Publications

2011

• Sub-picosecond Raman spectrometer for time-resolved studies of structural dynamics in heme proteins
  
  • Kruglik Sergei G.
  • Lambry Jean-Christophe
  • Martin Jean-Louis
  • Vos Marten H.
  • Négrerie Michel
  Journal of Raman Spectroscopy, Wiley, 2011, 42 (3), pp.265. We describe a pump-probe Raman spectrometer based on a femtosecond Ti:sapphire laser, an optical parametric generator and two optical parametric amplifiers for time-resolved studies, with emphasis on the structural dynamics in heme proteins. The system provides a 100-fs pump pulse tunable in the range 500-600 nm and a transform-limited sub-picosecond probe pulse tunable in the range 390-450 nm. The spectrometer has spectral (25 cm(-1)) and temporal (similar to 0.7 ps) resolutions which constitute an effective compromise for identifying transient heme protein species and for following their structural evolution by spontaneous Raman scattering in the time range 0.5 ps to 2 ns. This apparatus was applied to time-resolved studies of a broad range of heme proteins, monitoring the primary dynamics of photoinduced heme coordination state and structural changes, its interaction with protein side-chains and diatomic gaseous ligands, as well as heme vibrational cooling. The treatment of transient Raman spectra is described in detail, and the advantages and shortcomings of spontaneous resonance Raman spectroscopy for ultrafast heme proteins studies are discussed. We demonstrate the efficiency of the constructed spectrometer by measuring Raman spectra in the sub-picosecond and picosecond time ranges for the oxygen-storage heme protein myoglobin and for the oxygen-sensor heme protein FixLH in interaction with the diatomic gaseous ligands CO, NO, and O-2. Copyright (C) 2010 John Wiley and Sons, Ltd. (10.1002/jrs.2685)
  DOI : 10.1002/jrs.2685
• Deep and fast live imaging with two-photon scanned light-sheet microscopy
  
  • Truong T.V.
  • Supatto Willy
  • Koos D.S.
  • Choi J.M.
  • Fraser S.E.
  Nature Methods, Nature Publishing Group, 2011, 8 (9), pp.757. We implemented two-photon scanned light-sheet microscopy, combining nonlinear excitation with orthogonal illumination of light-sheet microscopy, and showed its excellent performance for in vivo, cellular-resolution, three-dimensional imaging of large biological samples. Live imaging of fruit fly and zebrafish embryos confirmed that the technique can be used to image up to twice deeper than with one-photon light-sheet microscopy and more than ten times faster than with point-scanning two-photon microscopy without compromising normal biology. Cop. 2011 Nature America, Inc. All rights reserved. (10.1038/nmeth.1652)
  DOI : 10.1038/nmeth.1652
• Dynamics of NO interacting with soluble guanylate cyclase from 1 ps to 0.1 s and induced structural transitions
  
  • Yoo Byung-Kuk
  • Lamarre Isabelle
  • Martin Jean-Louis
  • Rappaport Fabrice
  • Negrerie Michel
  BMC Pharmacology, BioMed Central, 2011, 11 (Suppl 1), pp.P77.
• Vibrational Motions Associated with Primary Processes in Bacteriorhodopsin Studied by Coherent Infrared Emission Spectroscopy
  
  • Groma Geza I.
  • Colonna Anne
  • Martin Jean-Louis
  • Vos Marten H.
  Biophysical Journal, Biophysical Society, 2011, 100 (6), pp.1578. The primary energetic processes driving the functional proton pump of bacteriorhodopsin take place in the form of complex molecular dynamic events after excitation of the retinal chromophore into the Franck-Condon state. These early events include a strong electronic polarization, skeletal stretching, and all-trans-to-13-cis isomerization upon formation of the J intermediate. The effectiveness of the photoreaction is ensured by a conical intersection between the electronic excited and ground states, providing highly nonadiabatic coupling to nuclear motions. Here, we study real-time vibrational coherences associated with these motions by analyzing light-induced infrared emission from oriented purple membranes in the 750-1400 cm(-1) region. The experimental technique applied is based on second-order femtosecond difference frequency generation on macroscopically ordered samples that also yield information on phase and direction of the underlying motions. Concerted use of several analysis methods resulted in the isolation and characterization of seven different vibrational modes assigned as C-C stretches, out-of-plane methyl rocks, and hydrogen out-of-plane wags, whereas no in-plane H rock was found. Based on their lifetimes and several other criteria, we deduce that the majority of the observed modes take place on the potential energy surface of the excited electronic state. In particular, the direction sensitivity provides experimental evidence for large intermediate distortions of the retinal plane during the excited-state isomerization process. (10.1016/j.bpj.2011.02.011)
  DOI : 10.1016/j.bpj.2011.02.011
• Combining rails and anchors with laser forcing for selective manipulation within 2D droplet arrays.
  
  • Fradet Etienne
  • Mcdougall Craig
  • Abbyad Paul
  • Dangla Rémi
  • Mcgloin David
  • Baroud Charles N.
  Lab on a Chip, Royal Society of Chemistry, 2011, 11 (24), pp.4228-4234. We demonstrate the combination of a rails and anchors microfluidic system with laser forcing to enable the creation of highly controllable 2D droplet arrays. Water droplets residing in an oil phase can be pinned to anchor holes made in the base of a microfluidic channel, enabling the creation of arrays by the appropriate patterning of such holes. The introduction of laser forcing, via laser induced thermocapillary forces to anchored droplets, enables the selective extraction of particular droplets from an array. We also demonstrate that such anchor arrays can be filled with multiple, in our case two, droplets each and that if such droplets have different chemical contents, the application of a laser at their interface triggers their merging and a chemical reaction to take place. Finally by adding guiding rails within the microfluidic structure we can selectively fill large scale arrays with monodisperse droplets with significant control over their contents. In this way we make a droplet array filled with 96 droplets containing different concentrations of fluorescent microparticles. (10.1039/C1LC20541B)
  DOI : 10.1039/C1LC20541B
• From Cilia Hydrodynamics to Zebrafish Embryonic Development
  
  • Supatto Willy
  • Vermot Julien
  , 2011, pp.vol. 95, 33-66. Embryonic development involves the cellular integration of chemical and physical stimuli. A key physical input is the mechanical stress generated during embryonic morphogenesis. This process necessitates tensile forces at the tissue scale such as during axis elongation and budding, as well as at the cellular scale when cells migrate and contract. Furthermore, cells can generate forces using motile cilia to produce flow. Cilia-driven flows are critical throughout embryonic development but little is known about the diversity of the forces they exert and the role of the mechanical stresses they generate. In this chapter, through an examination of zebrafish development, we highlight what is known about the role of hydrodynamics mediated by beating cilia and examine the physical features of flow fields from the modeling and experimental perspectives. We review imaging strategies to visualize and quantify beating cilia and the flow they generate in vivo. Finally, we describe the function of hydrodynamics during left-right embryonic patterning and inner ear development. Ideally, continued progress in these areas will help to address a key conceptual problem in developmental biology, which is to understand the interplay between environmental constraints and genetic control during morphogenesis (10.1016/B978-0-12-385065-2.00002-5)
  DOI : 10.1016/B978-0-12-385065-2.00002-5
• Strong Ligand-Protein Interactions Revealed by Ultrafast Infrared Spectroscopy of CO in the Heme Pocket of the Oxygen Sensor FixL
  
  • Nuernberger Patrick
  • Lee Kevin F.
  • Bonvalet Adeline
  • Bouzhir-Sima Latifa
  • Lambry Jean-Christophe
  • Liebl Ursula
  • Joffre Manuel
  • Vos Marten H.
  Journal of the American Chemical Society, American Chemical Society, 2011, 133 (43), pp.17110. In heme-based sensor proteins, ligand binding to heme in a sensor domain induces conformational changes that eventually lead to changes in enzymatic activity of an associated catalytic domain. The bacterial oxygen sensor FixL is the best-studied example of these proteins and displays marked differences in dynamic behavior with respect to model globin proteins. We report a mid-IR study of the configuration and ultrafast dynamics of CO in the distal heme pocket site of the sensor PAS domain FixLH, employing a recently developed method that provides a unique combination of high spectral resolution and range and high sensitivity. Anisotropy measurements indicate that CO rotates toward the heme plane upon dissociation, as is the case in globins. Remarkably, CO bound to the heme iron is tilted by similar to 30 degrees with respect to the heme normal, which contrasts to the situation in myoglobin and in present FixLH-CO X-ray crystal structure models. This implies protein-environment-induced strain on the ligand, which is possibly at the origin of a very rapid docking-site population in a single conformation. Our observations likely explain the unusually low affinity of FixL for CO that is at the origin of the weak ligand discrimination between CO and O(2). Moreover, we observe orders of magnitude faster vibrational relaxation of dissociated CO in FixL than in globins, implying strong interactions of the ligand with the distal heme pocket environment. Finally, in the R220H FixLH mutant protein, where CO is H-bonded to a distal histidine, we demonstrate that the H-bond is maintained during photolysis. Comparison with extensively studied globin proteins unveils a surprisingly rich variety in both structural and dynamic properties of the interaction of a diatomic ligand with the ubiquitous b-type heme-proximal histidine system in different distal pockets. (10.1021/ja204549n)
  DOI : 10.1021/ja204549n
• Measurement of circular dichroism dynamics in a nanosecond temperature-jump experiment
  
  • Khuc Mai-Thu
  • Mendonça Lucille
  • Sharma S.
  • Volk M.
  • Solinas Xavier
  • Hache François
  Review of Scientific Instruments, American Institute of Physics, 2011, 82 (5). The use of a fast temperature jump (T-jump) is a very powerful experiment aiming at studying protein denaturation dynamics. However, probing the secondary structure is a difficult challenge and rarely yields quantitative values. We present the technical implementation of far-UV circular dichroism in a nanosecond T-jump experiment and show that this experiment allows us to follow quantitatively the change in the helical fraction of a poly(glutamic acid) peptide during its thermal denaturation with 12 ns time resolution. Cop. 2011 American Institute of Physics. (10.1063/1.3592331)
  DOI : 10.1063/1.3592331