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8 results on '"Université Laval [Québec] (ULaval)-CHUQ Research Center"'

1. Effects of Long-term Serial Passaging on the Characteristics and Properties of Cell Lines Derived From Uveal Melanoma Primary Tumors

Author

Arnaud de la Fouchardière, Marjorie-Allison Bergeron, Frédéric Fournier, Arnaud Droit, Mohib W. Morcos, Karine Zaniolo, Solange Landreville, Sylvain L. Guérin, Frédéric Mouriaux, Cindy Weidmann, CHU Pontchaillou [Rennes], Université de Rennes - Faculté de Médecine (UR Médecine), Université de Rennes (UR), CHU de Québec, Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, Natural Sciences and Engineering Research Council of Canada (NSERC) [138624-2012], Fondation du CHU de Quebec, Fonds de Recherche du Quebec - Sante (FRQS), Vision Health Research Network (VHRN), Faculte de medecine de l'Universite Laval, Centre de recherche en organogenese experimentale de l'Universite Laval/LOEX, Université de Rennes 1 - Faculté de Médecine (UR1 Médecine), Université de Rennes 1 (UR1), and Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)

Subjects
Uveal Neoplasms, 0301 basic medicine, short tandem repeat, Blotting, Western, Cell, Mice, Nude, Cell Count, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, Polymerase Chain Reaction, Metastasis, Mice, 03 medical and health sciences, models, MART-1 Antigen, Cancer stem cell, Cell Line, Tumor, imatinib mesylate, establishment, medicine, Animals, Humans, metastasis, Microscopy, Phase-Contrast, RNA, Neoplasm, ocular melanoma, Melanoma, cancer stem-cells, Gene Expression Profiling, human choroidal melanoma, Neoplasms, Experimental, Gene signature, [SDV.MHEP.EM]Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism, medicine.disease, Immunohistochemistry, gene-expression, 3. Good health, Gene Expression Regulation, Neoplastic, Gene expression profiling, 030104 developmental biology, Imatinib mesylate, medicine.anatomical_structure, xenografts, Cell culture, Cancer research, Female, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition
Abstract

International audience; PURPOSE. Development of liver metastasis remains the most common cause of mortality in uveal melanoma (UM). A few cell lines cultured from primary UM tumors have been used widely to investigate the pathobiology of UM. However, the translation of basic knowledge to the clinic for the treatment of the metastatic disease has remained incremental at best. In this study, we examined whether the properties of UM cell lines at various passages were similar to their corresponding primary tumors. METHODS. Gene expression profiling by microarray was performed on UM primary tumors and derived cell lines cultured at varying passages. Expression of UM protein markers was monitored by immunohistochemical analyses and Western blotting. The in vivo tumorigenic properties of UM cultures were evaluated using athymic nude mice. RESULTS. Cell passaging severely reduced the expression of genes encoding markers typical of UM, including those of the prognostic gene signature. Marked differences between gene expression profiles of primary tumors and cell lines could be linked to the infiltrating immune and stromal cells in situ. In addition, the tumorigenic properties of UM cell lines also increased with cell passaging in culture as evaluated by their subcutaneous injection into athymic mice. CONCLUSIONS. Together, these findings demonstrate that the short-term UM primary cultures exhibit molecular features that resemble the respective surgical material and, thus, represent the best model for in vitro-assessed cancer treatments.

Published
2016

2. Proteomics reveals a switch in CDK1-associated proteins upon M-phase exit during the Xenopus laevis oocyte to embryo transition

Author

Jean-Philippe Gagné, Jacek Z. Kubiak, Gaëlle Marteil, Laurent Richard-Parpaillon, Ewa Borsuk, Guy G. Poirier, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS), Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, Department of Embryology [Warsaw], Institute of Zoology [Warsaw], Faculty of Biology [Warsaw], University of Warsaw (UW)-University of Warsaw (UW)-Faculty of Biology [Warsaw], University of Warsaw (UW)-University of Warsaw (UW), The authors were supported by grants from Ligue Contre le Cancer and Association pour la Recherche contre le Cancer to JZK. GM was a recipient of a fellowship from the French Ministère de la Recherche et de l'Enseignement supérieur and MED from the Lebanese Government., Centre National de la Recherche Scientifique (CNRS)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS), and De Villemeur, Hervé

Subjects
Proteomics, Embryo, Nonmammalian, Cyclin B, Xenopus, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, environment and public health, Biochemistry, MESH: Oocytes, Xenopus laevis, 03 medical and health sciences, 0302 clinical medicine, MESH: Xenopus laevis, Cyclin-dependent kinase, CDC2 Protein Kinase, [SDV.BDD] Life Sciences [q-bio]/Development Biology, medicine, Animals, Humans, MESH: Animals, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, [SDV.BDD]Life Sciences [q-bio]/Development Biology, 030304 developmental biology, 0303 health sciences, Cyclin-dependent kinase 1, MESH: Humans, biology, MESH: Proteomics, MESH: Embryo, Nonmammalian, Oocyte activation, Cell Biology, Cell cycle, biology.organism_classification, Oocyte, MESH: CDC2 Protein Kinase, Cell biology, enzymes and coenzymes (carbohydrates), medicine.anatomical_structure, Oocytes, biology.protein, MESH: Cell Division, Female, biological phenomena, cell phenomena, and immunity, MESH: Female, Cell Division, 030217 neurology & neurosurgery
Abstract

International audience; Cyclin-dependent kinase 1 (CDK1) is a major M-phase kinase which requires the binding to a regulatory protein, Cyclin B, to be active. CDK1/Cyclin B complex is called M-phase promoting factor (MPF) for its key role in controlling both meiotic and mitotic M-phase of the cell cycle. CDK1 inactivation is necessary for oocyte activation and initiation of embryo development. This complex process requires both Cyclin B polyubiquitination and proteosomal degradation via the ubiquitin-conjugation pathway, followed by the dephosphorylation of the monomeric CDK1 on Thr161. Previous proteomic analyses revealed a number of CDK1-associated proteins in human HeLa cells. It is, however, unknown whether specific partners are involved in CDK1 inactivation upon M-phase exit. To better understand CDK1 regulation during MII-arrest and oocyte activation, we immunoprecipitated (IPed) CDK1 together with its associated proteins from M-phase-arrested and M-phase-exiting Xenopus laevis oocytes. A mass spectrometry (MS) analysis revealed a number of new putative CDK1 partners. Most importantly, the composition of the CDK1-associated complex changed rapidly during M-phase exit. Additionally, an analysis of CDK1 complexes precipitated with beads covered with p9 protein, a fission yeast suc1 homologue well known for its high affinity for CDKs, was performed to identify the most abundant proteins associated with CDK1. The screen was auto-validated by identification of: (i) two forms of CDK1: Cdc2A and B, (ii) a set of Cyclins B with clearly diminishing number of peptides identified upon M-phase exit, (iii) a number of known CDK1 substrates (e.g. peroxiredoxine) and partners (e.g. HSPA8, a member of the HSP70 family) both in IP and in p9 precipitated pellets. In IP samples we also identified chaperones, which can modulate CDK1 three-dimensional structure, as well as calcineurin, a protein necessary for successful oocyte activation. These results shed a new light on CDK1 regulation via a dynamic change in the composition of the protein complex upon M-phase exit and the oocyte to embryo transition.

Published
2012

3. Ensconsin/Map7 promotes microtubule growth and centrosome separation in Drosophila neural stem cells

Author

Renaud Caous, Régis Giet, Guy G. Poirier, Jean-Philippe Gagné, Denis Chrétien, Sébastien Huet, Laurent Richard-Parpaillon, Emmanuel Gallaud, Aude Pascal, Franck Bazile, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, This work was funded by the Ligue Nationale Contre le Cancer, l'Association de la Recherche contre le Cancer, Biosit, and the University of Rennes 1. This work was supported by the Canadian Institutes of Health Research (grant No. MO-178013 to GGP). The authors wish to thank the Proteomics Platform of Quebec Genomic Center for analytical services and the Microscopy Rennes Imaging Center. G. G. Poirier holds a Canada Research Chair in Proteomics. E. Gallaud and R. Caous are doctoral fellows of the Region Bretagne and the French Ministry of Research, respectively., and Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )

Subjects
genetic structures, Microtubule-associated protein, ASYMMETRIC DIVISION, Kinesins, Mitosis, PROTEIN, STATISTICAL-MODEL, macromolecular substances, Spindle Apparatus, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, KINETOCHORE FIBERS, TRANSGENIC RNAI, Microtubules, Time-Lapse Imaging, Spindle pole body, NEUROBLASTS, Microtubule polymerization, Neural Stem Cells, Microtubule, Report, Chromosome Segregation, Animals, Drosophila Proteins, GENOME-WIDE ANALYSIS, Interphase, Research Articles, Cells, Cultured, Microtubule nucleation, Centrosome, ANTAGONISTIC ACTIVITIES, Cell Biology, Cell biology, Spindle apparatus, Drosophila melanogaster, METAPHASE SPINDLE, Microscopy, Fluorescence, MITOTIC SPINDLE, Protein Multimerization, Centrosome separation, Microtubule-Associated Proteins
Abstract

Ensconsin cooperates with its binding partner, Kinesin-1, during interphase to trigger centrosome separation, but it promotes microtubule polymerization independently of Kinesin-1 to control spindle length during mitosis., The mitotic spindle is crucial to achieve segregation of sister chromatids. To identify new mitotic spindle assembly regulators, we isolated 855 microtubule-associated proteins (MAPs) from Drosophila melanogaster mitotic or interphasic embryos. Using RNAi, we screened 96 poorly characterized genes in the Drosophila central nervous system to establish their possible role during spindle assembly. We found that Ensconsin/MAP7 mutant neuroblasts display shorter metaphase spindles, a defect caused by a reduced microtubule polymerization rate and enhanced by centrosome ablation. In agreement with a direct effect in regulating spindle length, Ensconsin overexpression triggered an increase in spindle length in S2 cells, whereas purified Ensconsin stimulated microtubule polymerization in vitro. Interestingly, ensc-null mutant flies also display defective centrosome separation and positioning during interphase, a phenotype also detected in kinesin-1 mutants. Collectively, our results suggest that Ensconsin cooperates with its binding partner Kinesin-1 during interphase to trigger centrosome separation. In addition, Ensconsin promotes microtubule polymerization during mitosis to control spindle length independent of Kinesin-1.

Published
2014

4. Proteomic investigation of phosphorylation sites in poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase

Author

Mélissa Chevalier-Paré, Jean-Philippe Gagné, Sylvie Bourassa, Arnaud Droit, Claude Prigent, Yves Labelle, Darin McDonald, Xavier Moreel, Guy G. Poirier, Michael J. Hendzel, Pierre Gagné, De Villemeur, Hervé, Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS), Department of Oncology, University of Alberta, Proteomics Platform, Québec Genomic Center-Laval University Medical Research Center, Université de Rennes 1 (UR1), and Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Phosphopeptides, MESH: Signal Transduction, Poly (ADP-Ribose) Polymerase-1, MESH: Amino Acid Sequence, Biochemistry, environment and public health, Mass Spectrometry, MESH: Recombinant Proteins, MESH: Protein Structure, Tertiary, 0302 clinical medicine, Protein phosphorylation, MESH: Animals, Phosphorylation, Threonine, 0303 health sciences, PARG, Mitogen-Activated Protein Kinase 3, Recombinant Proteins, MESH: Reproducibility of Results, 030220 oncology & carcinogenesis, Poly(ADP-ribose) Polymerases, Signal transduction, MESH: Mitogen-Activated Protein Kinase 3, Signal Transduction, inorganic chemicals, Glycoside Hydrolases, DNA damage, Poly ADP ribose polymerase, Molecular Sequence Data, MESH: Phosphopeptides, macromolecular substances, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, 03 medical and health sciences, MESH: Glycoside Hydrolases, Animals, Humans, Amino Acid Sequence, Poly(ADP-ribose) glycohydrolase, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, 030304 developmental biology, MESH: DNA Damage, MESH: Mass Spectrometry, MESH: Humans, MESH: Molecular Sequence Data, MESH: Phosphorylation, MESH: Poly(ADP-ribose) Polymerases, Reproducibility of Results, General Chemistry, Protein Structure, Tertiary, enzymes and coenzymes (carbohydrates), bacteria, DNA Damage
Abstract

International audience; Phosphorylation is a very common post-translational modification event known to modulate a wide range of biological responses. Beyond the regulation of protein activity, the interrelation of phosphorylation with other post-translational mechanisms is responsible for the control of diverse signaling pathways. Several observations suggest that phosphorylation of poly(ADP-ribose) polymerase-1 (PARP-1) regulates its activity. There is also accumulating evidence to suggest the establishment of phosphorylation-dependent assembly of PARP-1-associated multiprotein complexes. Although it is relatively straightforward to demonstrate phosphorylation of a defined target, identification of the actual amino acids involved still represents a technical challenge for many laboratories. With the use of a combination of bioinformatics-based predictions tools for generic and kinase-specific phosphorylation sites, in vitro phosphorylation assays and mass spectrometry analysis, we investigated the phosphorylation profile of PARP-1 and poly(ADP-ribose) glycohydrolase (PARG), two major enzymes responsible for poly(ADP-ribose) turnover. Mass spectrometry analysis revealed the phosphorylation of several serine/threonine residues within important regulatory domains and motifs of both enzymes. With the use of in vivo microirradiation-induced DNA damage, we show that altered phosphorylation at specific sites can modify the dynamics of assembly and disassembly of PARP-1 at sites of DNA damage. By documenting and annotating a collection of known and newly identified phosphorylation sites, this targeted proteomics study significantly advances our understanding of the roles of phosphorylation in the regulation of PARP-1 and PARG.

Published
2009

5. Proteome-wide identification of poly(ADP-ribose) binding proteins and poly(ADP-ribose)-associated protein complexes

Author

Guy G. Poirier, Valina L. Dawson, Sylvie Bourassa, Ted M. Dawson, Jean-Philippe Gagné, Michael J. Hendzel, Ken Sin Lo, Maxim Isabelle, Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS), Department of Oncology, University of Alberta, Cross Cancer Institute, Department of Biology, Johns Hopkins University (JHU), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS), and De Villemeur, Hervé

Subjects
Poly Adenosine Diphosphate Ribose, Proteome, DNA repair, Immunoprecipitation, DNA damage, In silico, Amino Acid Motifs, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Models, Biological, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Tandem Mass Spectrometry, Cell Line, Tumor, Ribose, Genetics, Humans, Electrophoresis, Gel, Two-Dimensional, Poly-ADP-Ribose Binding Proteins, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Molecular Biology, 030304 developmental biology, 0303 health sciences, Adenosine diphosphate ribose, Proteins, Biochemistry, chemistry, 030220 oncology & carcinogenesis, Carrier Proteins, Chromatography, Liquid, DNA Damage, HeLa Cells
Abstract

International audience; Poly(ADP-ribose) (pADPr) is a polymer assembled from the enzymatic polymerization of the ADP-ribosyl moiety of NAD by poly(ADP-ribose) polymerases (PARPs). The dynamic turnover of pADPr within the cell is essential for a number of cellular processes including progression through the cell cycle, DNA repair and the maintenance of genomic integrity, and apoptosis. In spite of the considerable advances in the knowledge of the physiological conditions modulated by poly(ADP-ribosyl)ation reactions, and notwithstanding the fact that pADPr can play a role of mediator in a wide spectrum of biological processes, few pADPr binding proteins have been identified so far. In this study, refined in silico prediction of pADPr binding proteins and large-scale mass spectrometry-based proteome analysis of pADPr binding proteins were used to establish a comprehensive repertoire of pADPr-associated proteins. Visualization and modeling of these pADPr-associated proteins in networks not only reflect the widespread involvement of poly(ADP-ribosyl)ation in several pathways but also identify protein targets that could shed new light on the regulatory functions of pADPr in normal physiological conditions as well as after exposure to genotoxic stimuli.

Published
2008

6. Differential proteomic screen to evidence proteins ubiquitinated upon mitotic exit in cell-free extract of Xenopus laevis embryos

Author

Jacek Z. Kubiak, Aude Pascal, Guy G. Poirier, Jean-Philippe Gagné, Julian Vasilescu, Franck Bazile, Franck Chesnel, Daniel Figeys, Geneviève Mercier, Ken Sin Lo, Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS), Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, Ottawa Institute of Systems Biology, University of Ottawa [Ottawa], De Villemeur, Hervé, Université de Rennes 1 (UR1), and Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Proteomics, Embryo, Nonmammalian, Proteome, Cell, Xenopus, Xenopus Proteins, Biochemistry, law.invention, MESH: Recombinant Proteins, Xenopus laevis, MESH: Histidine, Ubiquitin, law, MESH: Cell-Free System, MESH: Animals, MESH: Xenopus Proteins, 0303 health sciences, MESH: Proteomics, 030302 biochemistry & molecular biology, Cell cycle, Recombinant Proteins, MESH: Proteome, medicine.anatomical_structure, Recombinant DNA, Female, MESH: Ubiquitin, Mitosis, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, 03 medical and health sciences, Affinity chromatography, MESH: Xenopus laevis, medicine, Animals, Histidine, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, 030304 developmental biology, Cell-Free System, Ubiquitination, MESH: Embryo, Nonmammalian, General Chemistry, MESH: Mitosis, biology.organism_classification, Mitotic exit, biology.protein, MESH: Ubiquitination, MESH: Female
Abstract

International audience; Post-translational modification of proteins via ubiquitination plays a crucial role in numerous vital functions of the cell. Polyubiquitination is one of the key regulatory processes involved in regulation of mitotic progression. Here we describe a differential proteomic screen dedicated to identification of novel proteins ubiquitinated upon mitotic exit in cell-free extract of Xenopus laevis embryo. Mutated recombinant His6-tagged ubiquitin (Ubi (K48R)) was added to mitotic extract from which we purified conjugated proteins, as well as associated proteins in nondenaturing conditions by cobalt affinity chromatography. Proteins eluted from Ubi (K48R) supplemented and control extracts were compared by LC-MS/MS analysis after monodimensional SDS-PAGE. A total of 144 proteins potentially ubiquitinated or associated with them were identified. Forty-one percent of these proteins were shown to be involved in ubiquitination and/or proteasomal degradation pathway confirming the specificity of the screen. Twelve proteins, among them ubiquitin itself, were shown to carry a "GG" or "LRGG" remnant tag indicating their direct ubiquitination. Interestingly, sequence analysis of ubiquitinated substrates carrying these tags indicated that in Xenopus cell-free embryo extract supplemented with Ubi (K48R) the majority of polyubiquitination occurred through lysine-11 specific ubiquitin chain polymerization. The potential interest in this atypical form of ubiquitination as well as usefulness of our method in analyzing atypical polyubiquitin species is discussed.

Published
2008

7. Human Proteinpedia enables sharing of human protein data

Author

Ron Bose, Jean-Charles Sanchez, Young Jin Lee, Marcus Bantscheff, Pia Hønnerup Jensen, Yunping Zhu, Jonathan C. Trinidad, Juan Martínez-Heredia, Michael Moran, Samir K. Brahmachari, Pierre Gagné, Kripa Shankari, Jeffrey C. Smith, Jose-Manuel Vidal-Taboada, James P. DeLany, Shi Jun Sheng, Ragna Rönnholm, Xosé R. Bustelo, Helene L. Cardasis, Erik Björling, Ole N. Jensen, Pavel Gromov, Michael J. Dunn, Xiaoyue Wang, Guy G. Poirier, Greg T. Cantin, Richard J. Simpson, Kenny Helsens, Ming Zhou, Sumio Sugano, Samir M. Hanash, Prashantha Hebbar, Y. L. Ramachandra, Jennifer E. Van Eyk, Rafael Oliva, Philip C. Andrews, Lennart Martens, Julio E. Celis, B. Abdul Rahiman, Alexander Mehrle, Feixia Chu, Richard D. Smith, Philip A. Cole, Leroi V. DeSouza, Stefan Wiemann, Joseph A. Loo, Bernhard Kuster, Mauno Vihinen, Peter Jung, David C. Muddiman, Jayson A. Falkner, Osamu Ohara, Fredrik Levander, Gerald W. Hart, Mukhtar Ahmed, T. S. Keshava Prasad, Eric W. Deutsch, Riaz Mohmood, Indu Kheterpal, Jeffrey M. Gimble, John R. Yates, Catherine Fenselau, Timothy D. Veenstra, Julian Vasilescu, Brian M. Balgley, Heiko Hermeking, Johanna Salonen, Rainer Pepperkok, Michael Lefevre, William S. Hancock, Visith Thongboonkerd, Tao Xu, Beerelli Seshi, Christine A. Miller, Florian Gnad, Ravi Sirdeshmukh, Arnaud Droit, Renu Goel, Maarit Takatalo, Emanuel F. Petricoin, Mathias Uhlén, Vitor M. Faça, Billy Wu, Robert J. Cotter, Angelo M. De Marzo, Mark E. McComb, Alma S Burlingame, Oliver Hofmann, Martine Morzel, Rajasree Menon, Denis F. Hochstrasser, Peter James, Matthew J. Sullivan, Robin Wait, K. W. Michael Siu, H. C. Harsha, Hainard Alexandre, Megan S. Lim, Winston Hide, Kris Gevaert, Harald Mischak, Thierry Sayd, Matthias Mann, Blagoy Blagoev, Gerard Cagney, Xiangming Fang, Ralph H. Hruban, James D. Morgan, Joel S. Bader, Samuel O. Purvine, Fuchu He, Robert Moritz, Rob M. Ewing, Daniel Figeys, Min-Seok Kwon, Kumaran Kandasamy, Reiko F. Kikuno, Masaaki Oyama, Cecilia Gelfi, Gilbert S. Omenn, James P. McRedmond, Pierre Lescuyer, Kojo S.J. Elenitoba-Johnson, Akhilesh Pandey, Joël Vandekerckhove, Karin Hjernø, Subburaman Mohan, Jens Rick, Kyla Pennington, Raghothama Chaerkady, Henrik Molina, David M Horn, Faith A. Hays, Young Ki Paik, Balamurugan Periaswamy, Giulio Superti-Furga, Roman Körner, Gerard Drewes, Jun Zhong, E. Dransfield, Suresh Mathivanan, Robert H. Rice, David K. Crockett, Thomas A. Neubert, Minna Lehvaslaiho, K. Shivakumar, Catherine E. Costello, Hyoung Joo Lee, Christian Löbke, Keiryn L. Bennett, Nieves Ibarrola, Ramars Amanchy, Petra Zürbig, Vivekananda Shetty, Natalie G. Ahn, Ulrike Korf, J. Daniel Navarro, Anuradha Nalli, Prasanna Ramachandran, David J. States, Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Biological Systems Engineering, University of Wisconsin-Madison, CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria, Royal Institute of Technology [Stockholm] (KTH ), University of California [San Francisco] (UC San Francisco), University of California (UC), Boston University School of Medicine (BUSM), Boston University [Boston] (BU), Radiothérapie moléculaire (UMR 1030), Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Qualité des Produits Animaux (QuaPA), Institut National de la Recherche Agronomique (INRA), Université Laval [Québec] (ULaval), British Antarctic Survey (BAS), Natural Environment Research Council (NERC), Chemistry and Biochemistry Department (University of Maryland), University of Maryland [College Park], University of Maryland System-University of Maryland System, Ottawa Institute of Systems Biology, University of Ottawa [Ottawa], Cabinet de Médecine Générale (Cabinet PG), Ville, Flanders Institute for Biotechnology, Beijing Institute of Radiation medicine, South African National Bioinformatics Institute (SANBI), University of the Western Cape (UWC), Département de science des protéines humaines [Genève], Université de Genève = University of Geneva (UNIGE)-Faculté de médecine [Genève], Donders Institute for Brain, Cognition and Behaviour, Radboud University [Nijmegen], Fraunhofer German-Sino Lab for Mobile Communications (MCI), Fraunhofer Institute for Manufacturing Engineering and Automation (Fraunhofer IPA), Fraunhofer (Fraunhofer-Gesellschaft)-Fraunhofer (Fraunhofer-Gesellschaft), Max Planck Institute for Human Cognitive and Brain Sciences [Leipzig] (IMPNSC), Max-Planck-Gesellschaft, Technische Universität Munchen - Université Technique de Munich [Munich, Allemagne] (TUM), Hôpitaux Universitaires de Genève (HUG), Department of Pathology, University of Utah, University of California [Los Angeles] (UCLA), Universiteit Gent = Ghent University (UGENT), Protein Analysis for Clinical Diagnosis and Pharmaceutical Research, Mosaiques Diagnostics and Therapeutics AG, Proteomics Resource Center, Rockfeller University, Auteur indépendant, Section des Sciences de la Terre, Université de Genève = University of Geneva (UNIGE), RIKEN Research Center for Allergy and Immunology, Yokohama (RCAI), Research Unit for Immune Homeostasis, Yonsei Proteome Research Center and Department of Biochemistry, Yonsei University, European Molecular Biology Laboratory [Heidelberg] (EMBL), Department of Molecular Pathology and Microbiology, Center for Applied Proteomics and Molecular Medicine-George Mason University [Fairfax], Axe cancer, Université Laval [Québec] (ULaval)-CHUQ Research Center, The University of Arizona Medical Center, University of Arizona, The University of Tokyo (UTokyo), Research Center for Molecular Medicine of the Austrian Academy of Sciences [Vienna, Austria] (CeMM ), Austrian Academy of Sciences (OeAW), Faculty of Biological and Environmental Sciences [Helsinki], Helsingin yliopisto = Helsingfors universitet = University of Helsinki, Medical Molecular Biology Unit, Mahidol University [Bangkok], National Cancer Institute [Bethesda] (NCI-NIH), National Institutes of Health [Bethesda] (NIH), Department of Experimental Medical Science, Lund University [Lund], Tsinghua University [Beijing] (THU), Deutsches Krebsforschungszentrum, Institut d’Électronique, de Microélectronique et de Nanotechnologie - UMR 8520 (IEMN), Centrale Lille-Institut supérieur de l'électronique et du numérique (ISEN)-Université de Valenciennes et du Hainaut-Cambrésis (UVHC)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Université Polytechnique Hauts-de-France (UPHF), Institut Européen des membranes (IEM), Université Montpellier 2 - Sciences et Techniques (UM2)-Ecole Nationale Supérieure de Chimie de Montpellier (ENSCM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Mosaiques Diagnostics & Therapeutics AG, University of California [San Francisco] (UCSF), University of California, Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Gustave Roussy (IGR)-Université Paris-Sud - Paris 11 (UP11), University of the Western Cape, Université de Genève (UNIGE)-Faculté de médecine [Genève], Radboud university [Nijmegen], Universiteit Gent = Ghent University [Belgium] (UGENT), University of Geneva [Switzerland], University of Helsinki, Université Montpellier 2 - Sciences et Techniques (UM2)-Ecole Nationale Supérieure de Chimie de Montpellier (ENSCM)-Institut de Chimie du CNRS (INC)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Hochstrasser, Denis, Lescuyer, Pierre, Moritz, Robert, Pepperkok, Rainer, and Sanchez, Jean-Charles

Subjects
Proteomics, Proteomics methods, Internationality, Magnetic Resonance Spectroscopy, Proteome, Information storage, [SDV]Life Sciences [q-bio], Biomedical Engineering, Information Storage and Retrieval, Proteome/*chemistry/classification/*metabolism, Bioengineering, Computational biology, Biology, Bioinformatics, Databases, Protein, Applied Microbiology and Biotechnology, Data type, Peptide Mapping, 03 medical and health sciences, User-Computer Interface, ddc:576, ComputingMilieux_MISCELLANEOUS, Database Management Systems, Magnetic Resonance Spectroscopy/methods, 030304 developmental biology, 0303 health sciences, Internet, business.industry, Peptide mapping, Gene Expression Profiling, 030302 biochemistry & molecular biology, Information Storage and Retrieval/*methods, Peptide Mapping/methods, Gene Expression Profiling/*methods, Molecular Medicine, The Internet, Proteomics/methods, business, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition, Software, Biotechnology
Abstract

Proteomic technologies, such as yeast two-hybrid, mass spectrometry (MS), protein/peptide arrays and fluorescence microscopy, yield multi-dimensional data sets, which are often quite large and either not published or published as supplementary information that is not easily searchable. Without a system in place for standardizing and sharing data, it is not fruitful for the biomedical community to contribute these types of data to centralized repositories.

Published
2008

8. Expression, purification, crystallization and preliminary X-ray studies of the outer membrane efflux proteins OprM and OprN from Pseudomonas aeruginosa

Author

Isabelle Broutin, Marie-Bernard Lascombe, Arnaud Ducruix, Houssain Benabdelhak, Dimitri Lerouge, Xavier Moreel, Laboratoire de cristallographie et RMN biologiques (LCRB - UMR 8015), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5), Axe cancer, Université Laval-CHUQ Research Center, Université Paris Descartes - Paris 5 (UPD5)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Université Laval [Québec] (ULaval)-CHUQ Research Center

Subjects
[SDV]Life Sciences [q-bio], Molecular Sequence Data, Biophysics, Biology, medicine.disease_cause, Crystallography, X-Ray, Biochemistry, Models, Biological, Polymerase Chain Reaction, 03 medical and health sciences, Affinity chromatography, Structural Biology, Genetics, medicine, Outer membrane efflux proteins, Escherichia coli, 030304 developmental biology, DNA Primers, 0303 health sciences, Base Sequence, 030306 microbiology, Membrane transport protein, Pseudomonas aeruginosa, Membrane Transport Proteins, biochemical phenomena, metabolism, and nutrition, Condensed Matter Physics, biology.organism_classification, Recombinant Proteins, Crystallization Communications, biology.protein, Efflux, Bacterial outer membrane, Crystallization, Bacteria, Bacterial Outer Membrane Proteins
Abstract

OprM and OprN belong to the outer membrane factor family proteins. These approximately 52 kDa proteins are part of the tripartite efflux pumps found in Pseudomonas aeruginosa and are responsible in part for the antibiotic resistance observed in these bacteria. Both proteins have been expressed in Escherichia coli as His-tag proteins and purified accordingly by affinity chromatography in the presence of n-octyl-beta-D-glucopyranoside detergent. OprM and OprN were crystallized using PEG 20 000/ammonium citrate and ammonium sulfate as precipitating agents, respectively. Crystals belong to space group C2, with unit-cell parameters a = 152.6, b = 87.9, c = 355.9 A, beta = 98.9 degrees and a = 151.3, b = 87.6, c = 356.5 A, beta = 98.1 degrees for OprM and OprN, respectively. Using the ESRF synchrotron-radiation source, OprM diffraction data extended to 3.4 A.

Published
2005

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