Publications

2012

• Observing the Confinement Potential of Bacterial Pore-Forming Toxin Receptors Inside Rafts with Nonblinking Eu3+-Doped Oxide Nanoparticles
  
  • Türkcan Silvan
  • Masson Jean-Baptiste
  • Casanova Didier
  • Mialon Geneviève
  • Gacoin Thierry
  • Boilot Jean-Pierre
  • Popoff Michel R.
  Biophysical Journal, Biophysical Society, 2012, 102 (10), pp.2299-2308. We track single toxin receptors on the apical cell membrane of MDCK cells with Eu-doped oxide nanoparticles coupled to two toxins of the pore-forming toxin family: α-toxin of Clostridium septicum and -toxin of Clostridium perfringens. These nonblinking and photostable labels do not perturb the motion of the toxin receptors and yield long uninterrupted trajectories with mean localization precision of 30 nm for acquisition times of 51.3 ms. We were thus able to study the toxin-cell interaction at the single-molecule level. Toxins bind to receptors that are confined within zones of mean area 0.40 0.05 m2. Assuming that the receptors move according to the Langevin equation of motion and using Bayesian inference, we determined mean diffusion coefficients of 0.16 0.01 m2/s for both toxin receptors. Moreover, application of this approach revealed a force field within the domain generated by a springlike confining potential. Both toxin receptors were found to experience forces characterized by a mean spring constant of 0.30 0.03 pN/m at 37C. Furthermore, both toxin receptors showed similar distributions of diffusion coefficient, domain area, and spring constant. Control experiments before and after incubation with cholesterol oxidase and sphingomyelinase show that these two enzymes disrupt the confinement domains and lead to quasi-free motion of the toxin receptors. Our control data showing cholesterol and sphingomyelin dependence as well as independence of actin depolymerization and microtubule disruption lead us to attribute the confinement of both receptors to lipid rafts. These toxins require oligomerization to develop their toxic activity. The confined nature of the toxin receptors leads to a local enhancement of the toxin monomer concentration and may thus explain the virulence of this toxin family. (10.1016/j.bpj.2012.03.072)
  DOI : 10.1016/j.bpj.2012.03.072
• Genomic binding of Pol III transcription machinery and relationship with TFIIS transcription factor distribution in mouse embryonic stem cells
  
  • Carrière Lucie
  • Graziani Sébastien
  • Alibert Olivier
  • Ghavi-Helm Yad
  • Boussouar Fayçal
  • Humbertclaude Hélène
  • Jounier Sylvie
  • Aude Jean-Christophe
  • Keime Celine
  • Murvai Janos
  • Foglio Mario
  • Gut Marta
  • Gut Ivo
  • Lathrop Mark
  • Soutourina Julie
  • Gérard Matthieu
  • Werner Michel
  Nucleic Acids Research, Oxford University Press, 2012, 40 (1), pp.270-283. (10.1093/nar/gkr737)
  DOI : 10.1093/nar/gkr737
• Photoinduced dynamics of oxyluciferin analogues: Unusual enol "super"photoacidity and evidence for keto-enol isomerization
  
  • Solntsev K.M.
  • Laptenok Sergey P.
  • Naumov P.
  Journal of the American Chemical Society, American Chemical Society, 2012, 134 (40), pp.16452-16455. The first systematic pico-nanosecond time-resolved spectroscopic study of the firefly emitter oxyluciferin and two of its chemically modified analogues revealed that in the excited state the enol group is more acidic than the phenol group. The 6'-dehydroxylated derivative, in which only the 4-enolic hydroxyl proton is acidic, has an experimentally determined pK a* of 0.9 in dimethyl sulfoxide and an estimated pK a* of -0.3 in water. Moreover, this compound provided direct evidence that in a nonpolar, basic environment the keto form in the excited state can tautomerize into the enol, which subsequently undergoes excited-state proton transfer (ESPT) to produce enolate ion. This observation presents the first experimental evidence of excited-state keto-enol tautomerization of a firefly fluorophore, and it could be important in resolving the enol-keto conundrum related to the color-tuning mechanism of firefly bioluminescence. The 6'-dehydroxylated form of oxyluciferin adds a very rare case of a stable enol to the family of "super"photoacids. Cop. 2012 American Chemical Society. (10.1021/ja3045212)
  DOI : 10.1021/ja3045212
• Adaptive Optics for Biomedical Microscopy
  
  • Booth Martin J.
  • Débarre Delphine
  • Jesacher Alexander
  Optics and photonics news, Optical Society of America - OSA Publishing, 2012, 23 (1), pp.22-29. Over the last decade, researchers have applied adaptive optics--a technology that was originally conceived for telescopes--to high-resolution microscopy in order to overcome the problems caused by specimen-induced aberrations. © 2012 Optical Society of America (10.1364/OPN.23.1.000022)
  DOI : 10.1364/OPN.23.1.000022
• Mechanistic and structural basis for inhibition of thymidylate synthase ThyX
  
  • Basta Tamara
  • Boum Yap
  • Briffotaux Julien
  • Becker Hubert F.
  • Lamarre-Jouenne Isabelle
  • Lambry Jean-Christophe
  • Skouloubris Stéphane
  • Liebl Ursula
  • Graille Marc
  • van Tilbeurgh Herman
  • Myllykallio Hannu
  Open Biology, Royal Society, 2012, 2 (-). Nature has established two mechanistically and structurally unrelated families of thymidylate synthases that produce de novo thymidylate or dTMP, an essential DNA precursor. Representatives of the alternative flavin-dependent thymidylate synthase family, ThyX, are found in a large number of microbial genomes, but are absent in humans. We have exploited the nucleotide binding pocket of ThyX proteins to identify non-substrate-based tight-binding ThyX inhibitors that inhibited growth of genetically modified Escherichia coli cells dependent on thyX in a manner mimicking a genetic knockout of thymidylate synthase. We also solved the crystal structure of a viral ThyX bound to 2-hydroxy-3-(4-methoxybenzyl)-1,4-naphthoquinone at a resolution of 2.6 angstrom. This inhibitor was found to bind within the conserved active site of the tetrameric ThyX enzyme, at the interface of two monomers, partially overlapping with the dUMP binding pocket. Our studies provide new chemical tools for investigating the ThyX reaction mechanism and establish a novel mechanistic and structural basis for inhibition of thymidylate synthesis. As essential ThyX proteins are found e. g. in Mycobacterium tuberculosis and Helicobacter pylori, our studies have also potential to pave the way towards the development of new anti-microbial compounds. (10.1098/rsob.120120)
  DOI : 10.1098/rsob.120120
• Mechanistic and structural basis for inhibition of thymidylate synthase ThyX
  
  • Basta Tamara
  • Boum Yap
  • Briffotaux Julien
  • Becker Hubert F
  • Lamarre-Jouenne Isabelle
  • Lambry Jean-Christophe
  • Skouloubris Stephane
  • Liebl Ursula
  • Graille Marc
  • van Tilbeurgh Herman
  • Myllykallio Hannu
  Open Biology, Royal Society, 2012, 2 (10), pp.120120. Nature has established two mechanistically and structurally unrelated families of thymidylate synthases that produce de novo thymidylate or dTMP, an essential DNA precursor. Representatives of the alternative flavin-dependent thymidylate synthase family, ThyX, are found in a large number of microbial genomes, but are absent in humans. We have exploited the nucleotide binding pocket of ThyX proteins to identify non-substrate-based tight-binding ThyX inhibitors that inhibited growth of genetically modified Escherichia coli cells dependent on thyX in a manner mimicking a genetic knockout of thymidylate synthase. We also solved the crystal structure of a viral ThyX bound to 2-hydroxy-3-(4-methoxybenzyl)-1,4-naphthoquinone at a resolution of 2.6 Å. This inhibitor was found to bind within the conserved active site of the tetrameric ThyX enzyme, at the interface of two monomers, partially overlapping with the dUMP binding pocket. Our studies provide new chemical tools for investigating the ThyX reaction mechanism and establish a novel mechanistic and structural basis for inhibition of thymidylate synthesis. As essential ThyX proteins are found e.g. in Mycobacterium tuberculosis and Helicobacter pylori, our studies have also potential to pave the way towards the development of new anti-microbial compounds. (10.1098/rsob.120120)
  DOI : 10.1098/rsob.120120
• A Bayesian Inference Scheme to Extract Diffusivity and Potential Fields from Confined Single-Molecule Trajectories
  
  • Tuerkcan Silvan
  • Alexandrou Antigoni
  • Masson Jean-Baptiste
  Biophysical Journal, Biophysical Society, 2012, 102 (10), pp.2288-2298. Currently used techniques for the analysis of single-molecule trajectories only exploit a small part of the available information stored in the data. Here, we apply a Bayesian inference scheme to trajectories of confined receptors that are targeted by pore-forming toxins to extract the two-dimensional confining potential that restricts the motion of the receptor. The receptor motion is modeled by the overdamped Langevin equation of motion. The method uses most of the information stored in the trajectory and converges quickly onto inferred values, while providing the uncertainty on the determined values. The inference is performed on the polynomial development of the potential and on the diffusivities that have been discretized on a mesh. Numerical simulations are used to test the scheme and quantify the convergence toward the input values for forces, potential, and diffusivity. Furthermore, we show that the technique outperforms the classical mean-square-displacement technique when forces act on confined molecules because the typical mean-square-displacement analysis does not account for them. We also show that the inferred potential better represents input potentials than the potential extracted from the position distribution based on Boltzmann statistics that assumes statistical equilibrium. (10.1016/j.bpj.2012.01.063)
  DOI : 10.1016/j.bpj.2012.01.063
• Nitric oxide binding to the cardiolipin complex of ferric cytochrome
  
  • Silkstone G.
  • Kapetanaki Sofia M.
  • Husu I.
  • Vos Marten H.
  • Wilson M.T.
  Biochemistry, American Chemical Society, 2012, 51 (34), pp.6760-6766. Cardiolipin, a phospholipid specific to the mitochondrion, interacts with the small electron transfer heme protein cytochrome c through both electrostatic and hydrophobic interactions. Once in a complex with cardiolipin, cytochrome c has been shown to undergo a conformational change that leads to the rupture of the bond between the heme iron and the intrinsic sulfur ligand of a methionine residue and to enhance the peroxidatic properties of the protein considered important to its apoptotic activity. Here we report that the ferric cytochrome c/cardiolipin complex binds nitric oxide tightly through a multistep process in which the first step is the relatively slow displacement (5 s-1) from heme coordination of an intrinsic ligand that replaces methionine in the complex. Nanosecond photolysis of the nitrosyl adduct demonstrated that a fraction of the nitric oxide escapes from the heme pocket and subsequently recombines to the heme in second-order processes (k = 1.8 × 106 and 5.5 × 105 M-1 s-1) that, under these conditions, were much faster than recombination of the intrinsic ligand with which they compete. Ultrafast (femtosecond) laser photolysis showed that the geminate recombination of nitric oxide to the heme occurred with time constants (? = 22 and 72 ps) and that ~23% of the photolyzed nitric oxide escaped into the bulk phase. This high value for the escape fraction relative to other heme proteins indicates the open nature of the heme pocket in this complex. These results are summarized in a scheme and are discussed in terms of the possible modulation of the apoptotic activity of cytochrome c by nitric oxide. Cop. 2012 American Chemical Society. (10.1021/bi300596)
  DOI : 10.1021/bi300596
• Asynchronous optical sampling with arbitrary detuning between laser repetition rates
  
  • Antonucci Laura
  • Solinas Xavier
  • Bonvalet Adeline
  • Joffre Manuel
  Optics Express, Optical Society of America - OSA Publishing, 2012, 20 (16), pp.17928-17937. A method of asynchronous optical sampling based on free-running lasers with no requirement on the repetition rates is presented. The method is based on the a posteriori determination of the delay between each pair of pulses. A resolution better than 400 fs over 13 ns total delay scan is demonstrated. In addition to the advantages of conventional asynchronous sampling techniques, this method allows a straightforward implementation on already-existing laser systems using a fiber-based setup and an appropriate acquisition procedure. (C) 2012 Optical Society of America (10.1364/OE.20.017928)
  DOI : 10.1364/OE.20.017928
• Diffraction from a subwavelength elliptic aperture: Analytic approximate aperture fields
  
  • Masson Jean-Baptiste
  • Gallot Guilhem
  Journal of the Optical Society of America. A Optics, Image Science, and Vision, Optical Society of America, 2012, 29 (9), pp.2005-2014. An analytical approximate solution of the electromagnetic field on a subwavelength elliptical hole in a thin perfectly conducting screen is presented. Illumination is a linear polarized, normally incident plane wave. A polynomial development method is used and allows one to obtain an easy-to-use analytical solution of the fields, which can be used to build analytical expressions of aperture fields for apertures in anisotropic structures. © 2012 Optical Society of America. (10.1364/josaa.29.002005)
  DOI : 10.1364/josaa.29.002005
• Glotaran: A Java-based graphical user interface for the R package TIMP
  
  • Snellenburg Joris J.
  • Laptenok Sergey
  • Seger Ralf
  • Mullen Katharine M.
  • van Stokkum Ivo H.M.
  Journal of Statistical Software, University of California, Los Angeles, 2012, 49 (3). In this work the software application called Glotaran is introduced as a Java-based graphical user interface to the R package TIMP, a problem solving environment for fitting superposition models to multi-dimensional data. TIMP uses a command-line user interface for the interaction with data, the specification of models and viewing of analysis results. Instead, Glotaran provides a graphical user interface which features interactive and dynamic data inspection, easier -- assisted by the user interface -- model specification and interactive viewing of results. The interactivity component is especially helpful when working with large, multi-dimensional datasets as often result from time-resolved spectroscopy measurements, allowing the user to easily pre-select and manipulate data before analysis and to quickly zoom in to regions of interest in the analysis results. Glotaran has been developed on top of the NetBeans rich client platform and communicates with R through the Java-to-R interface Rserve. The background and the functionality of the application are described here. In addition, the design, development and implementation process of Glotaran is documented in a generic way.
• Amplification and temporal filtering during gradient sensing by nerve growth cones probed with a microfluidic assay
  
  • Morel Mathieu
  • Shynkar Vasyl
  • Galas Jean-Christophe
  • Dupin Isabelle
  • Bouzigues Cédric
  • Studer Vincent
  • Dahan Maxime
  Biophysical Journal, Biophysical Society, 2012, 103 (8), pp.1648-1656. Nerve growth cones (GCs) are chemical sensors that convert graded extracellular cues into oriented axonal motion. To ensure a sensitive and robust response to directional signals in complex and dynamic chemical landscapes, GCs are presumably able to amplify and filter external information. How these processing tasks are performed remains however poorly known. Here, we probe the signal-processing capabilities of single GCs during ?-Aminobutyric acid (GABA) directional sensing with a shear-free microfluidic assay that enables systematic measurements of the GC output response to variable input gradients. By measuring at the single molecule level the polarization of GABA A chemoreceptors at the GC membrane, as a function of the external GABA gradient, we find that GCs act as i), signal amplifiers over a narrow range of concentrations, and ii), low-pass temporal filters with a cutoff frequency independent of stimuli conditions. With computational modeling, we determine that these systems-level properties arise at a molecular level from the saturable occupancy response and the lateral dynamics of GABA A receptors. Cop. 2012 Biophysical Society. (10.1016/j.bpj.2012.08.040)
  DOI : 10.1016/j.bpj.2012.08.040
• Optical activity of metallic helices in the terahertz domain: A theoretical investigation
  
  • Hache François
  • Gallot Guilhem
  Journal of the Optical Society of America B, Optical Society of America, 2012, 29 (10), pp.2675-2684. Optical activity in the terahertz spectral domain has recently seen a growing interest, but fine understanding of these phenomena is not yet developed. In this article, we study analytically the response of a metallic helix in the terahertz regime and present a full nonlocal calculation of its chiroptical response. Because we do not use multipolar expansion, this calculation is very general and applies to the case where the helix size is comparable to the wavelength of the light. We calculate the circular birefringence and dichroism in three configurations: propagation along or perpendicular to the helix axis and response of an isotropic distribution of such helices. We obtain analytical expressions and can examine the consequence of the breakdown of the multipolar expansion and the wavelength-dependence of the chiroptical response, as well as give orders of magnitude that compare favorably with experimental data. This calculation is also comforted by a finite element calculation. Cop. 2012 Optical Society of America. (10.1364/JOSAB.29.002675)
  DOI : 10.1364/JOSAB.29.002675
• Ultrafast Ligand Dynamics in the Heme-Based GAF Sensor Domains of the Histidine Kinases DosS and DosT from Mycobacterium tuberculosis.
  
  • Vos Marten H.
  • Bouzhir-Sima Latifa
  • Lambry Jean-Christophe
  • Luo Hao
  • Eaton-Rye Julian J.
  • Ioanoviciu Alexandra
  • Ortiz de Montellano Paul
  • Liebl Ursula
  Biochemistry, American Chemical Society, 2012, 51 (1), pp.159-166. The transcriptional regulator DosR from M. tuberculosis plays a crucial role in the virulence to dormancy transition of the pathogen. DosR can be activated by DosT and DosS, two histidine kinases with heme-containing sensor GAF domains, capable of diatomic ligand binding. To investigate the initial processes occurring upon ligand dissociation, we performed ultrafast time-resolved absorption spectroscopy of the isolated sensor domains ligated with O2, NO, and CO. The results reveal a relatively closed heme pocket for both proteins. For DosT the yield of O2 escape from the heme pocket on the picoseconds time scale upon photodissociation was found to be very low (1.5%), similar to other heme-based oxygen sensor proteins, implying that this sensor acts as an effective O2 trap. Remarkably, this yield is an order of magnitude higher in DosS (18%). For CO, by contrast, the fraction of CO rebinding within the heme pocket is higher in DosS. Experiments with mutant DosT sensor domains and molecular dynamics simulations indicate an important role in ligand discrimination of the distal tyrosine, present in both proteins, which forms a hydrogen bond with heme-bound O2. We conclude that despite their similarity, DosT and DosS display ligand-specific different primary dynamics during the initial phases of intraprotein signaling. The distal tyrosine, present in both proteins, plays an important role in these processes. (10.1021/bi201467c)
  DOI : 10.1021/bi201467c
• Multicolor two-photon tissue imaging by wavelength mixing
  
  • Mahou Pierre
  • Zimmerley Maxwell
  • Loulier Karine
  • Matho Katherine S.
  • Labroille Guillaume
  • Morin Xavier
  • Supatto Willy
  • Livet Jean
  • Débarre Delphine
  • Beaurepaire Emmanuel
  Nature Methods, Nature Publishing Group, 2012, 9 (8), pp.815-818. We achieve simultaneous two-photon excitation of three chromophores with distinct absorption spectra using synchronized pulses from a femtosecond laser and an optical parametric oscillator. The two beams generate separate multiphoton processes, and their spatiotemporal overlap provides an additional two-photon excitation route, with submicrometer overlay of the color channels. We report volume and live multicolor imaging of 'Brainbow'-labeled tissues as well as simultaneous three-color fluorescence and third-harmonic imaging of fly embryos. (10.1038/nmeth.2098)
  DOI : 10.1038/nmeth.2098
• Transcriptional Activation and Cell Cycle Block Are the Keys for 5-Fluorouracil Induced Up-Regulation of Human Thymidylate Synthase Expression
  
  • Ligabue Alessio
  • Marverti G.
  • Liebl Ursula
  • Myllykallio Hannu
  PLoS ONE, Public Library of Science, 2012, 7 (10), pp.e47318. Background: 5-fluorouracil, a commonly used chemotherapeutic agent, up-regulates expression of human thymidylate synthase (hTS). Several different regulatory mechanisms have been proposed to mediate this up-regulation in distinct cell lines, but their specific contributions in a single cell line have not been investigated to date. We have established the relative contributions of these previously proposed regulatory mechanisms in the ovarian cancer cell line 2008 and the corresponding cisplatin-resistant and 5-FU cross-resistant-subline C13*. Methodology/Principal Findings: Using RNA polymerase II inhibitor DRB treated cell cultures, we showed that 70-80% of up-regulation of hTS results from transcriptional activation of TYMS mRNA. Moreover, we report that 5-FU compromises the cell cycle by blocking the 2008 and C13* cell lines in the S phase. As previous work has established that TYMS mRNA is synthesized in the S and G 1 phase and hTS is localized in the nuclei during S and G 2-M phase, the observed cell cycle changes are also expected to affect the intracellular regulation of hTS. Our data also suggest that the inhibition of the catalytic activity of hTS and the up-regulation of the hTS protein level are not causally linked, as the inactivated ternary complex, formed by hTS, deoxyuridine monophosphate and methylenetetrahydrofolate, was detected already 3 hours after 5-FU exposure, whereas substantial increase in global TS levels was detected only after 24 hours. Conclusions/Significance: Altogether, our data indicate that constitutive TYMS mRNA transcription, cell cycle-induced hTS regulation and hTS enzyme stability are the three key mechanisms responsible for 5-fluorouracil induced up-regulation of human thymidylate synthase expression in the two ovarian cancer cell lines studied. As these three independent regulatory phenomena occur in a precise order, our work provides a feasible rationale for earlier observed synergistic combinations of 5-FU with other drugs and may suggest novel therapeutic strategies. Cop. 2012 Ligabue et al. (10.1371/journal.pone.0047318)
  DOI : 10.1371/journal.pone.0047318
• Hyperglycemia-induced abnormalities in rat and human corneas: the potential of second harmonic generation microscopy.
  
  • Latour Gaël
  • Kowalczuk Laura
  • Savoldelli Michèle
  • Bourges Jean-Louis
  • Plamann Karsten
  • Behar-Cohen Francine
  • Schanne-Klein Marie-Claire
  PLoS ONE, Public Library of Science, 2012, 7 (11), pp.e48388. BACKGROUND: Second Harmonic Generation (SHG) microscopy recently appeared as an efficient optical imaging technique to probe unstained collagen-rich tissues like cornea. Moreover, corneal remodeling occurs in many diseases and precise characterization requires overcoming the limitations of conventional techniques. In this work, we focus on diabetes, which affects hundreds of million people worldwide and most often leads to diabetic retinopathy, with no early diagnostic tool. This study then aims to establish the potential of SHG microscopy for in situ detection and characterization of hyperglycemia-induced abnormalities in the Descemet's membrane, in the posterior cornea. METHODOLOGY/PRINCIPAL FINDINGS: We studied corneas from age-matched control and Goto-Kakizaki rats, a spontaneous model of type 2 diabetes, and corneas from human donors with type 2 diabetes and without any diabetes. SHG imaging was compared to confocal microscopy, to histology characterization using conventional staining and transmitted light microscopy and to transmission electron microscopy. SHG imaging revealed collagen deposits in the Descemet's membrane of unstained corneas in a unique way compared to these gold standard techniques in ophthalmology. It provided background-free images of the three-dimensional interwoven distribution of the collagen deposits, with improved contrast compared to confocal microscopy. It also provided structural capability in intact corneas because of its high specificity to fibrillar collagen, with substantially larger field of view than transmission electron microscopy. Moreover, in vivo SHG imaging was demonstrated in Goto-Kakizaki rats. CONCLUSIONS/SIGNIFICANCE: Our study shows unambiguously the high potential of SHG microscopy for three-dimensional characterization of structural abnormalities in unstained corneas. Furthermore, our demonstration of in vivo SHG imaging opens the way to long-term dynamical studies. This method should be easily generalized to other structural remodeling of the cornea and SHG microscopy should prove to be invaluable for in vivo corneal pathological studies. (10.1371/journal.pone.0048388)
  DOI : 10.1371/journal.pone.0048388
• Red Blood Cell Sickling During Oxygen Cycles in a Microdroplet Device
  
  • Abbyad Paul
  • Dangla Remi
  • Tharaux Pierre-Louis
  • Baroud Charles
  • Alexandrou Antigoni
  Biophysical Journal, Biophysical Society, 2012, 102 (3), pp.29A-29A. We have developed a novel microfluidic device to study repetitive sickling on individual red blood cells by replicating the physiological oxygen cycling of the vascular circulatory system (Abbyad et al., Lab Chip, 2011, 11, 813). A small number of red blood cells from sickle cell patients are encapsulated in an array of aqueous microdroplets. These microdroplets are anchored and arranged in a 2-dimensional array against the flow of the carrier oil. Precise spatial and temporal changes in oxygen concentration are obtained through gas exchange with the inert oil flowing outside the droplets. (10.1016/j.bpj.2011.11.184)
  DOI : 10.1016/j.bpj.2011.11.184
• Absorption band III kinetics probe the picosecond heme iron motion triggered by nitric oxide binding to hemoglobin and myoglobin.
  
  • Yoo Byung-Kuk
  • Kruglik Sergei G.
  • Lamarre Isabelle
  • Martin Jean-Louis
  • Negrerie Michel
  Journal of Physical Chemistry B, American Chemical Society, 2012, 116 (13), pp.4106-4114. To study the ultrafast movement of the heme iron induced by nitric oxide (NO) binding to hemoglobin (Hb) and myoglobin (Mb), we probed the picosecond spectral evolution of absorption band III (∼760 nm) and vibrational modes (iron-histidine stretching, ν(4) and ν(7) in-plane modes) in time-resolved resonance Raman spectra. The time constants of band III intensity kinetics induced by NO rebinding (25 ps for hemoglobin and 40 ps for myoglobin) are larger than in Soret bands and Q-bands. Band III intensity kinetics is retarded with respect to NO rebinding to Hb and to Mb. Similarly, the ν((Fe-His)) stretching intensity kinetics are retarded with respect to the ν(4) and ν(7) heme modes and to Soret absorption. In contrast, band III spectral shift kinetics do not coincide with band III intensity kinetics but follows Soret kinetics. We concluded that, namely, the band III intensity depends on the heme iron out-of-plane position, as theoretically predicted ( Stavrov , S. S. Biopolymers 2004 , 74 , 37 - 40 ). (10.1021/jp300849y)
  DOI : 10.1021/jp300849y
• Systèmes Femtoseconde : optique et phénomènes ultrarapides
  
  • Mottin Stephane
  • Monmayrant Antoine
  • Ruchon Thierry
  • Bonvalet Adeline
  • Lee Kevin
  • Joffre Manuel
  • Chiche Ronic
  • Jehanno Didier
  • Picqué Nathalie
  • Soskov Viktor
  • Variola Allessandro
  • Zomer Fabian
  • Vauthey Eric
  • Vallée Fabrice
  • del Fatti Natalia
  • Marie Xavier
  • Urbaszek Bernhard
  • Renucci Pierre
  • Gilliot Pierre
  • Druon Frédéric
  • Amand Thierry
  , 2012. Sommaire : P. 9 Préface / Béatrice Chatel P. 11 Introduction / Pierre Gilliot P. 13 Production d'impulsions femtoseconde et nouveaux matériaux pour le pompage par diode / Frédéric Druon. P. 51 Façonnage et caractérisation d‟impulsions ultra-rapides / Antoine Monmayrant. P. 81 Introduction to the synthesis and the measurement of attosecond pulses / Thierry Ruchon. P. 109 Impulsions infrarouges femtosecondes : génération, caractérisation et applications / Adeline Bonvalet, Kevin F Lee, Manuel Joffre. P. 119 Les Cavités Fabry-Perot en mode pulsé et leurs récentes applications / Ronic Chiche, Didier Jehanno, Nathalie Picqué, Viktor Soskov, Allessandro Variola, Fabian Zomer. P. 171 Les réactions de transfert d'électron photoinduites étudiées par spectroscopie ultrarapide / Eric Vauthey. P. 185 Ultrafast spectroscopy of a single metal nanoparticle / Fabrice Vallée, Natalia Del Fatti. P. 209 Time-resolved optical measurements of the spin coherence in semiconductor nanostructures / Xavier Marie, Bernhard Urbaszek, Pierre Renucci, Thierry Amand.
• In vivo structural imaging of the cornea by polarization-resolved second harmonic microscopy.
  
  • Latour Gaël
  • Gusachenko Ivan
  • Kowalczuk Laura
  • Lamarre Isabelle
  • Schanne-Klein Marie-Claire
  Biomedical optics express, Optical Society of America - OSA Publishing, 2012, 3 (1), pp.1-15. The transparency and mechanical strength of the cornea are related to the highly organized three-dimensional distribution of collagen fibrils. It is of great interest to develop specific and contrasted in vivo imaging tools to probe these collagenous structures, which is not available yet. Second Harmonic Generation (SHG) microscopy is a unique tool to reveal fibrillar collagen within unstained tissues, but backward SHG images of cornea fail to reveal any spatial features due to the nanometric diameter of stromal collagen fibrils. To overcome this limitation, we performed polarization-resolved SHG imaging, which is highly sensitive to the sub-micrometer distribution of anisotropic structures. Using advanced data processing, we successfully retrieved the orientation of the collagenous fibrils at each depth of human corneas, even in backward SHG homogenous images. Quantitative information was also obtained about the submicrometer heterogeneities of the fibrillar collagen distribution by measuring the SHG anisotropy. All these results were consistent with numerical simulation of the polarization-resolved SHG response of cornea. Finally, we performed in vivo SHG imaging of rat corneas and achieved structural imaging of corneal stroma without any labeling. Epi-detected polarization-resolved SHG imaging should extend to other organs and become a new diagnosis tool for collagen remodeling. 2011 Optical Society of America (10.1364/BOE.3.000001)
  DOI : 10.1364/BOE.3.000001