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Biochemistry, Biophysics and Structural Biology department

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  • Monday 13 April 2015 11:00-12:00 - Harold P. Erickson - Department of Cell Biology, Duke University, NC USA

    Curved FtsZ protofilaments bend the bacterial membrane, and generate the constriction force for cytokinesis.

    Lieu : Bibliothèque, Bâtiment 34 - Campus de Gif

  • Wednesday 18 November 2015 11:30-12:30 - Olivia du Roure - PMMH, ESPCI, Paris

    Mechanics and growth of dense branched actin networks

    Résumé : Cell mechanics is fundamental in many cellular processes both in physiological and pathological situations. The actin cytoskeleton is responsible for the main part of this mechanics. In cells, actin filaments are very dynamic polymers spatially organized by many different partners. We recently developed a new approach to study at the same time the mechanics and the dynamics of growing Arp2/3-branched actin networks. This new technique combines the use of the biochemical machinery at play in the leading edge of a cell and magnetic particles to apply mechanical constraints. The idea is to use dipolar attractive forces that develop between superparamagnetic micro-sized objects to deform, in a controlled way, the dense branched actin networks grown from the surface of magnetic particles. One of the main advantages of this technique is its high throughput that allows reliable measurements to be performed. We carried out a first study that established the link between elastic properties of these networks and their architecture in assembly from a mix of purified proteins. In a new version of the technique, we developed new particles with flat surfaces that allow non-linear measurements to be done and growth under constraints to be followed. In this talk, I will present our recent results obtained with this approach showing that depending on the degree of branching the actin networks may or may not experience a transition from an elastic growing gel to a liquid-like flowing material.

    Lieu : Bibliothèque - Bâtiment 34, campus Gif-sur-Yvette

  • Wednesday 3 February 2016 11:00-12:00 - Dr. Stéphane Roméro - Centre Interdiscipliaire de Recherche en Biologie, Collège de France, Paris

    Sensing of the environment: how actin reorganization rules filopodia dynamics

    Résumé : Filopodia are functional units described as finger-like membrane extensions acting as sensor organelles to probe the environment and initiate contacts, transmit signals or guide cell migration. How cells integrate external signal to reorganize the actin cytoskeleton in physiological and physiopathological context is crucial for the sensing function of filopodia. Our work shows that actin polymerization is regulated by extracellular signals to control filopodia dynamics, and that Shigella hijacks filopodia function and cellular actin reorganization to invade epithelial cells.
    Invited by Christophe Le Clainche, Team Cytoskeleton Dynamics and Motility

    Lieu : Bibliothèque, 2ème étage - Bâtiment 34, Campus de Gif

  • Friday 8 April 2016 11:00-12:00 - Emmanuelle Schmitt - Laboratoire de Biochimie, UMR7654 Ecole Polytechnique, CNRS

    Structural and functional studies of the translation initiation factor e/alF2

    Résumé : Eukaryotic and archaeal translation initiation complexes have in common a functional core containing mRNA, the ternary initiation complex (e/aIF2, GTP, Met-tRNAiMet), e/aIF1 and e/aIF1A bound to the small ribosomal subunit. In eukaryotes, the functional core is made more complex by many additional factors, most of them being involved in a long-range scanning of mRNA, necessary to decipher the initiation codon. In archaea, long-range scanning does not occur thanks to the occurrence of Shine-Dalgarno sequences or of very short 5’ untranslated regions on mRNA. Concomitantly, archaeal translation initiation only requires the core complexes. Within the core complex, e/aIF2, in its GTP-bound form, is responsible for selecting the methionylated initiator tRNA and bringing it to the small ribosomal subunit. The establishment of the connection between the start codon on the mRNA and the anticodon of the initiator tRNA is also coupled to the release of one molecule of phosphate resulting from the hydrolysis of GTP bound to eIF2. The function of the e/aIF2 protein in eukaryotes and archaea is therefore crucial for translation initiation. We have been studying factor e/aIF2 for the last few years. In particular, using eukaryotic or archaeal versions of this factor, many data have been accumulated to help understand how it operates. Finally, using purified versions of the archaeal translation initiation complexes, we recently solved the structure of two stages of archaeal translation initiation by Cryo-EM. The two snapshots highlight a new network of interactions crucial for translation initiation in archaea. According to the conservation of the core complex in eukaryotes, this network of interactions is also anticipated to be relevant for the eukaryotic translation initiation process.
    Invited by Marie-Hélène Le Du, Nuclear enveloppe, Telomeres and DNA repair team

    Lieu : Bâtiment 144 – Service de Biologie Intégrative et Génétique Moléculaire - CEA Saclay

  • Friday 15 April 2016 11:00-12:00 - Malene Ringkjobing-Jensen - Institut de Biologie Structurale, UMR5075 CEA/CNRS/Univ. Grenoble Alpes

    Understanding the conformational behaviour of intrinsically disordered proteins and their dynamic complexes using NMR spectroscopy

    Résumé : Invited by Sophie Zinn-Justin, Nuclear enveloppe, Telomeres and DNA repair team

    Lieu : Bâtiment 144 – Service de Biologie Intégrative et Génétique Moléculaire - CEA Saclay

  • Thursday 12 May 2016 14:00-15:00 - Maud Hertzog - Laboratoire de Microbiologie et Génétique Moléculaire, CNRS UMR 5100, TOULOUSE

    Molecular characterization of pneumococcal RecASp assembly.

    Résumé : Homologous Recombination (HR) is crucial for genome maintenance and dynamics in all kingdoms of life. HR is universally catalyzed by recombinases, i.e., RecA in bacteria, Rad51 in eukaryotes, and RadA in archaea. HR recombinases catalyze the exchange between complementary DNA strands via its polymerization along the DNA substrates. Originally discovered in the human pathogen Streptococcus pneumoniae (Sp), bacterial transformation is a genetically programmed process leading to genetic diversity. This process is proposed to favor spreading of antibiotic resistance or vaccine escape of this human pathogen. This horizontal gene transfer process, which relies on the physiological shift of bacteria into natural competence, consists in the integration of exogenous DNA into the genome by HR. Our aim is to dissect the different steps of the mechanism of action of RecA from Sp (RecASp) during genetic transformation. We are using fluorescent-labeled proteins to reconstitute RecASp filaments along ssDNA molecules by combining TIRF (Total Internal Reflected Fluorescence) microscopy and microfluidics (collaboration with Kowalczykowski S). For the first time, we did a direct visualization of RecASp filament assembly on single molecules of ssDNA. We have observed that the intrinsic length of the RecASp filaments was significantly shorter than the RecAEc (E. coli) filaments. Only the use of non-hydrolysable analogue (ATPɣS) stabilizes the filaments, which are shorter than RecAEc in the same conditions. By electronic microscopy, RecASp was also found to form shorter filaments on ssDNA than RecAEc (collaboration with P. Dupaigne-E. Le Cam’s lab). Using Fluorescence Correlation Spectroscopy (FCS), we have been able to measure the kinetic of assembly showing a half time of polymerization similar between RecAEc and RecASp (collaboration E. Margeat-CBS Montpellier). Despite the different intrinsic length between the RecAEc and RecASp filament, we measured an equivalent ATP hydrolysis activity for RecASp or RecAEc proteins suggesting that the coupling between depolymerization and ATP hydrolysis could differ between the 2 filaments. Finally in vivo, we have shown that RecAEc is unable to substitute RecASpboth for genome maintenance and genetic transformation. All together these results show that RecASp has unique polymerizing properties required for the HR reaction dedicated to the genetic transformation in Streptococcus pneumoniae.

    Lieu : Bibliothèque, 2ème étage - Bâtiment 34, Campus de Gif

  • Friday 8 July 2016 11:00-12:00 - Rémi Veneziano

    Designer DNA nanoparticles for spatially-oriented macromolecular assemblies

    Résumé : Organizing macromolecules at the nanometer scale to mimic specific cellular assemblies will enable a better understanding of natural molecular mechanisms and potentially allow the synthesis of new biomimetic systems. In the last several years this formidable challenge has been partially addressed with the emergence of DNA as a promising material that can be specifically programmed at the nanometer scale to scaffold nanoarchitecture assemblies. More recently DNA origami nanostructures have demonstrated major potential to serve as a versatile medium to program and organize complex molecular architectures at the nanoscale. These spatially addressable DNA nanostructures with finite size have been used in various applications such as enzyme cascade reconstitution, membrane nanopore formation, delivery vehicles, and excitonic devices, among others. While potential applications of these objects have grown significantly during the last couple of years, their manual design and their increasing complexity have limited this technology to experts in the field. To respond to this, we have developed a fully automated top-down approach to easily design and directly synthesize arbitrary programmable 3D DNA nanoparticles that can be used by non-experts, needing only a user-defined geometry as input. The characterization methods used, including cryo-electron microscopy and atomic force microscopy, show robust and monodisperse nanoparticles with well-defined 3D structures. These particles are stable in a variety of physiological buffers and are easy to modify, offering a real alternative to reconstruct complex macromolecular assemblies to enable diverse biological applications such as targeted drug delivery or excitonic circuits.
    Recent references:
    Designer nanoscale DNA assemblies programmed from the top down.
    Veneziano R, Ratanalert S, Zhang K, Zhang F, Yan H, Chiu W, Bathe M.
    Science. 2016 Jun 24;352(6293):1534.
    Bordetella pertussis adenylate cyclase toxin translocation across a tethered lipid bilayer.
    Veneziano R, Rossi C, Chenal A, Devoisselle JM, Ladant D, Chopineau J.
    Proc Natl Acad Sci U S A. 2013 Dec 17;110(51):20473-8.

    Lieu : Bibliothèque, 2ème étage - Bâtiment 34, campus de gif

  • Tuesday 4 October 2016 14:00-15:30 - MENON Anant - Weill Cornell Medical College, New York

    Phospholipid scramblases - new names to match old activities

    Résumé : Polar lipids must flip rapidly, and often bi-directionally, across membranes to support cellular life. As flipping is energetically costly, specialized transporters increase its intrinsically low rate to a physiologically relevant level. While some of these transporters couple ATP hydrolysis to lipid movement, many function without any discernible metabolic energy input. The molecular identification of these latter ‘ATP-independent flippases’, also termed ‘scramblases’, has eluded researchers for decades - until recently. I will discuss the first examples of this novel class of proteins: afTMEM16, a fungal homolog of the TMEM16 family of ion channels is a Ca2+-dependent phospholipid scramblase [1] and rhodopsin, a prototypical G protein-coupled receptor, is a constitutively active phospholipid scramblase responsible for the homeostasis of photoreceptor disc membranes [2-4]. New data on the possible mechanisms of lipid scrambling will be presented.
    1. Malvezzi, M. Chalat, M.N., Janjusevic, R., Picollo, A., Terashima, H., Menon, A.K., Accardi, A. (2013) Ca2+-dependent phospholipid scrambling by a reconstituted TMEM16 ion channel. Nature Commun. 4, 2367.
    2. Menon, I., Huber, T., Sanyal, S., Banerjee, S., Barré, P., Canis, S., Warren, J.D., Hwa, J., Sakmar, T.P., Menon, A.K. (2011) Opsin is a phospholipid flippase. Curr. Biol. 21, 149-153.
    3. Goren, M.A., Morizumi, T., Menon, I., Joseph, J.S., Dittman, J.S., Cherezov, V., Stevens, R., Ernst, O.P., Menon, A.K. (2014) Constitutive phospholipid scramblase activity of a G protein-coupled receptor. Nature Commun. 5, 5115.
    4. Ploier, B., L. N. Caro, T. Morizumi, K. Pandey, J. N. Pearring, M. A. Goren, S. C. Finnemann, J. Graumann, V. Y. Arshavsky, J. S. Dittman, O. P. Ernst, and A. K. Menon (2016) Dimerization deficiency of enigmatic retinitis pigmentosa-linked rhodopsin mutants. Nature Commun. 7,12832.
    Invited by Guillaume Lenoir, Membrane Proteins and Membrane Systems Laboratory

    Lieu : Auditorium - Bâtiment 21, campus de gif

  • Friday 20 January 2017 11:00-12:00 - Pr. Patrice Soumillion - Laboratoire Biochimie Biophysique et Génétique des Microorganismes Institut des Sciences de la Vie, UC Louvain

    Accelerated evolution of enzymes: traps, hurdles and secret passages

    Résumé : Directed evolution of enzymes is a modern field of research in biochemistry and molecular biology aiming at generating biocatalysts endowed with new or improved properties. Although engineering enzymes with new specificities has been extensively reported, the evolution of new and efficient catalytic mechanisms within an enzyme active site remains extremely challenging. Here, we will focus on the accelerated evolution of a D-alanyl-D-alanine-peptidase (DD-peptidase) into a beta-lactamase as a model system. In nature, these two phylogenetically related enzyme families share common fold and active site motifs although beta-lactamases have acquired an additional hydrolytic machinery. Through success and failures, we have finally succeeded in converting a DD-peptidase into a beta-lactamases by following a counter-intuitive evolutionary trajectory that involves an initial neutralization of the starting activity. This neutralization occurs by drift selection without any pressure for activity and results in variants featuring a reshaped active site but with conservation of all the essential residues for catalysis. Interestingly, the neutralized mutants have acquired the potential to further evolve into beta-lactamases. Expression of the newly born enzymes is however associated with a fitness cost indicating that, besides antibiotic resistance, the new hydrolytic activity is creating a problem to the bacteria. Although natural DD-peptidases and beta-lactamases are phylogenetically related, these activities appear mutually exclusive. Overall, our results are shedding light on an unexplored property of natural enzymes, i.e. their security against interfering activities. Securing enzymes may constitute an important obstacle for their natural evolution as well as for their engineering in the laboratory.

    Lieu : Salle Olivier Kahn - Bâtiment 410, Campus d’Orsay

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  • Friday 18 September 2015 13:30-16:30 - Abbas El Sahili - I2BC - Département B3S

    Structural and functional characterization of acc operon from A. tumefaciens C58

    Résumé : Agrobacterium tumefaciens is a soil bacterium responsible of the crown gall in plants once it possess the Ti plasmid also called virulence plasmid. The bacterium transfers a piece of DNA from the pTi in the plant genome. The transferred DNA codes on one hand for plant hormone synthesis, leading to tumor formation. Those tumors are colonized by bacteria. On the other hand, the transferred DNA code for the synthesis of small molecules called opines that are used as nutrients by A. tumefaciens. The opine agrocinopine A induces the production of quorum sensing signals leading to the spread of the virulence plasmid from pathogenic to nonpathogenic bacterium. Agrobacterium radiobacter K84, a nonpathogenic bacterium, produces an antibiotic called agrocin 84 that kills A. tumefaciens.
    Import and catabolism of agrocinopine A are operated by acc operon, present on the pTi. The periplasmic binding protein AccA (PBP AccA) associated with the ABC transporter imports the opine in the periplasm where it is degraded by AccF and AccG. AccR regulates the expression of the acc operon and TraR transcription factor, central in quorum sensing signaling. AccA also allow the import of agrocin 84, which is activated by AccF. My PhD work allowed through structure-function studies the definition of AccA and AccF specificity and to initiate the study of transcription factor AccR. The structure study of AccA in complex with agrocinopine A, agrocin 84 and derivatives from these molecules revealed that the pyranose-2-phosphate motif, common in the two molecules, only was recognized. Microcalorimetry and autofluorescence measurements confirmed these conclusion. The pyranose-2-phosphate motif would allow any compound to be transported. The structure of the enzyme AccF showed that again only the pyranose-2-phosphate group is recognized. From the structure and molecular modelisation of the substrate in the active site, an original mechanism of the rupture of the phosphodiester bond is proposed. Microcalorumetry affinity measures showed that only arabinose-2-phosphate and glucose-2-phosphate are capable of interacting with AccR. in cellulo experiments showed also that only these compound can regulate the expression of quorum sensing.
    My work sheds a new light on import and use of agrocinopine in A. tumefaciens. Recognition specificity of PBP AccA for a part of the imported molecule is observed in other PBPs and opens new ways for rational design of antibiotic compounds that, just like agrocin 84, would use the “Trojan horse” strategy.
    Keywords: Agrobacterium tumefaciens, Agrocinopine A, Agrocine 84, quorum sensing, PBP, phosphodiesterase, transcription regulation.

    Lieu : Bibliothèque, Bâtiment 34 - Campus de Gif

  • Tuesday 22 September 2015 14:30-17:00 - Guillaume Gaullier - I2BC - Département B3S

    Etude structurale de l’assemblage du complexe télomérique humain TRF2/RAP1

    Résumé : Les télomères sont les extrémités des chromosomes linéaires des eucaryotes. Ils sont constitués de répétitions en tandem d’un motif court riche en guanine, et liés par des protéines spécifiques. Chez les vertébrés ces protéines forment un complexe appelé le shelterin et dont l’intégrité est critique pour assurer la réplication correcte des extrémités des chromosomes, et pour les protéger contre une prise en charge illicite par les voies de réparation des cassures double-brin de l’ADN. Des dysfonctions des télomères engendrent une instabilité du génome qui peut conduire à la sénescence ou au cancer. Les télomères représentent une région subnucléaire où les protéines du shelterin sont fortement enrichies, ce qui permet l’implication dans les fonctions biologiques d’interactions de basse affinité. Parmi les protéines du shelterin, la protéine de liaison aux répétitions télomériques TRF2 et son partenaire constitutif RAP1 sont les facteurs majeurs responsables de la protection des extrémités. Nous avons étudié en détails l’assemblage du complexe TRF2/RAP1 par des approches intégrées de biologie structurale, de biophysique et de biochimie. Nous avons montré que cet assemblage s’accompagne d’importants ajustements de conformation des deux protéines, et implique une interaction de basse affinité qui engage de grandes régions des deux protéines et affecte leurs propriétés d’interactions.

    Lieu : INSTN - Saclay

  • Wednesday 7 October 2015 14:00-16:30 - Isaline Herrada - I2BC - Département B3S

    Etude des interactions protéine-protéine à l’enveloppe nucléaire

    Lieu : Amphithéâtre Bloch de l'Orme des merisiers - Bâtiment 772, Route de l’Orme, 91190 Saint-Aubin

  • Friday 9 October 2015 14:00-16:30 - Dyana Sanchez - I2BC - Département B3S

    Etude structurale et fonctionnelle de la régulation de la compétence et du processus de transformation chez Streptococcus pneumoniae

    Résumé : The natural genetic transformation contributes to the maintenance and the evolution of the genomes in bacteria; it is a key mechanism to adapt to their environment. It allows the integration of exogenous DNA into the bacterial chromosome by homologous recombination during a particular state called competence.
    My thesis focused on the regulation of the competence state in S. pneumoniae (ComD, ComE), and on the interactions between the proteins involved in the uptake, the processing and recombination of exogenous DNA (DprA, RecA). In this bacterium, the opening of the competence is under the control of the two-component system ComD-ComE, who induces the transcription of target genes. DprA is one of the protein induced during the competence state, it is very conserved into the bacterial kingdom, and is involved in the closure of competence via direct interaction with ComE. DprA is also a key transformation protein involved in processing the incoming DNA, protection against nucleases, and recruitment of the RecA recombinase. SAXS analysis of the ComD-ComE, resolution of the crystallographic structure of ComE REC domain study of the interactions between ComE and its promoter regions allowed us to understand the choreography of competence opening in S. pneumoniae. Meanwhile, we studied spDprA interactions with DNA and with RecA. These data allowed us to propose an interaction model between DprA and RecA in S. pneumoniae and to propose a mechanism for RecA’s loading on the ssDNA by DprA. I focused too on H. pylori DprA participating on the resolution of the 3D structure of the C-terminal domain by NMR and studying its interaction with the dsDNA.
    Keywords: Natural transformation, competence, two components systems, protein-protein and nucleic acids interactions, high and low structure resolution, ComD, ComE, DprA, RecA, S. pneumoniae, H. pylori.

    Lieu : Salle de Lederer - Bâtiment 430, Campus d’Orsay

  • Wednesday 4 November 2015 14:30-16:30 - Nahuel PERROT - Co-directeurs : Nadege JAMIN et Manuel GARRIGOS

    Production dans Escherichia coli de vésicules enrichies en cavéoline-1(32-178) canine ou son fragment (76-178)

    Résumé : La cavéoline-1, une petite protéine membranaire de 21 kDa, est la protéine membranaire majoritaire d’invaginations de la membrane cytoplasmique appelées cavéoles. Enrichis en cholestérol et sphingolipides, ces domaines ont un rôle important dans de nombreux aspects de la vie cellulaire et constituent une véritable plateforme d’interactions protéiques et lipidiques. Ces dernières années, malgré le nombre important de travaux concluant à l’implication de la cavéoline-1, ou des cavéoles, dans de nombreux processus cellulaires ou pathologiques, les données quant à l’organisation de cette protéine au sein de la membrane plasmique restent très éparses. Aussi, l’objectif principal de ce travail est de contribuer à l’acquisition de données structurales sur cette protéine. Ce travail se base sur les expressions hétérologues d’une isoforme de la cavéoline-1 canine ou de l’un de ses fragments, dans un hôte bactérien, sous la forme de protéines de fusion associées à la Maltose Binding Protein. Ces expressions induisent la formation de vésicules intracytosoliques composées majoritairement de la protéine exprimée. Aussi, la première partie du travail est consacré à la mise en place d’un protocole de purification de ces vésicules dans des conditions natives, répondant au critère de réaliser une étude structurale de cette protéine n’impliquant pas l’usage de détergent. La deuxième partie porte sur une application potentielle de ces vésicules, et en particulier pour des essais de caractérisation d’une enzyme membranaire, la glutathion-S-transférase microsomale de rat. Enfin une troisième partie est dédiée à l’analyse de simulations de dynamique moléculaire dans le cadre d’études d’interactions au sein de systèmes membranaires.
    Mots-clés : cavéoline,protéine,membranes.

    Lieu : Auditorium - Bat 21, Campus de gif

  • Monday 9 May 2016 14:00-17:00 - Esra Karakas Dagli - Département B3S

    Molecular Dynamics simulations of the light-harvesting 2 complex of a purple bacterium in different micellar and membrane environments.

    Résumé : Purple photosynthetic bacteria, such as Rhodopseudomonas acidophila strain 10050 have a synthetic apparatus which is composed by membrane protein complexes (LH1 and LH2) with specialized pigments to harvesting the light. This latter is mainly absorbed by photosynthetic pigments of the LH2 complex and the resulting excitation energy is transferred to LH1 complex, then to reaction center. Fluorescence spectroscopy experiments carried out with a single LH2 complexes have shown that the intensity and the position of the electronic transition of complexes can strongly fluctuate with the time describing a « dynamic disorder ». This disorder is strongly related to the biological function of the LH2 complex. The molecular description of the dynamic disorder of LH2 complex is great interest to get a precise comprehension about the ability to use of solar energy of these complexes. In order to obtain a precise picture of the dynamical disorder at the atomic level, we combined classical and quantum molecular dynamics simulations. We first model the LH2 complex in different biomimetic environments composed of detergents (dimethyldodecylamine-N-oxide, LDAO) and the β octyl glucoside, bOG)) or a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, POPC membrane. To validate the LH2-detergents complex models, small angle X-ray scattering (SAXS) experiments were performed. We find a good agreement of the SAXS results with simulation in particular with LH2-bOG models. In the second part of this thesis, we specifically studied with MD simulations the LH2 structure as well as the interactions between peptide and pigment and pigment-pigment in the different environments. Our results revealed a significant impact of the environment on the complex structure and dynamic of the interactions between pigments and pigment-protein depending on the environment.
    Finally to relate these variations with the molecular disorder we performed additional quantum calculation to model the absorbance variations changes by taken some representative snapshots of the simulations.
    Keywords : molecular dynamics, LH2 complex, POPC, LDAO, bOG, membrane, SAXS, dynamic disorder
    Team : Laboratory of Bioenergetics, Metalloproteins and Stress
    PhD Director : Bruno Robert

    Lieu : Auditorium - Bâtiment 21, Campus de Gif

  • Friday 16 September 2016 14:00-17:00 - Loïc Marty

    Structures et spécificité de protéines périplasmiques de fixation (PBP) pour les mannityl-opines chez Agrobacterium tumefaciens

    Résumé : L’agent pathogène Agrobacterium tumefaciens induit, chez les plantes, le développement de tumeurs dans lesquelles il prolifère, en intégrant un fragment de son plasmide Ti de virulence dans le génome de son hôte. Les tissus transformés synthétisent des composés originaux, appelés opines, qui sont utilisés comme nutriments spécifiques par la bactérie. Une vingtaine d’opines sont connues à ce jour, et chacune d’elles peut être métabolisée par des souches d’Agrobacterium tumefaciens possédant les gènes de transport et de catabolisme qui lui sont associés, ce qui apparait comme un avantage compétitif dans la colonisation de la tumeur. La présence de ces gènes dépend du type de plasmide Ti que la souche pathogène possède.
    Agrobacterium tumefaciens B6 possède un pTi de type octopine, qui porte les gènes de métabolisme des mannityl-opines, qui sont la mannopine, l’acide mannopinique, l’agropine et l’acide agropinique. La mannopine et l’acide mannopinique sont synthétisés par la même enzyme, et ont pour précurseurs respectivement la désoxy-fructosyl-glutamine (DFG) et le désoxy-fructosyl-glutamate (DFGA), tous deux opines de la famille de la chrysopine. La DFG est aussi un composé d’Amadori répandu et assimilable par de nombreux organismes. La mannopine sert de précurseur pour la synthèse de l’agropine. Enfin, la mannopine, l’acide mannopinique et l’agropine peuvent toutes trois se lactamiser spontanément en acide agropinique.
    Malgré la similarité chimique de ces quatre opines, chacune est transportée par une protéine périplasmique de fixation (PBP) associée à un transporteur ATP-binding Cassette (ABC) différent. La PBP sélectionne et fixe une opine pour l’apporter au transporteur ABC, qui permet le passage de l’opine dans le cytoplasme grâce à l’hydrolyse de deux molécules d’ATP. La spécificité du transporteur entier est déterminée par la PBP.
    Des études génétiques chez des souches possédant un pTi de type octopine ont montré que le système PBP-transporteur ABC AgaABCD est spécifique de l’acide agropinique, AgtABCD spécifique de l’agropine, MoaABCD spécifique de l’acide mannopinique et que MotABCD transporte la mannopine et également l’acide mannopinique. Chez la souche C58, qui ne possède pas un pTi de type octopine, le système de transport SocAB, codé par des gènes situés sur le plasmide cryptique At, transporte la DFG comme nutriment, et semble aussi capable d’importer la mannopine.
    Mon travail de thèse a permis, dans un premier temps, de caractériser les fortes affinités et la spécificité des PBP AgaA et AgtB pour l’acide agropinique, de la PBP MoaA pour l’acide mannopinique et de la PBP SocA pour la DFG, mais aussi la non spécificité de MotA pour la mannopine, l’acide mannopinique et la DFG, ce qui remet en question les affinités précédemment décrites pour AgtB et SocA. Dans un deuxième temps, ce travail a apporté les bases moléculaires et structurales des complexes PBP-mannityl-opines, complexes jamais caractérisés auparavant. Enfin, dans un troisième temps, la structure de la PBP AttC chez la souche C58, annotée comme mannopine-like, a été déterminée, et les expériences d’interaction ont montré qu’elle n’interagit avec aucune mannityl-opine, ce qui conduit à une révision de son annotation.
    Mes travaux apportent un éclairage nouveau sur l’import des mannityl-opines chez Agrobacterium tumefaciens. Le fait qu’aucun des transporteurs étudiés ne permette l’import de l’agropine laisse penser qu’il existe une autre PBP ou un autre système de transport encore inconnu assurant cette fonction, ouvrant la voie vers de nouvelles études sur les pTi de type octopine et agropine.

    Lieu : Bibliothèque, 2ème étage - Bâtiment 34, Campus de Gif

  • Tuesday 27 September 2016 14:00-17:30 - LuYan Cao

    Structural Basis of Kinesin Motility

    Résumé : Directeurs de thèse: Marcel Knossow et Benoit Gigant

    Lieu : Bibliothèque, 2ème étage - Bâtiment 34, campus de Gif

  • Wednesday 28 September 2016 14:00-17:30 - Hassina Azouaoui-Boumzar

    Étude fonctionnelle d’un transporteur de lipides (flippase) de la levure S. cerevisiae: l’ATPase P4 Drs2p et sa sous-unité associée Cdc50p

    Lieu : Auditorium - Bâtiment 21, campus de Gif

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  • Monday 18 December 2017 09:00-18:00 -

    B3S Department scientific Day

    Résumé :

    Programme Journée B3S - 18 décembre 2017

    Ci-joint, le programme quasi-définitif de la journée.
    Les inscriptions sont encore possibles jusqu’à jeudi (7 décembre) en remplissant le sondage à l’adresse :
    Si vous souhaitez présenter un poster, merci d’envoyer un titre et une liste d’auteurs (en précisant qui présentera le poster le 18/12) à Jessica Andreani

    Lieu : Auditorium - Bâtiment 21, campus Gif-sur-Yvette

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