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Institutes/ Institute of Biology and Technology Saclay (iBiTec-S)/ Units/ Bioenergetics, structural biology, and mechanisms (SB2SM/URA2096 CNRS)/ Structural biology and radiation biology laboratory (LBSR)/ Molecular assemblies and signaling (R. Guérois & F. Ochsenbein)

Molecular assemblies and signaling

Raphaël GUEROIS & Françoise OCHSENBEIN
CEA Saclay/Bât. 144
91 191 Gif sur Yvette cedex
Tél : 01 69 08 67 17 / 01 69 08 96 79
raphael.guerois@cea.fr / françoise.ochsenbein@cea.fr


Prediction, characterization and inhibition of protein-protein complex structures combining experimental and bioinformatic strategies.


Human resources
Raphaël GUEROIS , Group Leader
Françoise OCHSENBEIN , Group Leader
Gwenaëlle MOAL-RAISIN, Technician
Jessica ANDREANI, PhD Student
Nicolas RICHET, PhD Student



Research Programs

Our team combines computational and experimental strategies to unravel the molecular bases underlying the dynamic assembly of specific protein networks. Most of signaling pathways rely on transient interactions between proteins so as to switch off and on specific cell responses. Our main concern is to characterize at high resolution the structure of the complexes and elucidate the mechanisms underlying their assembly. The possibility to disrupt these protein/protein interactions by specific exogenous compounds opens novel and fascinating perspectives. In that scope, we have undertaken the rational design of compounds that inhibit protein assemblies activated after a genotoxic stress.

©cea/R.Guérois

 
We are particularly interested in chaperoning processes regulating the assembly of various complexes. Chaperones are targets of major interest because of their central role in controling of the assembly itself. We are presently focusing on two assembly processes : (i) the early assembly of chromatin by the histone chaperone Asf1 (ii) the assembly of complexes involved in the ubiquitination and degradation of proteins. Both processes play an active role in maintaining genome integrity after various stresses, in particular after the exposure to ionizing agents. Exposure to natural radiations, to endogenous oxydative stresses or to ionizing agents activates processes that protect the genomic information. In particular, complex DNA repair machineries are essential for the recovery of the original DNA. When these systems are not working properly severe pathologies such as cancer may be induced. Understanding the way, cell systems manage to resist to these stresses constitute a major concern of the CEA research.

 

In that context, we are actively collaborating with several teams of cell biologists and geneticists in and outside the CEA. These collaborations are an essential ground to connect our molecular investigations with complex phenotypes observed in the cell context. Among these, for the research project dedicated to the study of chromatin assembly and DNA damage signalling, we benefit from a long-term collaboration with the teams of Carl Mann and of Marie-Claude Marsolier-Kergoat and more recently with the groups of Ariel Prunell, Marc Lipinski and Vasily Ogryzko. For the project dedicated to the regulation of the assembly of the ubiquitin and degradation complexes, we are synergising our efforts with Ken Shirasu’ lab at the RIKEN Institute of Yokohama and more recently with Laurent Noël at the CEA Cadarache.

The specificity of our approaches is to combine predictive strategies based on the development of original methodologies in structural bioinformatics (link to our webtools) with a wide range of experimental techniques. Among them, NMR (nuclear magnetic resonance), biochemistry and various biophysical spectroscopy’s together with molecular biology. Each topic benefits from the expertise of both team leaders. Raphaël Guerois is mainly involved in the bioinformatics development. He initially developed the FoldX program in Luis Serrano’s lab at EMBL Heidelberg and has further specialized in the development of remote homology detection tools, in structure prediction and in the prediction and design of protein-protein interactions. Françoise Ochsenbein supervises the experimental part of the team. In particular, she is expert in the determination of protein structures and of their complexes by NMR. She previously worked in the functional design of specific protein domains derived from annexins (3 patents, creation of the Bionexis company). Through her experience she gained the skills required for the fundamental characterization of protein-protein interactions and for the design of compounds to inhibit these interactions.


Key words
Protein structure, NMR spectroscopy, bioinformatic, molecular modelling, DNA damage signalling, assembly chaperone, protein-protein interactions


Publications

Barrault MB, Richet N, Godard C, Murciano B, Le Tallec B, Rousseau E, Legrand P, Charbonnier JB, Le Du MH, Guérois R, Ochsenbein F, Peyroche A. (2012) Dual functions of the Hsm3 protein in chaperoning and scaffolding regulatory particle subunits during the proteasome assembly. Proc Natl Acad Sci U S A. [Epub ahead of print]

Jiao, Y., Seeger, K., Lautrette, A., Gaubert, A., Mousson, F., Guérois, R., Mann, C., and Ochsenbein, F. (2012) Surprising complexity of the Asf1 histone chaperone-Rad53 kinase interaction. Proc Natl Acad Sci U S A 109(8):2866-71.

Faure G, Andreani J, Guérois R. (2012). InterEvol database: exploring the structure and evolution of protein complex interfaces. Nucleic Acids Res. 40,:D847-56.

Stuttmann J, Hubberten HM, Rietz S, Kaur J, Muskett P, Guérois R, Bednarek P, Hoefgen R, Parker JE. (2011). Perturbation of Arabidopsis amino acid metabolism causes incompatibility with the adapted biotrophic pathogen Hyaloperonospora arabidopsidis. Plant Cell . 23 (7):2788-803.

Ropars V, Drevet P, Legrand P, Baconnais S, Amram J, Faure G, Márquez JA, Piétrement O, Guérois R, Callebaut I, Le Cam E, Revy P, de Villartay JP, Charbonnier JB. (2011). Structural characterization of filaments formed by human Xrcc4-Cernunnos/XLF complex involved in nonhomologous DNA end-joining. Proc Natl Acad Sci U S A 108(31):12663-8.

García-Ortíz MV, Marsin S, Arana ME, Gasparutto D, Guérois R, Kunkel TA, Radicella JP. (2011). Unexpected role for Helicobacter pylori DNA polymerase I as a source of genetic variability. PLoS Genet 7(6):e1002152.

Martin G, Burke B, Thaï R, Dey AK, Combes O, Ramos OH, Heyd B, Geonnotti AR, Montefiori DC, Kan E, Lian Y, Sun Y, Abache T, Ulmer JB, Madaoui H, Guérois R, Barnett SW, Srivastava IK, Kessler P, Martin L. (2011). Stabilization of HIV-1 envelope in the CD4-bound conformation through specific cross-linking of a CD4 mimetic. J Biol Chem 286(24):21706-16.

Aucher W, Becker E, Ma E, Miron S, Martel A, Ochsenbein F, Marsolier-Kergoat MC, Guérois R. (2010) A strategy for interaction site prediction between phospho-binding modules and their partners identified from proteomic data. Mol Cell Proteomics. 9(12):2745-59.

Fourquet S, Guérois R, Biard D, Toledano MB. (2010) Activation of NRF2 by nitrosative agents and H2O2 involves KEAP1 disulfide formation. J Biol Chem. 285, 8463-71.
Kadota Y, Shirasu K, Guérois R. (2010). NLR sensors meet at the SGT1-HSP90 crossroad. Trends BiochemSci. 35, 199-207.

Lopes A, Amarir-Bouhram J, Faure G, Petit MA, Guérois R. (2010) Detection of novel recombinases in bacteriophage genomes unveils Rad52, Rad51 and Gp2.5 remote homologs. Nucleic Acids Res. 38, 3952-62.

Malivert L, Ropars V, Nunez M, Drevet P, Miron S, Faure G, Guérois R, Mornon JP, Revy P, Charbonnier JB, Callebaut I, de Villartay JP. (2010) Delineation of the Xrcc4-interacting region in the globular head domain of cernunnos/XLF. J Biol Chem. 285, 26475-83.

Marsin S, Lopes A, Mathieu A, Dizet E, Orillard E, Guérois R, Radicella JP. (2010) Genetic dissection of Helicobacter pylori AddAB role in homologous recombination. FEMS Microbiol Lett. (Sous Presse).

Mousseau G, Raffy Q, Thomas O P, Agez M, Thai R, Renault J P, Pin S, Ochsenbein F, Cintrat J C, Rousseau B. (2010). Footprinting of Protein Interactions by Tritium Labeling. Biochemistry. 49, 4297-4299.

Cariou B, Ouguerram K, Zaïr Y, Guerois R, Langhi C, Kourimate S, Benoit I, Le May C, Gayet C, Belabbas K, Dufernez F, Chétiveaux M, Tarugi P, Krempf M, Benlian P, Costet P. (2009) PCSK9 dominant negative mutant results in increased LDL catabolic rate and familial hypobetalipoproteinemia. Arterioscler Thromb Vasc Biol. 29, 2191-7.

Le Tallec B, Barrault MB, Guérois R, Carré T, Peyroche A. (2009). Hsm3/S5b participates in the assembly pathway of the 19S regulatory particle of the proteasome. Mol Cell. 33, 389-399.

Stuttmann J, Lechner E, Guérois R, Parker JE, Nussaume L, Genschik P, Noël LD. (2009). COP9 signalosome- and 26S Proteasome-dependent regulation of SCFTIR1 accumulation in Arabidopsis. J Biol Chem. 284, 7920-2930.

Galvani A, Courbeyrette R, Agez M, Ochsenbein F, Mann C, Thuret JY. (2008). In vivo study of the nucleosome assembly functions of ASF1 histone chaperones in human cells. Mol Cell Biol. 28, 3672-85.

Kadota Y, Amigues B, Ducassou L, Madaoui H, Ochsenbein F, Guérois R and Shirasu K (2008) Structural and functional analysis of SGT1-HSP90 core complex required for innate immunity in plants. EMBO Reports. 9, 1209-1215.

Madaoui H, Guérois R (2008). Coevolution at protein complex interfaces can be detected by the complementarity trace with important impact for predictive docking. Proc Natl Acad Sci U S A. 105, 7708-13.

Marsin S, Mathieu A, Kortulewski T, Guérois R, and Radicella JP. (2008) Unveiling Novel RecO Distant Orthologues Involved In Homologous Recombination. PLoS Genetics, 4: e1000146.

Agez M, Chen J, Guérois R, van Heijenoort C, Thuret JY, Mann C, Ochsenbein F. (2007). Structure of the histone chaperone asf1 bound to the histone h3 C-terminal helix and functional insights Structure (Camb).15, 191-199.

Becker E, Cotillard A, Meyer V, Madaoui H, Guérois R (2007). HMM-Kalign : a tool for generating sub-optimal HMM alignments. Bioinformatics. 23, 3095-7.

Botër M, Amigues B, Peart J, Breuer C, Kadota Y, Casais C, Moore G, Kleanthous C, Ochsenbein F, Shirasu K, Guérois R. (2007).Structural and Functional Analysis of SGT1 Reveals That Its Interaction with HSP90 Is Required for the Accumulation of Rx, an R Protein Involved in Plant Immunity. Plant Cell. 19, 3791-804.

Guillemain G, Ma E, Mauger S, Miron S, Thai R, Guérois R, Ochsenbein F, Marsolier-Kergoat MC. (2007). Mechanisms of checkpoint kinase Rad53 inactivation after a double-strand break in Saccharomyces cerevisiae. Mol Cell Biol.27, 3378-3389.

Heise CT, Duff CS, Boter M, Casais C, Airey JE, Leech AP, Amigues B, Guérois R, Moore GR, Shirasu K, Kleanthous C. (2007). Biochemical Characterization of RAR1 Cysteine- and Histidine-Rich Domains (CHORDs): A Novel Class of Zinc-Dependent Protein-Protein Interaction Modules. Biochemistry.46, 1612-1623.

Mousson F, Ochsenbein F, Mann C. (2007). The histone chaperone Asf1 at the crossroads of chromatin and DNA checkpoint pathways. Chromosoma.116, 79-93.

Odaert B, Saida F, Aliprandi P, Durand S, Crechet JB, Guérois R, Laalami S, Uzan M, Bontems F. (2007). Structural and functional studies of RegB, a new member of a family of sequence-specific ribonucleases involved in mRNA inactivation on the ribosome. J Biol Chem.282, 2019-2028.

Le Tallec B, Barrault MB, Courbeyrette R, Guérois R, Marsolier-Kergoat MC, Peyroche A. (2007). 20S proteasome assembly is orchestrated by two distinct pairs of chaperones in yeast and in mammals. Mol Cell. 27, 660-74.

Becker E, Meyer V, Madaoui H, Guérois R. (2006). Detection of a tandem BRCT in Nbs1 and Xrs2 with functional implications in the DNA damage response. Bioinformatics.22, 1289-129.

Haubertin DY, Madaoui H, Sanson A, Guérois R, Orlowski S. (2006). Molecular dynamics simulations of E. coli MsbA transmembrane domain: formation of a semipore structure. Biophys J.91, 2517-2531.

Madaoui H, Becker E, Guérois R. (2006). Sequence search methods and scoring functions for the design of protein structures. Methods Mol Biol.340, 183-206.

Tresaugues L, Dehe PM, Guérois R, Rodriguez-Gil A, Varlet I, Salah P, Pamblanco M, Luciano P, Quevillon-Cheruel S, Sollier J, Leulliot N, Couprie J, Tordera V, Zinn-Justin S, Chavez S, van Tilbeurgh H, Geli V. (2006). Structural Characterization of Set1 RNA Recognition Motifs and their Role in Histone H3 Lysine 4 Methylation. J Mol Biol.359, 1170-1181.

Mousson F, Lautrette A, Thuret JY, Agez M, Courbeyrette R, Amigues B, Becker E, Neumann JM, Guérois R, Mann C, Ochsenbein F. (2005). Structural basis for the interaction of Asf1 with histone H3 and its functional implications Proc Natl Acad Sci U S A. 102, 5975-5980.

Charier G, Couprie J, Alpha-Bazin B, Meyer V, Quéméneur E, Guérois R, Callebaut I, Gilquin B, Zinn-Justin S. (2004). The Tudor tandem of 53BP1: a new structural motif involved in DNA and RG-rich peptide binding. Structure (Camb). 12, 1551-1562.

Mousson F, Couprie J, Thuret JY, Neumann JM, Mann C, Ochsenbein F. (2004). 1H, (13)C and (15)N Resonance Assignments of the Conserved Core of hAsf1 A. J Biomol NMR. 29, 413-414.

Leroy C, Lee SE, Vaze MB, Ochsenbein F, Guérois R, Haber JE, Marsolier-Kergoat MC. (2003). PP2C Phosphatases Ptc2 and Ptc3 Are Required for DNA Checkpoint Inactivation after a Double-Strand Break. Mol Cell. 11, 827-835.

Micheelsen MA, Rischel C, Borg J, Guérois R, Serrano L. (2003) Mean first-passage time analysis reveals rate-limiting steps, parallel pathways and dead ends in a simple model of protein folding. Europhys. Let. 61,561-66.

Barth P, Savarin P, Gilquin B, Lagoutte B, Ochsenbein F. (2002). Solution NMR Structure and Backbone Dynamics of the PsaE Subunit of Photosystem I from Synechocystis sp. PCC 6803 Biochemistry. 41, 13902-13914.

Dubacq C, Guérois R, Courbeyrette R, Kitagawa K, Mann C. (2002). Sgt1p contributes to cyclic AMP pathway activity and physically interacts with theadenylyl cyclase Cyr1p/Cdc35p in budding yeast. Eukaryot Cell. 1, 568-582.

Guérois R, Nielsen JE, Serrano L. (2002). Predicting changes in the stability of proteins and protein complexes: a study of more than 1000 mutations. J Mol Biol. 320, 369-387. Guérois R, Mendes J, Serrano L. (2002). Energy estimation in protein design. Curr Opin Struct Biol. 12, 441-446.

Montaville P, Neumann JM, Russo-Marie F, Ochsenbein F, Sanson A. (2002). A new consensus sequence for phosphatidylserine recognition by annexins. J Biol Chem. 277, 24684-24693.

Ochsenbein F, Neumann JM, Guittet E, van Heijenoort C. (2002). Dynamical characterization of residual and non native structures in a partially folded protein by 15N-NMR relaxation using a model based on a distribution of correlation times. Protein Sci. 11, 957-964.

Ochsenbein F, Guérois R, Neumann JM, Sanson A, Guittet E, Van Heijenoort C. (2001). 15N NMR relaxation as a probe for helical intrinsic propensity : the case of the unfolded D2 domain of annexin I J Biomol NMR. 19, 3-18.

Guérois R, Serrano L. (2001). Protein design based on folding models. Curr Opin Struct Biol.11,101-6.

Guérois R, Serrano L. (2000). The SH3-fold family: experimental evidence and prediction of variations in the folding pathways. J Mol Biol. 304,967-82.

Beswick V, Guérois R, Cordier-Ochsenbein F, Coic YM, Tam HD, Tostain J, Noël JP, Sanson A, Neumann JM. (1999). Dodecylphosphocholine micelles as a membrane-like environment : new results from NMR relaxation and paramagnetic relaxation enhancement analysis Eur Biophys J. 28, 48-58.

Cordier-Ochsenbein F, Guérois R, Russo-Marie F, Neumann JM, Sanson A. (1998). Exploring the folding pathways of annexin I, a multidomain protein. II . Hierarchy in domain folding propensities may govern the folding process J Mol Biol. 279, 1177-1185.

Cordier-Ochsenbein F, Guérois R, Baleux F, Huynh-Dinh T, Lirsac PN, Russo-Marie F, Neumann JM, Sanson A. (1998). Exploring the folding pathways of annexin I, a multidomain protein. I. Non-native structures stabilize the partially folded state of the isolated domain 2 of annexin I J Mol Biol. 279, 1163-1175.

Guérois R, Cordier-Ochsenbein F, Baleux F, Huynh-Dinh T, Neumann JM, Sanson A. (1998). A conformational equilibrium in a protein fragment caused by two consecutive capping boxes: 1H-, 13C-NMR, and mutational analysis. Protein Sci. 7, 1506-1515.

Agez, M., Lautrette, A., Mousson, F., Mann, C., and Ochsenbein, F. (2006). Structural biology of histone associated proteins, Chromatin: structure and function, Research Signpost, Kerala, India

Guérois R, Mendes J, Serrano L. (2004). Predicting the effect of mutations on protein stability in "Handbook of Protein Folding" Wiley/Verlag Chemie Ed. Brevet Français étendu à l’Europe et aux US.

Sanson A., Russo-Marie F., Neumann J.M., Cordier-Ochsenbein F., Guérois R. Structure chimique ayant une affinité pour un phospholipide, et composé de marquage, trousse de diagnostic, et médicament comprenant cette structure. N°98-12366 au nom du CEA et Université Paris VI. (1998).

Brevet Français : Sanson A., Ochsenbein F. Peptides ayant une affinité pour un phospholipide et utilisations. N° 02 08202 au nom du CEA et de l’Université Paris VI. (2002).

Brevet Français : Sanson A., Ochsenbein F., Dollé F. Peptides marqués ayant une affinité pour un phospholipide et utilisations. N° 0208204 au nom du CEA et de l’Université Paris VI. (2002).