Accueil > Research > Highlights > Archives > 2013
How do fluorescent proteins die ?Fluorescent proteins are widespread markers in cellular imaging, providing a highly flexible toolbox to investigate live cells. Unfortunately, contrary to organic dyes, fluorescent proteins are particularly sensitive to the photobleaching phenomenon, the definitive loss of fluorescence following photo-induced destruction of the chromophore. Photobleaching is particularly problematic in super-resolution microscopy techniques, which are being rapidly developed today, limiting the resolution that can be achieved. By combining kinetic crystallography, optical and Raman spectroscopy, molecular dynamics simulations, mass spectrometry, and super resolution microscopy, we have investigated the photophysical mechanisms leading to photobleaching of the fluorescent protein IrisFP. We have shown that depending on the illumination intensity used for the imaging experiment, two completely different photobleaching mechanisms show up. At low laser intensity, typical of a standard widefield microscopy experiment, an oxygen-dependent mechanism predominates. On the contrary, at high laser intensity, typical of super-resolution microscopy experiments, a redox-dependent mechanism prevails. The first mechanism, which generates reactive oxygen species (ROS) in the cell is thus expected to be more cytotoxic than the second mechanism, which does not generate such species. Thus, this work suggests in a counterintuitive manner that by increasing laser intensity at constant those, less cellular damages would be created. This hypothesis now needs to be experimentally verified.
Structural evidence for a two-regime photobleaching mechanism in a reversibly switchable fluorescent protein. |
Choreographed origami : folding ribosomal RNA involves paired tagging sequenceBuilding ribosomes – the cell’s protein factories – is like a strictly choreographed dance. Other ‘machines’ inside the cell have to produce specific RNA molecules and fold them into the right shape, then combine the folded RNA with proteins to form a working ribosome. The study combined nuclear magnetic resonance experiments performed at EMBL and neutron scattering experiments performed at the ILL in Grenoble, France. The structure of the box C/D enzyme reveals regulation of RNA methylation. |
Modelling of chemical reactions in the spotlighThe 2013 Nobel Prize in Chemistry awarded for the development of multiscale models for complex chemical systems Chemists used to create models of molecules using plastic balls and sticks. Today the modelling is carried out in computers, thanks in large part to the work done in the 1970s by Martin Karplus, Michael Levitt and Arieh Warshel. They developed powerful computer models that are used all around the world to understand and predict chemical processes. Applications of their discoveries are endless for both research and industry. " Detailed knowledge about chemical processes permits the optimization of catalysts for cars or the design of drugs and materials for solar cells" the Swedish Royal Academy of Sciences said. The theoretical work of the Nobel Laureates focused on molecular dynamics in general, and hybrid methods in particular. Martin Field (IBS/DYNAMOP), a postdoctorate in the laboratory of Martin Karplus, played a very important role in the initial development of hybrid methods (method coupling classical and quantum mechanics). Today his DYNAMO team and Patricia Amara (IBS/METALLO) continue this work in the IBS to study the structure and function of proteins and other biological macromolecules and their complexes. |
UJF prize awarded to F.X. GallatFrançois-Xavier Gallet won the Biology-Chemistry interface PhD thesis prize of the Chemistry and Biology Faculty at the Université Joseph Fourier (UJF). This prize honours PhD students who published their work as first authors in high-profile journals. During his PhD thesis, François-Xavier studied the dynamics of intrinsically disordered proteins and of protein-polymer nanohybrids by neutron scattering and complementary methods. |
Move to EPN Campus underway : exciting time ahead
From its initial size of 100 people, within 20 years the IBS has grown to an institute of over 240 individuals and with its installation on EPN campus (European Photon and Neutron science campus), continues on an international track. The new building of 9500 m2 is now ready to host new teams or start-up companies and will provide closer proximity to the European PSB partners (Partnership for Structural Biology). For one month the IBS will be a hive of activity with the move of researchers and platforms. Please note our new address : |
The NMR-Bio development project awarded by ERCTeams of Jerome Boisbouvier (IBS/NMR) and Olivier Hamelin (iRTSV) jointly develop technology solutions in isotopic labelling to push forward Biomolecular NMR frontiers. With support from CNRS, CEA and Grenoble Alps Innovation, the consortium had transformed these basic research findings into innovative market products that are now distributed by the CEA (www.nmr-bio.com). The project has been selected by the NMR-Bio is the third project from IBS Biomolecular NMR spectroscopy group selected by ERC (after SeeNanoLifeInAction ERC-consolidator grant awarded to J. Boisbouvier in 2010 and ProtDyn2Function ERC-starting grant awarded to Paul Schanda in 2012). Contact IBS researcher : : Jérome Boisbouvier |
Creation of an International Associated Laboratory between France (Grenoble) and Brazil (Campinas)An agreement creating an International Associated Laboratory (LIA) involving Ibs (for the French side) and LNBio and CNPEM (for the Brazilian side) was signed. This intenational associated laboratory named "BACWALL" aims to study the assembly and structure of macromolecular complexes involved in the synthesis of bacterial cell wall and virulence. This work may lead to significant advances in the understanding of bacterial virulence and thus develop new antibiotics. |
Coping mechanisms of pneumococcusHow cell division is ensured within pneumococcus ? IBS researchers show it’s based on the interaction of two key proteins. Their results could be used to find new antibiotics. Bacterial cell division requires the co-ordinated action of machineries composed by peptidoglycan biosynthetic enzymes and cell morphogenesis proteins. The regulation of these processes, notably in ovococci remains largely uncharacterized. The conserved eukaryotic-like Ser/Thr protein kinase of Streptococcus pneumoniae (StkP) plays a major role in cell shape and division. The molecular mechanisms underlining the regulatory function(s) of StkP through its interaction with the essential actor of septal peptidoglycan synthesis Penicillin-Binding Protein 2x (PBP2x) was investigated. We show that StkP and PBP2x interact directly in vitro and are present in the same membrane-associated complex in S. pneumoniae. We further show that they both display a late-division localization pattern at the division site and that their positioning is independent from each other. We demonstrate that StkP and PBP2x interaction is mediated by their extracellular regions and that the complex is dissociated in presence of muropeptides. All together, these data suggest a model in which StkP regulates cell division in the pneumococcus through the control of the septal peptidoglycan synthesis machinery. Interaction of Penicillin-Binding Protein 2x and Ser/Thr protein kinase StkP, two key players in Streptococcus pneumoniae R6 morphogenesis. Morlot C, Bayle L, Jacq M, Fleurie A, Tourcier G, Galisson F, Vernet T, Grangeasse C, Di Guilmi AM. Molecular Microbiology, 2013 Oct ;90(1):88-102 |
The 2013 Walter Hälg Prize awarded to Joe ZaccaiGiuseppe Zaccai (IBS) has just been awarded the prestigious Walter Hälg prize for an outstanding, coherent work in neutron scattering with long-term impact on scientific and/or technical neutron scattering applications. The European Neutron Scattering Association underlined his “pioneering contributions to the application of neutron scattering to a range of biophysical and biochemical problems in biology, which has provided important insights in the debate on the relationship between molecular structure and dynamics and biological function, and for his leading advocacy of the role of neutron scattering in biological research.” |
10th anniversary celebrations for Grenoble’s Partnership for Structural Biology (PSB)On 4th June more than 150 specialists from all across Europe gathered in Grenoble to celebrate the 10th anniversary of the Partnership for Structural Biology. The day was centred around talks by prestigious guests including David Stuart from the University of Oxford, UK, and Patrick Cramer from the Gene Center, University of Munich, Germany. Structural biology is one of the central tenets of research in Grenoble along with micro/nano technologies and energy. In 2003, three European and two French institutes joined forces to create the PSB and pool their knowledge and state-of-the-art equipment to study structural biology.
Members of the PSB : |
Recent insights into the function of human complement component C1q
Expression of recombinant human complement C1q allows identification of the C1r/C1s-binding sites, Bally I, Ancelet S, Moriscot C, Gonnet F, Mantovani A, Daniel R, Schoehn G, Arlaud GJ, Thielens NM. Proc Natl Acad Sci U S A, 2013 May 21 ;110(21):8650-5 |
Looking at bacterial cell walls with NMRThe cell wall of bacteria is composed of several biopolymers building a shell around the cell, which allows recognition and adhesion to hosts, as well as regulation of other important cellular functions. A better understanding of the complex cell wall structure and interactions may help scientist to develop new antibiotics. Solid-state nuclear magnetic resonance (NMR) offers an interesting tool to look at this huge and complex molecular system in intact conditions, as it is principally not limited by the size of the object and do not require any crystallization. However, it suffers from its intrinsic low sensitivity, especially for the case of cell wall studies in living cells. Using high-field dynamic nuclear polarization (DNP), an emerging technique aimed at improving sensitivity of NMR, researchers of INAC and IBS demonstrate that signal intensity from the bacterial cell wall can be 24-fold enhanced, opening the possibility of atomic-scale studies of cell-wall interactions. Solid-State NMR on Bacterial Cells : Selective Cell Wall Signal Enhancement and Resolution Improvement using Dynamic Nuclear Polarization. Hiroki Takahashi, Isabel Ayala, Michel Bardet, Gaël De Paëpe, Jean-Pierre Simorre, and Sabine Hediger. J Am Chem Soc. 2013 Feb 12 |
Cryo-electron microscopy three-dimensional structure of a phage infecting a bacteia
Cryo-electron microscopy three-dimensional structure of the jumbo phage ΦRSL1 infecting the phytopathogen Ralstonia solanacearum. Effantin G, Hamasaki R, Kawasaki T, Bacia M, Moriscot C, Weissenhorn W, Yamada T, Schoehn G. Structure, 2013 Feb 5 ;21(2):298-305 |
Features responsible for O2 tolerance of membrane-bound NiFe-hydrogenases
1. Crystal structure of the O2-tolerant membrane-bound hydrogenase 1 from Escherichia coli in complex with its cognate cytochrome b. Volbeda A, Darnault C, Parkin A, Sargent F, Armstrong FA and Fontecilla-Camps JC, Structure, 21 : 184-190 (2013). |
Creation of the “Integrated Structural Biology, Grenoble” mixed research instituteThe Unité Mixte de Service (mixed research institute) UMS 3518 (CNRS-CEA-UJF-EMBL) was created on January 1st 2013, in Grenoble. This new unit brings together facilities from the IBS and UVHCI, and represents an impressive collection of state-of-the-art equipment for integrated structural biology. The unit is open to national and international scientific communities, as well as industry. Development of integrated structural biology at the Grenoble and European scale In general, it is now accepted that this type of biology requires a collection of facilities corresponding to the equivalent of large instruments. The aim of the European initiative ESFRI is to establish such infrastructures. Within this context, the project Instruct (Integrating Structural Biology), which began in 2011, defines 15 reference centers for integrated structural biology in Europe and 5 affiliated centers. Grenoble, and more specifically the combination of the two French units of the PSB (IBS and UVHCI), represents one of the major reference centers for Instruct. The project FRISBI (French Infrastructure for Integrated Structural Biology), comprised of the two French Instruct centers (Grenoble and Strasbourg) as well as three other nationally important structural biology centers, was selected as part of the program “Investissement d’Avenir”. Grenoble has thus received 11.2 million € to support major improvements in most of the PSB facilities, notably including the purchase of new NMR spectrometers and electron microscopes. The missions of the new UMS Darren Hart (EMBL) has been appointed director of the new Unité Mixte de Service, and is assisted by Yvette Gaude in financial administration. The governance of the UMS will involve the management of IBS and UVHCI within a local steering committee. The vast majority of scientific and technical personnel working at the facilities will keep their affiliation with the IBS and UVHCI research teams, to allow the best possible interaction between facility and research. The equipment present on the day of the creation of the UMS will thus evolve according to the needs of scientific projects and the UMS will ensure the sustained operation of the facilities by guaranteeing optimum access to all types of users. |