of aromatic amino acids may allow very fast long-range electron transfer also in nonphotoactive proteins. Copyright Cop. 2008 American Chemical Society. (10.1021/ja805261m)
    DOI : 10.1021/ja805261m
  • Third-harmonic generation microscopy with focus-engineered beams: a numerical study
    • Olivier Nicolas
    • Beaurepaire Emmanuel
    Optics Express, Optical Society of America - OSA Publishing , 2008, 16 (19), pp.14703-14715 . We use a vector field model to analyze third-harmonic generation (THG) from model geometries (interfaces, slabs, periodic structures) illuminated by Hermite-Gaussian (HG) and Laguerre-Gaussian (LG) beams focused by a high NA lens. Calculations show that phase matching conditions are significantly affected by the tailoring of the field distribution near focus. In the case of an interface parallel to the optical axis illuminated by an odd HG mode, the emission patterns and signal level reflect the relative orientation of the interface and the focal field structure. In the case of slabs and periodic structures, the emission patterns reflect the interplay between focal field distribution (amplitude and phase) and sample structure. Forward-to-backward emission ratios using different beam shapes provide sub-wavelength information about sample spatial frequencies. © 2008 Optical Society of America (10.1364/OE.16.014703)
    DOI : 10.1364/OE.16.014703
  • Organic functionalization of luminescent oxide nanoparticles towards their application as biological probes
    • Giaume Domitille
    • Poggi Mélanie
    • Casanova Didier
    • Mialon Geneviève
    • Lahlil Khalid
    • Alexandrou Antigoni
    • Gacoin Thierry
    • Boilot Jean-Pierre
    Langmuir, American Chemical Society , 2008, 24 (19), pp.11018 . Luminescent inorganic nanoparticles are now widely studied for their applications as biological probes for in vitro or in vivo experiments. The functionalization of the particles is a key step toward these applications, since it determines the control of the coupling between the particles and the biological species of interest. This paper is devoted to the case of rare earth doped oxide nanoparticles and their functionalization through their surface encapsulation with a functional polysiloxane shell. The first step of the process is the adsorption of silicate ions that will act as a primary layer for the further surface polymerization of the silane, either aminopropyltriethoxysilane (APTES) or glycidoxypropyltrimethoxysilane (GPTMS). The amino- or epoxy- functions born by the silane allow the versatile coupling of the particles with bio-organic species following the chemistry that is commonly used in biochips. Special attention is paid to the careful characterization of each step of the functionalization process, especially concerning the average number of organic functions that are available for the final coupling of the particles with proteins. The surface density of amino or epoxy functions was found to be 0.4 and 1.9 functions per square nanometer for GPTMS and APTES silanized particles, respectively. An example of application of the amino-functionalized particles is given for the coupling with α-bungarotoxins. The average number (up to 8) and the distribution of the number of proteins per particle are given, showing the potentialities of the functionalization process for the labeling of biological species. (10.1021/la8015468)
    DOI : 10.1021/la8015468
  • Anomalies in the disappearance of the extraordinary electromagnetic transmission in subwavelength hole arrays
    • Masson Jean-Baptiste
    • Podzorov Alexander
    • Gallot Guilhem
    Optics Express, Optical Society of America - OSA Publishing , 2008, 16, pp.4719 .
  • Low-loss polymers for terahertz applications
    • Podzorov Alexander
    • Gallot Guilhem
    Applied optics, Optical Society of America , 2008, 47, pp.3254 .
  • Ligand dynamics and early signalling events in the heme domain of the sensor protein Dos from Escherichia coli
    • Yamashita Taku
    • Bouzhir-Sima Latifa
    • Lambry Jean-Christophe
    • Liebl Ursula
    • Vos Marten H.
    Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology , 2008, 283 (4), pp.2344-2352 . In the heme-based sensor Dos from Escherichia coli, the ferrous heme is coordinated by His-77 and Met-95. The latter residue is replaced upon oxygen binding or oxidation of the heme. Here we investigate the early signaling processes upon dissociation of the distal ligand using ultrafast spectroscopy and site-directed mutagenesis. Geminate CO rebinding to the heme domain DosH appears insensitive to replacement of Met-95, in agreement with the notion that this residue is oriented out of the heme pocket in the presence of external ligands. A uniquely slow 35-ps phase in rebinding of the flexible methionine side chain after dissociation from ferrous DosH is completely abolished in rebinding of the more rigid histidine side chain in the M95H mutant protein, where only the 7-ps phase, common to all 6-coordinate heme proteins, is observed. Temperature-dependence studies indicate that all rebinding of internal and external ligands is essentially barrierless, but that CfigsO escape from the heme pocket is an activated process. Solvent viscosity studies combined with molecular dynamics simulations show that there are two configurations in the ferrous 6-coordinate protein, involving two isomers of the Met-95 side chain, of which the structural changes extend to the solvent-exposed backbone, which is part of the flexible FG loop. One of these configurations has considerable motional freedom in the Met-95-dissociated state. We suggest that this configuration corresponds to an early signaling intermediate state, is responsible for the slow rebinding, and allows small ligands in the protein to efficiently compete for binding with the heme. (10.1074/jbc.M708123200)
    DOI : 10.1074/jbc.M708123200
  • Ultrafast ligand binding dynamics in the active site of native bacterial nitric oxide reductase
    • Kapetanaki Sofia
    • Field Sara J.
    • Hughes Ross
    • Watmough Nicholas
    • Liebl Ursula
    • Vos Marten H.
    Biochimica biophysica acta (BBA) - Bioenergetics, Elsevier , 2008, 1777 (7-8), pp.919-924 . The active site of nitric oxide reductase from Paracoccus denitrificans contains heme and non-heme iron and is evolutionarily related to heme-copper oxidases. The CO and NO dynamics in the active site were investigated using ultrafast transient absorption spectroscopy. We find that, upon photodissociation from the active site heme, 20% of the CO rebinds in 170 ps, suggesting that not all the CO transiently binds to the non-heme iron. The remaining 80% does not rebind within 4 ns and likely migrates out of the active site without transient binding to the non-heme iron. Rebinding of NO to ferrous heme takes place in approximately 13 ps. Our results reveal that heme-ligand recombination in this enzyme is considerably faster than in heme-copper oxidases and are consistent with a more confined configuration of the active site.
  • Characterization of mid-infrared femtosecond pulses [Invited]
    • Lee Kevin F.
    • Kubarych Kevin J.
    • Bonvalet Adeline
    • Joffre Manuel
    Journal of the Optical Society of America B, Optical Society of America , 2008, 25 (6), pp.A54 . We review different methods for characterizing mid-infrared femtosecond pulses, including linear methods such as electro-optic sampling, time-and frequency-domain interferometry and nonlinear self-referenced methods such as frequency-resolved optical gating (FROG) and spectral phase interferometry for direct electric-field reconstruction (SPIDER). Of particular interest are methods based on upconversion through non-linear mixing with chirped 800 nm pulses, enabling a complete measurement of mid-infrared pulses with visible-light spectrometers. Cop. 2008 Optical Society of America. (10.1364/JOSAB.25.000A54)
    DOI : 10.1364/JOSAB.25.000A54
  • Are cyanobacteria involved in Ciguatera Fish Poisoning-like outbreaks in New-Caledonia?
    • Laurent Dominique
    • Kerbrat A.S.
    • Daruis H.T.
    • Girard Emmanuelle
    • Golubic S.
    • Benoit Evelyne
    • Sauviat Martin-Pierre
    • Chinain M.
    • Molgó Jordi
    • Pauillac S.
    Harmful Algae, Elsevier , 2008, 7 (6), pp.827-838 . From 2001 to 2005, numerous cases of seafood poisonings were reported in a tribe from Lifou (Loyalty Islands Province, New Caledonia) of which 35 were thoroughly examined. Observations outlined by the epidemiological and clinical data (including severity and rapid onset of certain symptoms following consumption of either giant clams (Tridacna spp.) or grazing and molluscivorous fish together with the apparent inefficacy of traditional remedies, were not in favour of a classical Ciguatera Fish Poisoning (CFP) outbreak. From 2005 onwards, an environmental offshore survey of the affected area was conducted. Screening of the damaged coral area revealed the presence of large populations of cyanobacteria identified as Hydrocoleum Kützing, but the absence of Gambierdiscus spp., the well-known dinoflagellate causative agent of CFP. In vivo and in vitro toxicological studies of extracts obtained from cyanobacteria and giant clams, strongly suggested the co-occurrence of ciguatoxin-like, anatoxin-like and paralytic shellfish toxins in these samples. These new findings shed new light on the complexity of the CFP symptomatology and treatment and also on the diversity and origin of the CFP toxins. Furthermore they provide new evidence of the overall variability of seafood poisonings following the ingestion of different sea products living in a marine environment where significant harmful populations of microalgae and cyanobacteria coexist. This is the first report on the involvement of cyanobacteria in CFP-like outbreaks following the consumption of giant clams or fish specimens. Consequently, it is recommended that CFP risk assessment programs now include monitoring of cyanobacteria besides the obvious screening of CFP-promoting dinoflagellates (10.1016/j.hal.2008.04.005)
    DOI : 10.1016/j.hal.2008.04.005
  • Flavin-dependent thymidylate synthase ThyX activity: implications for the folate cycle in bacteria.
    • Leduc Damien
    • Escartin Frédéric
    • Nijhout H Frederik
    • Reed Michael C
    • Liebl Ursula
    • Skouloubris Stéphane
    • Myllykallio Hannu
    Journal of Bacteriology, American Society for Microbiology , 2007, 189 (23), pp.8537-45 . Although flavin-dependent ThyX proteins show thymidylate synthase activity in vitro and functionally complement thyA defects in heterologous systems, direct proof of their cellular functions is missing. Using insertional mutagenesis of Rhodobacter capsulatus thyX, we constructed the first defined thyX inactivation mutant. Phenotypic analyses of the obtained mutant strain confirmed that R. capsulatus ThyX is required for de novo thymidylate synthesis. Full complementation of the R. capsulatus thyX::spec strain to thymidine prototrophy required not only the canonical thymidylate synthase ThyA but also the dihydrofolate reductase FolA. Strikingly, we also found that addition of exogenous methylenetetrahydrofolate transiently inhibited the growth of the different Rhodobacter strains used in this work. To rationalize these experimental results, we used a mathematical model of bacterial folate metabolism. This model suggests that a very low dihydrofolate reductase activity is enough to rescue significant thymidylate synthesis in the presence of ThyX proteins and is in agreement with the notion that intracellular accumulation of folates results in growth inhibition. In addition, our observations suggest that the presence of flavin-dependent thymidylate synthase X provides growth benefits under conditions in which the level of reduced folate derivatives is compromised. (10.1128/JB.01380-07)
    DOI : 10.1128/JB.01380-07
  • Signal epidetection in third-harmonic generation microscopy of turbid media
    • Débarre Delphine
    • Olivier Nicolas
    • Beaurepaire Emmanuel
    Optics Express, Optical Society of America - OSA Publishing , 2007, 15, pp.8913-8924 . Third-harmonic generation (THG) imaging of thick samples or large organisms requires TH light to be epicollected through the focusing objective. In this study we first estimate the amount of backward-to-forward TH radiation created by an isolated object as a function of size and spatial frequencies in the object. Theory and model experiments indicate that no significant signal can be epidetected from a (biological) dielectric structure embedded in a transparent medium. In contrast, backward emission is observed from metal nanoparticles where THG is partly a surface effect. We then address the case of an object embedded in a turbid medium. Experiments and Monte Carlo simulations show that epidetection is possible when the absorption mean free path of harmonic light in the medium exceeds its reduced scattering length, and that epicollection efficiency critically depends on the microscope field-of-view even at shallow depths, because backscattered light is essentially diffusive. These observations provide guidelines for optimizing epidetection in third-harmonic, second-harmonic, or CARS imaging of thick tissues. (10.1364/OE.15.008913)
    DOI : 10.1364/OE.15.008913
  • A NADPH substitute for selective photo-initiation of reductive bioprocesses via two-photon induced electron transfer.
    • Robin Anne-Claire
    • Gmouh Saïd
    • Mongin Olivier
    • Jouikov Viatcheslav
    • Werts Martinus
    • Gautier Clément
    • Slama-Schwok Anny
    • Blanchard Desce Mireille
    Chemical Communications, Royal Society of Chemistry , 2007 (13), pp.1334-6 . A NADPH substitute where the nicotinamide moiety is replaced by a chromophoric unit having much larger two-photon absorption cross-section and able to transfer electrons to flavins only upon excitation is described as an effective two-photon nanotrigger for selective photo-activation of electron transfer in bioreductive processes. (10.1039/b615628b)
    DOI : 10.1039/b615628b
  • Second harmonic imaging and scoring of collagen in fibrotic tissues.
    • Strupler Mathias
    • Pena Ana-Maria
    • Hernest Monica
    • Tharaux Pierre-Louis
    • Martin Jean-Louis
    • Beaurepaire Emmanuel
    • Schanne-Klein Marie-Claire
    Optics Express, Optical Society of America - OSA Publishing , 2007, 15 (7), pp.4054-4065 . We compare second harmonic generation (SHG) to histological and immunohistochemical techniques for the visualization and scoring of collagen in biological tissues. We show that SHG microscopy is highly specific for fibrillar collagens and that combined SHG and two-photon excited fluorescence (2PEF) imaging can provide simultaneous three-dimensional visualization of collagen synthesis and assembly sites in transgenic animal models expressing GFP constructs. Finally, we propose several scores for characterizing collagen accumulation based on SHG images and appropriate for different types of collagen distributions. We illustrate the sensitivity of these scores in a murine model of renal fibrosis using a morphological segmentation of the tissue based on endogenous 2PEF signals. © 2007 Optical Society of America (10.1364/OE.15.004054)
    DOI : 10.1364/OE.15.004054
  • Synchronous photoinitiation of endothelial NO synthase activity by a nanotrigger targeted as its NADPH site
    • Beaumont Edward
    • Lambry Jean-Christophe
    • Gautier Clément
    • Robin Anne-Claire
    • Gmouh Saïd
    • Berka Vladimir
    • Tsai Ah-Lim
    • Blanchard Desce Mireille
    • Slama-Schwok Anny
    Journal of the American Chemical Society, American Chemical Society , 2007, 129 (7), pp.2178-2186 . We designed a new nanotrigger to synchronize and monitor an enzymatic activity interacting specifically with the conserved NADPH binding site. The nanotrigger (NT) combines a docking moiety targeting the NADPH site and a chromophore moiety responsive to light excitation for efficient electron transfer to the protein. Specific binding of the nanotrigger to the reductase domain of the endothelial nitric oxide synthase (eNOSred) was demonstrated by competition between NADPH and the nanotrigger on the reduction of eNOSred flavin. A micromolar Ki was estimated. We had monitored initiation of eNOSred activity by ultrafast transient spectroscopy. The transient absorption spectrum recorded at 250 ps fits the expected sum of the reduced and oxidized species, independently obtained by other chemical methods, in agreement with a photoinduced electron transfer from the excited nanotrigger to the flavin moiety of eNOSred. The rate of electron transfer from the excited state of the nanotrigger (NT*) to the protein is estimated to be kET = (7 ± 2) × 109 s-1 using the decay of oxidized eNOSred-bound nanotrigger compared against prereduced eNOSred or glucose 6-P dehydrogenase as controls. This fast electron transfer bypasses the slow hydride transfer to initiate NOS catalysis as shown by ultrafast kinetics using the eNOSred mutated in the regulatory F1160 residue. The selective targeting of the nanotrigger to NADPH sites should allow controlled initiation of the enzymatic activity of numerous proteins containing an NADPH site. (10.1021/ja067543e)
    DOI : 10.1021/ja067543e
  • Quantitative characterization of biological liquids for third-harmonic generation microscopy.
    • Débarre Delphine
    • Beaurepaire Emmanuel
    Biophysical Journal, Biophysical Society , 2007, 92 (2), pp.603-12 . Third-harmonic generation (THG) microscopy provides images of unstained biological samples based on spatial variations in third-order nonlinear susceptibility, refractive index, and dispersion. In this study, we establish quantitative values for the third-order nonlinear susceptibilities of several solvents (water, ethanol, glycerol), physiological aqueous (ions, amino acids, polypeptides, bovine serum albumin, glucose) and lipid (triglycerides, cholesterol) solutions as a function of solute concentration in the 1.05-1.25 microm excitation range. We use these data in conjunction with imaging experiments to show that THG imaging with approximately 1.2 microm excitation lacks specificity and sensitivity to detect physiological ion concentration changes, and that nonaqueous structures such as lipid bodies provide a more robust source of signal. Finally, we illustrate the impact of index-matching liquids in THG images. These data provide a basis for interpreting biological THG images and for developing additional applications. (10.1529/biophysj.106.094946)
    DOI : 10.1529/biophysj.106.094946
  • Nitric oxide synthase reduces nitrite to NO under anoxia
    • Vanin Anatoly
    • Bevers L.
    • Slama-Schwok Anny
    • van Fassen Ernst
    Cellular and Molecular Life Sciences, Springer Verlag , 2007, 64 (1), pp.96-103 . Cultured bEND.3 endothelial cells show a marked increase in NO production when subjected to anoxia, even though the normal arginine pathway of NO formation is blocked due to absence of oxygen. The rate of anoxic NO production exceeds basal unstimulated NO synthesis in normoxic cells. The anoxic release of NO is mediated by endothelial nitric oxide synthase (eNOS), can be abolished by inhibitors of NOS and is accompanied by consumption of intracellular nitrite. The anoxic NO release is unaffected by the xanthine oxidase inhibitor oxypurinol. The phenomenon is attributed to anoxic reduction of intracellular nitrite by eNOS, and its magnitude and duration suggests that the nitrite reductase activity of eNOS is relevant for fast NO delivery in hypoxic vascular tissues. (10.1007/s00018-006-6374-2)
    DOI : 10.1007/s00018-006-6374-2
  • Why does curly hair get less tangled than straight hair?
    • Masson Jean-Baptiste
    American Journal of Physics, American Association of Physics Teachers , 2007, 75 (8), pp.701 . We address the question of hair tangles and show experimentally that curly hair tends to become less tangled than straight hair. A statistical model based on geometry confirms our findings. The model gives an interesting geometric approach to hair behavior. Cop. 2007 American Association of Physics Teachers. (10.1119/1.2733683)
    DOI : 10.1119/1.2733683
  • Alterations of offspring heart muscle electrical activity transferred by rat male genitors chronically treated with lindane (7-hexachlorocyclohexane) trace concentrations
    • Sauviat Martin-Pierre
    • Godeau Gaston
    • Pages Nicole
    Pesticide Biochemistry and Physiology, Elsevier , 2007, 87 (2), pp.131 . Using intracellular microelectrodes, we Studied transmembrane resting and action potentials (AP) of left ventricle papillary muscles isolated from the heart of adult lindane-treated (TMG) and untreated (UMG) male genitor rat offspring, obtained by mating untreated female with males chronically treated and untreated with lindane (2 ppb) trace concentrations through beverage. The AP magnitude and duration (APD) were similar in both groups and their response to low temperature (22 degrees C) unchanged. Lowering the external Ca2+ concentration from 2.5 to 0.625 mM prolonged APD in the TMG group but not in the UMG group. In the TMG group, (i) cumulative addition of Sr2+ (1 mM) to the physiological Solution prolonged APD; (ii) apamin (4 mu M) and charybdotoxin (4 mu M) prolonged the APD. In conclusion, our data revealed that an altered sensitivity of the Ca2+-induced Ca2+ inactivation of L-type Ca2+ channels and of Ca2+-activated K+ channels to Ca2+ has been transferred to TMG offspring. (cop) 2006 Elsevier Inc. All rights reserved. (10.1016/j.pestbp.2006.07.002)
    DOI : 10.1016/j.pestbp.2006.07.002