40 results on '"Corinne Vivès"'
Search Results
2. Identification of a two-component regulatory system involved in antimicrobial peptide resistance in Streptococcus pneumoniae.
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Aissatou Maty Diagne, Anaïs Pelletier, Claire Durmort, Agathe Faure, Kerstin Kanonenberg, Céline Freton, Adeline Page, Frédéric Delolme, Jaroslav Vorac, Sylvain Vallet, Laure Bellard, Corinne Vivès, Franck Fieschi, Thierry Vernet, Patricia Rousselle, Sébastien Guiral, Christophe Grangeasse, Jean-Michel Jault, and Cédric Orelle
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
Two-component regulatory systems (TCS) are among the most widespread mechanisms that bacteria use to sense and respond to environmental changes. In the human pathogen Streptococcus pneumoniae, a total of 13 TCS have been identified and many of them have been linked to pathogenicity. Notably, TCS01 strongly contributes to pneumococcal virulence in several infection models. However, it remains one of the least studied TCS in pneumococci and its functional role is still unclear. In this study, we demonstrate that TCS01 cooperates with a BceAB-type ABC transporter to sense and induce resistance to structurally-unrelated antimicrobial peptides of bacterial origin that all target undecaprenyl-pyrophosphate or lipid II, which are essential precursors of cell wall biosynthesis. Even though tcs01 and bceAB genes do not locate in the same gene cluster, disruption of either of them equally sensitized the bacterium to the same set of antimicrobial peptides. We show that the key function of TCS01 is to upregulate the expression of the transporter, while the latter appears the main actor in resistance. Electrophoretic mobility shift assays further demonstrated that the response regulator of TCS01 binds to the promoter region of the bceAB genes, implying a direct control of these genes. The BceAB transporter was overexpressed and purified from E. coli. After reconstitution in liposomes, it displayed substantial ATPase and GTPase activities that were stimulated by antimicrobial peptides to which it confers resistance to, revealing new functional features of a BceAB-type transporter. Altogether, this inducible defense mechanism likely contributes to the survival of the opportunistic microorganism in the human host, in which competition among commensal microorganisms is a key determinant for effective host colonization and invasive path.
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- 2022
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3. DC/L-SIGN recognition of spike glycoprotein promotes SARS-CoV-2 trans-infection and can be inhibited by a glycomimetic antagonist.
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Michel Thépaut, Joanna Luczkowiak, Corinne Vivès, Nuria Labiod, Isabelle Bally, Fátima Lasala, Yasmina Grimoire, Daphna Fenel, Sara Sattin, Nicole Thielens, Guy Schoehn, Anna Bernardi, Rafael Delgado, and Franck Fieschi
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
The efficient spread of SARS-CoV-2 resulted in a unique pandemic in modern history. Despite early identification of ACE2 as the receptor for viral spike protein, much remains to be understood about the molecular events behind viral dissemination. We evaluated the contribution of C-type lectin receptors (CLRS) of antigen-presenting cells, widely present in respiratory mucosa and lung tissue. DC-SIGN, L-SIGN, Langerin and MGL bind to diverse glycans of the spike using multiple interaction areas. Using pseudovirus and cells derived from monocytes or T-lymphocytes, we demonstrate that while virus capture by the CLRs examined does not allow direct cell infection, DC/L-SIGN, among these receptors, promote virus transfer to permissive ACE2+ Vero E6 cells. A glycomimetic compound designed against DC-SIGN, enable inhibition of this process. These data have been then confirmed using authentic SARS-CoV-2 virus and human respiratory cell lines. Thus, we described a mechanism potentiating viral spreading of infection.
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- 2021
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4. Targeting of the C-Type Lectin Receptor Langerin Using Bifunctional Mannosylated Antigens
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Rui-Jun Eveline Li, Tim P. Hogervorst, Silvia Achilli, Sven C. M. Bruijns, Sander Spiekstra, Corinne Vivès, Michel Thépaut, Dmitri V. Filippov, Gijs A. van der Marel, Sandra J. van Vliet, Franck Fieschi, Jeroen D. C. Codée, and Yvette van Kooyk
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mannoside ,tumor associated antigens ,peptide conjugate ,vaccine model ,glyco-antigen ,Langerhans cell ,Biology (General) ,QH301-705.5 - Abstract
Langerhans cells (LCs) are antigen-presenting cells that reside in the skin. They uniquely express high levels of the C-type lectin receptor Langerin (CD207), which is an attractive target for antigen delivery in immunotherapeutic vaccination strategies against cancer. We here assess a library of 20 synthetic, well-defined mannoside clusters, built up from one, two, and three of six monomannosides, dimannosides, or trimannosides, appended to an oligopeptide backbone, for binding with Langerin using surface plasmon resonance and flow cytometric quantification. It is found that Langerin binding affinity increases with increasing number of mannosides. Hexavalent presentation of the mannosides resulted in binding affinities ranging from 3 to 12 μM. Trivalent presentation of the dimannosides and trimannosides led to Langerin affinity in the same range. The model melanoma gp100 antigenic peptide was subsequently equipped with a hexavalent cluster of the dimannosides and trimannosides as targeting moieties. Surprisingly, although the bifunctional conjugates were taken up in LCs in a Langerin-dependent manner, limited antigen presentation to cytotoxic T cells was observed. These results indicate that targeting glycan moieties on immunotherapeutic vaccines should not only be validated for target binding, but also on the continued effects on biology, such as antigen presentation to both CD8+ and CD4+ T cells.
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- 2020
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5. Bacteriophage T5 tail tube structure suggests a trigger mechanism for Siphoviridae DNA ejection
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Charles-Adrien Arnaud, Grégory Effantin, Corinne Vivès, Sylvain Engilberge, Maria Bacia, Pascale Boulanger, Eric Girard, Guy Schoehn, and Cécile Breyton
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Science - Abstract
Host cell recognition is mediated by the phage tail tip proteins, which then triggers viral genome delivery via the phage tail. Here, the authors combine crystallography and cryoEM to structurally characterise the bacteriophage T5 tail tube structure before and after interaction with its host receptor.
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- 2017
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6. Systematic Dual Targeting of Dendritic Cell C-Type Lectin Receptor DC-SIGN and TLR7 Using a Trifunctional Mannosylated Antigen
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Rui-Jun Eveline Li, Tim P. Hogervorst, Silvia Achilli, Sven C. Bruijns, Tim Arnoldus, Corinne Vivès, Chung C. Wong, Michel Thépaut, Nico J. Meeuwenoord, Hans van den Elst, Herman S. Overkleeft, Gijs A. van der Marel, Dmitri V. Filippov, Sandra J. van Vliet, Franck Fieschi, Jeroen D. C. Codée, and Yvette van Kooyk
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DC-SIGN ,TLR7 ,glyco-antigen ,vaccine model ,peptide conjugate ,tumor-associated antigens ,Chemistry ,QD1-999 - Abstract
Dendritic cells (DCs) are important initiators of adaptive immunity, and they possess a multitude of Pattern Recognition Receptors (PRR) to generate an adequate T cell mediated immunity against invading pathogens. PRR ligands are frequently conjugated to tumor-associated antigens in a vaccination strategy to enhance the immune response toward such antigens. One of these PPRs, DC-SIGN, a member of the C-type lectin receptor (CLR) family, has been extensively targeted with Lewis structures and mannose glycans, often presented in multivalent fashion. We synthesized a library of well-defined mannosides (mono-, di-, and tri-mannosides), based on known “high mannose” structures, that we presented in a systematically increasing number of copies (n = 1, 2, 3, or 6), allowing us to simultaneously study the effect of mannoside configuration and multivalency on DC-SIGN binding via Surface Plasmon Resonance (SPR) and flow cytometry. Hexavalent presentation of the clusters showed the highest binding affinity, with the hexa-α1,2-di-mannoside being the most potent ligand. The four highest binding hexavalent mannoside structures were conjugated to a model melanoma gp100-peptide antigen and further equipped with a Toll-like receptor 7 (TLR7)-agonist as adjuvant for DC maturation, creating a trifunctional vaccine conjugate. Interestingly, DC-SIGN affinity of the mannoside clusters did not directly correlate with antigen presentation enhancing properties and the α1,2-di-mannoside cluster with the highest binding affinity in our library even hampered T cell activation. Overall, this systematic study has demonstrated that multivalent glycan presentation can improve DC-SIGN binding but enhanced binding cannot be directly translated into enhanced antigen presentation and the sole assessment of binding affinity is thus insufficient to determine further functional biological activity. Furthermore, we show that well-defined antigen conjugates combining two different PRR ligands can be generated in a modular fashion to increase the effectiveness of vaccine constructs.
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- 2019
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7. C-terminal engineering of CXCL12 and CCL5 chemokines: functional characterization by electrophysiological recordings.
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Antoine Picciocchi, Lina Siaučiūnaiteė-Gaubard, Isabelle Petit-Hartlein, Rabia Sadir, Jean Revilloud, Lydia Caro, Michel Vivaudou, Franck Fieschi, Christophe Moreau, and Corinne Vivès
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Medicine ,Science - Abstract
Chemokines are chemotactic cytokines comprised of 70-100 amino acids. The chemokines CXCL12 and CCL5 are the endogenous ligands of the CXCR4 and CCR5 G protein-coupled receptors that are also HIV co-receptors. Biochemical, structural and functional studies of receptors are ligand-consuming and the cost of commercial chemokines hinders their use in such studies. Here, we describe methods for the expression, refolding, purification, and functional characterization of CXCL12 and CCL5 constructs incorporating C-terminal epitope tags. The model tags used were hexahistidines and Strep-Tag for affinity purification, and the double lanthanoid binding tag for fluorescence imaging and crystal structure resolution. The ability of modified and purified chemokines to bind and activate CXCR4 and CCR5 receptors was tested in Xenopus oocytes expressing the receptors, together with a Kir3 G-protein activated K(+) channel that served as a reporter of receptor activation. Results demonstrate that tags greatly influence the biochemical properties of the recombinant chemokines. Besides, despite the absence of any evidence for CXCL12 or CCL5 C-terminus involvement in receptor binding and activation, we demonstrated unpredictable effects of tag insertion on the ligand apparent affinity and efficacy or on the ligand dissociation. These tagged chemokines should constitute useful tools for the selective purification of properly-folded chemokines receptors and the study of their native quaternary structures.
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- 2014
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8. Mannobioside biomimetics that trigger DC-SIGN binding selectivity
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Irene Herrera-González, Michel Thépaut, Elena M. Sánchez-Fernández, Antonio di Maio, Corinne Vivès, Javier Rojo, José M. García Fernández, Franck Fieschi, Pedro M. Nieto, Carmen Ortiz Mellet, Department of Organic Chemistry, Faculty of Chemistry, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Instituto de Investigaciones Químicas (IIQ), Universidad de Sevilla / University of Sevilla-Consejo Superior de Investigaciones Científicas [Madrid] (CSIC), Instituto de Investigaciones Quimicas, C.S.I.C-Universidad de Sevilla, Glycosystems Laboratory, Instituto de Investigaciones Químicas, Centro de Investigaciones Científicas Isla de la Cartuja, and Facultad de Química, Universidad de Sevilla
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Binding Sites ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Metals and Alloys ,General Chemistry ,Catalysis ,Surfaces, Coatings and Films ,Electronic, Optical and Magnetic Materials ,Mannose-Binding Lectins ,Biomimetics ,Antigens, CD ,Materials Chemistry ,Ceramics and Composites ,Lectins, C-Type ,Protein Binding - Abstract
International audience; Selective DC-SIGN targeting vs. langerin might lead to anti-infective agents, given their counteracting effects upon infection by some pathogens. Here we show that multivalent sp2-iminosugar-containing mannobioside analogs can achieve total DC-SIGN selectivity by levering the canonic binding mode towards high-mannose oligosaccharide ligands, behaving as factual biomimics
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- 2022
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9. Membrane-Bound Flavocytochrome MsrQ Is a Substrate of the Flavin Reductase Fre in Escherichia coli
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Corinne Vivès, Vincent Nivière, Christelle Caux, Hawra Hassoune, Bruno Guigliarelli, Franck Fieschi, Frédéric Biaso, Isabelle Petit-Hartlein, Céline Juillan-Binard, Marius Horeau, and Stéphane Torelli
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0303 health sciences ,Hemeprotein ,biology ,Chemistry ,Stereochemistry ,030302 biochemistry & molecular biology ,Flavin mononucleotide ,Active site ,General Medicine ,Periplasmic space ,Biochemistry ,Cofactor ,03 medical and health sciences ,chemistry.chemical_compound ,Flavin reductase ,biology.protein ,Molecular Medicine ,Methionine sulfoxide reductase ,Heme ,030304 developmental biology - Abstract
MsrPQ is a new type of methionine sulfoxide reductase (Msr) system found in bacteria. It is specifically involved in the repair of periplasmic methionine residues that are oxidized by hypochlorous acid. MsrP is a periplasmic molybdoenzyme that carries out the Msr activity, whereas MsrQ, an integral membrane-bound hemoprotein, acts as the physiological partner of MsrP to provide electrons for catalysis. Although MsrQ (YedZ) was associated since long with a protein superfamily named FRD (ferric reductase domain), including the eukaryotic NADPH oxidases and STEAP proteins, its biochemical properties are still sparsely documented. Here, we have investigated the cofactor content of the E. coli MsrQ and its mechanism of reduction by the flavin reductase Fre. We showed by electron paramagnetic resonance (EPR) spectroscopy that MsrQ contains a single highly anisotropic low-spin (HALS) b-type heme located on the periplasmic side of the membrane. We further demonstrated that MsrQ holds a flavin mononucleotide (FMN) cofactor that occupies the site where a second heme binds in other members of the FDR superfamily on the cytosolic side of the membrane. EPR spectroscopy indicates that the FMN cofactor can accommodate a radical semiquinone species. The cytosolic flavin reductase Fre was previously shown to reduce the MsrQ heme. Here, we demonstrated that Fre uses the FMN MsrQ cofactor as a substrate to catalyze the electron transfer from cytosolic NADH to the heme. Formation of a specific complex between MsrQ and Fre could favor this unprecedented mechanism, which most likely involves transfer of the reduced FMN cofactor from the Fre active site to MsrQ.
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- 2021
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10. Front Cover: Precision Glycodendrimers for DC‐SIGN Targeting (Eur. J. Org. Chem. 29/2022)
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Giulio Goti, Cinzia Colombo, Silvia Achilli, Corinne Vivès, Michel Thépaut, Joanna Luczkowiak, Nuria Labiod, Rafael Delgado, Franck Fieschi, and Anna Bernardi
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Organic Chemistry ,Physical and Theoretical Chemistry - Published
- 2022
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11. Precision Glycodendrimers for DC‐SIGN Targeting**
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Giulio Goti, Cinzia Colombo, Silvia Achilli, Corinne Vivès, Michel Thépaut, Joanna Luczkowiak, Nuria Labiod, Rafael Delgado, Franck Fieschi, Anna Bernardi, Università degli Studi di Milano = University of Milan (UNIMI), Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Instituto de Investigación Hospital Universitario 12 de Octubre, and European Project: 642870,H2020,H2020-MSCA-ITN-2014,IMMUNOSHAPE(2015)
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Carbohydrates ,DC-SIGN ,Dendrimers ,Glycomimetics ,Langerin ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Organic Chemistry ,Settore CHIM/06 - Chimica Organica ,Physical and Theoretical Chemistry - Abstract
International audience; Multivalent ligands of the C‐type lectin receptor DC‐SIGN have emerged as effective antiadhesive agents against various pathogens. Some years ago, we described a hexavalent DC‐SIGN ligand, Polyman‐26, designed to bridge two of the four binding sites displayed by the receptor. In this work, we present our efforts to accomplish simultaneous coordination of all four carbohydrate binding sites of DC‐SIGN through the synthesis of cross‐shaped glycodendrimers. The tailored rigid scaffold allowed multivalent presentation of glycomimetics in a spatially defined fashion, while providing good water solubility to the constructs. Evaluation of the biological activity by SPR assays revealed strong binding avidity towards DC‐SIGN and increased selectivity over langerin. Inhibition of DC‐SIGN binding to SARS‐CoV‐2 spike protein and of DC‐SIGN mediated Ebola virus trans‐infection testifies for the glycodendrimers potential application in infection diseases. The tetravalent platform described here is easily accessible and can be used in modular fashion with different ligands, thus lending itself to multiple applications.Multivalent antagonists able to reach the four carbohydrate recognition domains (CRD) of DC‐SIGN have been prepared. The extended rigid core of these glycodendrimers allows multivalent presentation of glycomimetic molecules in a spatially defined fashion, providing high affinity towards DC‐SIGN and selectivity over other C‐type lectins featuring distinct CRD arrangements. The constructs successfully inhibit DC‐SIGN binding to SARS‐CoV‐2 spike protein and DC‐SIGN mediated trans‐infection by Ebola virus.
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- 2022
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12. Lectin recognition and hepatocyte endocytosis of GalNAc-decorated nanostructured lipid carriers
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Mireille Chevallet, Franck Fieschi, Francois Bulteau Bulteau, Christelle Gateau, Pascale Delangle, Aurélien Deniaud, Michel Thépaut, Isabelle Texier, Laura Gauthier, Corinne Vivès, Chimie Interface Biologie pour l’Environnement, la Santé et la Toxicologie (CIBEST ), SYstèmes Moléculaires et nanoMatériaux pour l’Energie et la Santé (SYMMES), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Département Microtechnologies pour la Biologie et la Santé (DTBS), Commissariat à l'énergie atomique et aux énergies alternatives - Laboratoire d'Electronique et de Technologie de l'Information (CEA-LETI), Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Métaux et Organes (MET&OR), Laboratoire de Chimie et Biologie des Métaux (LCBM - UMR 5249), Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017)
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Acetylgalactosamine ,surface functionalisation ,N-acetylgalactosamine ,nanostructured lipid carriers ,Pharmaceutical Science ,[SDV.BC]Life Sciences [q-bio]/Cellular Biology ,02 engineering and technology ,Endocytosis ,macrophage galactose-lectin ,targeted drug delivery ,N-Acetylgalactosamine ,03 medical and health sciences ,chemistry.chemical_compound ,asialoglycoprotein receptor ,0302 clinical medicine ,Lectins ,medicine ,Humans ,Drug Carriers ,biology ,Lectin ,[SDV.MHEP.HEG]Life Sciences [q-bio]/Human health and pathology/Hépatology and Gastroenterology ,Hep G2 Cells ,[SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences ,021001 nanoscience & nanotechnology ,Lipids ,Nanostructures ,Cell biology ,carbohydrates (lipids) ,medicine.anatomical_structure ,chemistry ,Targeted drug delivery ,030220 oncology & carcinogenesis ,Hepatocyte ,Drug delivery ,biology.protein ,hepatocytes ,lipids (amino acids, peptides, and proteins) ,Asialoglycoprotein receptor ,Nanocarriers ,0210 nano-technology - Abstract
International audience; Liver is the main organ for metabolism but is also subject to various pathologies, from viral, genetic, cancer or metabolic origin. There is thus a crucial need to develop efficient liver-targeted drug delivery strategies. Asialoglycoprotein receptor (ASGPR) is a C-type lectin expressed in the hepatocyte plasma membrane that efficiently endocytoses glycoproteins exposing galactose (Gal) or N-acetylgalactosamine (GalNAc). Its targeting has been successfully used to drive the uptake of small molecules decorated with three or four GalNAc, thanks to an optimisation of their spatial arrangement. Herein, we assessed the biological properties of highly stable nanostructured lipid carriers (NLC) made of FDA-approved ingredients and formulated with increasing amounts of GalNAc. Cellular studies showed that a high density of GalNAc was required to favour hepatocyte internalisation via the ASGPR pathway. Interaction studies using surface plasmon resonance and the macrophage galactose-lectin as GalNAc-recognising lectin confirmed the need of high GalNAc density for specific recognition of these NLC. This work is the first step for the development of efficient nanocarriers for prolonged liver delivery of active compounds.
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- 2020
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13. Targeting Tn-Antigen-Positive Human Tumors with a Recombinant Human Macrophage Galactose C-Type Lectin
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François Bulteau, Michel Thépaut, Maxime Henry, Amandine Hurbin, Laetitia Vanwonterghem, Corinne Vivès, Aline Le Roy, Christine Ebel, Olivier Renaudet, Franck Fieschi, Jean-Luc Coll, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Département de Chimie Moléculaire (DCM), and Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)
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MESH: HT29 Cells ,Pharmaceutical Science ,MESH: Spheroids, Cellular ,Mice, Nude ,MESH: Flow Cytometry ,MESH: Recombinant Proteins ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Spheroids, Cellular ,Drug Discovery ,Tn antigen ,MESH: Mice, Nude ,cancer ,MESH: Microscopy, Confocal ,Animals ,Humans ,MESH: Animals ,Antigens, Tumor-Associated, Carbohydrate ,Lectins, C-Type ,MESH: Mice ,030304 developmental biology ,0303 health sciences ,MESH: Humans ,Microscopy, Confocal ,MESH: Antigens, Tumor-Associated, Carbohydrate ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Surface Plasmon Resonance ,Flow Cytometry ,Recombinant Proteins ,3. Good health ,MESH: Surface Plasmon Resonance ,C-type lectin ,A549 Cells ,030220 oncology & carcinogenesis ,Molecular Medicine ,Female ,MESH: A549 Cells ,MESH: Female ,HT29 Cells ,MESH: Neoplasm Transplantation ,MESH: Lectins, C-Type ,Neoplasm Transplantation - Abstract
International audience; Alterations in glycosylation cause the emergence of tumor-associated carbohydrate antigens (TACAs) during tumorigenesis. Truncation of O-glycans reveals the Thomsen nouveau (Tn) antigen, an N-acetylgalactosamine (GalNAc) frequently attached to serine or threonine amino acids, that is accessible on the surface of cancer cells but not on healthy cells. Interestingly, GalNac can be recognized by macrophage galactose lectin (MGL), a type C lectin receptor expressed in immune cells. In this study, recombinant MGL fragments were tested in vitro for their cancer cell-targeting efficiency by flow cytometry and confocal microscopy and in vivo after administration of fluorescent MGL to tumor-bearing mice. Our results demonstrate the ability of MGL to target Tn-positive human tumors without inducing toxicity. This outcome makes MGL, a fragment of a normal human protein, the first vector candidate for in vivo diagnosis and imaging of human tumors and, possibly, for therapeutic applications.
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- 2021
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14. Membrane-Bound Flavocytochrome MsrQ Is a Substrate of the Flavin Reductase Fre in
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Christelle, Caux, Bruno, Guigliarelli, Corinne, Vivès, Frédéric, Biaso, Marius, Horeau, Hawra, Hassoune, Isabelle, Petit-Hartlein, Céline, Juillan-Binard, Stephane, Torelli, Franck, Fieschi, and Vincent, Nivière
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Kinetics ,Flavin Mononucleotide ,Escherichia coli Proteins ,Electron Spin Resonance Spectroscopy ,Escherichia coli ,Membrane Proteins ,Enzymes ,Substrate Specificity - Abstract
MsrPQ is a new type of methionine sulfoxide reductase (Msr) system found in bacteria. It is specifically involved in the repair of periplasmic methionine residues that are oxidized by hypochlorous acid. MsrP is a periplasmic molybdoenzyme that carries out the Msr activity, whereas MsrQ, an integral membrane-bound hemoprotein, acts as the physiological partner of MsrP to provide electrons for catalysis. Although MsrQ (YedZ) was associated since long with a protein superfamily named FRD (ferric reductase domain), including the eukaryotic NADPH oxidases and STEAP proteins, its biochemical properties are still sparsely documented. Here, we have investigated the cofactor content of the
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- 2021
15. Controlled density glycodendron microarrays for studying carbohydrate–lectin interactions
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Michel Thépaut, Corinne Vivès, Franck Fieschi, Antonio Di Maio, Niels-C. Reichardt, Anna Cioce, Silvia Achilli, Javier Rojo, Instituto de Investigaciones Químicas (IIQ), Universidad de Sevilla / University of Sevilla-Consejo Superior de Investigaciones Científicas [Madrid] (CSIC), Carbon Bionanotechnology Laboratory (CICbiomaGUNE), Centro de Investigación Cooperativa en Biomateriales (CIC biomaGUNE), Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Centro Investigacion Biomedica en Red Bioingenieria, Biomateriales y Nanomedicina - CIBER-BBN (SPAIN), ANR-10-INBS-0005,FRISBI,Infrastructure Française pour la Biologie Structurale Intégrée(2010), ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017), ANR-15-IDEX-0002,UGA,IDEX UGA(2015), European Project: 642870,H2020,H2020-MSCA-ITN-2014,IMMUNOSHAPE(2015), European Research Council, Ministerio de Economía y Competitividad (España), and Consejo Superior de Investigaciones Científicas [Madrid] (CSIC)-Universidad de Sevilla
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Glycan ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,010405 organic chemistry ,Chemistry ,Organic Chemistry ,Valency ,Lectin ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Combinatorial chemistry ,Cycloaddition ,3. Good health ,0104 chemical sciences ,Indium tin oxide ,Hydrophobic effect ,chemistry.chemical_compound ,Lectins ,Dendrimer ,biology.protein ,Azide ,Physical and Theoretical Chemistry - Abstract
Glycodendron microarrays with defined valency have been constructed by on-chip synthesis on hydrophobic indium tin oxide (ITO) coated glass slides and employed in lectin–carbohydrate binding studies with several plant and human lectins. Glycodendrons presenting sugar epitopes at different valencies were prepared by spotwise strain-promoted azide–alkyne cycloaddition (SPAAC) between immobilised cyclooctyne dendrons and azide functionalised glycans. The non-covalent immobilisation of dendrons on the ITO surface by hydrophobic interaction allowed us to study dendron surface density and SPAAC conversion rate by in situ MALDI-TOF MS analysis. By diluting the dendron surface density we could study how the carbohydrate–lectin interactions became exclusively dependant on the valency of the immobilised glycodendron.
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- 2021
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16. DC/L-SIGN recognition of spike glycoprotein promotes SARS-CoV-2 trans-infection and can be inhibited by a glycomimetic antagonist
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Daphna Fenel, Fátima Lasala, Anna Bernardi, Guy Schoehn, Michel Thépaut, Joanna Luczkowiak, Rafael L. Delgado, Isabelle Bally, Sara Sattin, Franck Fieschi, Nicole M. Thielens, Corinne Vivès, Yasmina Grimoire, Nuria Labiod, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Instituto de Investigación Hospital Universitario 12 de Octubre, Università degli Studi di Milano = University of Milan (UNIMI), and Università degli Studi di Milano [Milano] (UNIMI)
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RNA viruses ,Viral Diseases ,Pulmonology ,Coronaviruses ,viruses ,Cell ,Jurkat cells ,Biochemistry ,Pathogenesis ,Jurkat Cells ,0302 clinical medicine ,Medical Conditions ,Glycomimetic ,Animal Cells ,Lectins ,Chlorocebus aethiops ,Biology (General) ,Receptor ,Immune Response ,Lung ,Pathology and laboratory medicine ,Mannan-binding lectin ,chemistry.chemical_classification ,0303 health sciences ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Medical microbiology ,Cell biology ,3. Good health ,medicine.anatomical_structure ,Infectious Diseases ,030220 oncology & carcinogenesis ,Mannosides ,Viruses ,Spike Glycoprotein, Coronavirus ,SARS CoV 2 ,Pathogens ,Cellular Types ,Research Article ,Protein Binding ,Langerin ,SARS coronavirus ,QH301-705.5 ,Immune Cells ,Immunology ,Antigen-Presenting Cells ,Receptors, Cell Surface ,Respiratory Mucosa ,Microbiology ,Virus ,Cell Line ,03 medical and health sciences ,Respiratory Disorders ,Antigens, CD ,Virology ,medicine ,Genetics ,Animals ,Humans ,Lectins, C-Type ,Protein Interactions ,Molecular Biology ,Vero Cells ,030304 developmental biology ,Medicine and health sciences ,Innate immune system ,Biology and life sciences ,SARS-CoV-2 ,Organisms ,Viral pathogens ,Proteins ,COVID-19 ,Covid 19 ,Cell Biology ,Dendritic Cells ,RC581-607 ,Microbial pathogens ,Mannose-Binding Lectins ,chemistry ,Cell culture ,Respiratory Infections ,Vero cell ,biology.protein ,Parasitology ,Immunologic diseases. Allergy ,Glycoprotein ,Cell Adhesion Molecules ,030217 neurology & neurosurgery - Abstract
The efficient spread of SARS-CoV-2 resulted in a unique pandemic in modern history. Despite early identification of ACE2 as the receptor for viral spike protein, much remains to be understood about the molecular events behind viral dissemination. We evaluated the contribution of C-type lectin receptors (CLRS) of antigen-presenting cells, widely present in respiratory mucosa and lung tissue. DC-SIGN, L-SIGN, Langerin and MGL bind to diverse glycans of the spike using multiple interaction areas. Using pseudovirus and cells derived from monocytes or T-lymphocytes, we demonstrate that while virus capture by the CLRs examined does not allow direct cell infection, DC/L-SIGN, among these receptors, promote virus transfer to permissive ACE2+ Vero E6 cells. A glycomimetic compound designed against DC-SIGN, enable inhibition of this process. These data have been then confirmed using authentic SARS-CoV-2 virus and human respiratory cell lines. Thus, we described a mechanism potentiating viral spreading of infection., Author summary For their infectious effectiveness, viruses often use host attachment factors to improve their adhesion to the cell surface. This will mainly increase viruses concentration at cell surfaces, potentiating access and final engagement with their real entry receptors. This mechanism enhances viral infection of target cells or even allow viruses to be captured by non-permissive cells for secondary presentation to permissive cells by a process called trans-infection. While, the entry mechanism of SARS-CoV-2 using the ACE2 entry receptor, is well defined, little is known about additional factors explaining the high transmission rate of this virus. The level of glycosylations on the SARS-CoV-2 Spike protein prompted us to assess whether CLRs of immune cells, regularly diverted by pathogens, could play a role in the spread of SARS-CoV-2. Here, we show that these receptors are able to recognize Spike envelope protein of SARS-CoV-2 and two receptors among the four tested, DC-SIGN and L-SIGN, are able to promote virus trans-infection. This work identifies a new family of alternative SARS-CoV-2 cell receptors involved in uncharacterized dissemination mechanism. Moreover, considering the role of CLRs in immunomodulation, their early involvement opens avenues for understanding the imbalanced innate immune response observed in COVID-19 pathogenesis.
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- 2021
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17. Chemo‐Enzymatic Synthesis of S. mansoni O‐Glycans and Their Evaluation as Ligands for C‐Type Lectin Receptors MGL, DC‐SIGN, and DC‐SIGNR
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Anna Cioce, Niels-Christian Reichardt, Corinne Vivès, Anna Bernardi, Alvaro Hernández, Julie Pham, Franck Fieschi, Silvia Achilli, Giulio Goti, Michel Thépaut, CIC BiomaGUNE, CIC BiomaGUNE [Espagne], Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Università degli Studi di Milano = University of Milan (UNIMI), and Dipartimento di chimica, Universita degli Studi di Milano
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Glycan ,Fucosyltransferase ,Glycoconjugate ,enzymes ,carbohydrates ,Receptors, Cell Surface ,Ligands ,010402 general chemistry ,01 natural sciences ,Catalysis ,Polysaccharides ,C-type lectin ,Humans ,Lectins, C-Type ,Receptor ,microarrays ,chemistry.chemical_classification ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,010405 organic chemistry ,Chemistry ,Organic Chemistry ,glycoconjugates ,lectins ,Lectin ,General Chemistry ,biology.organism_classification ,0104 chemical sciences ,3. Good health ,DC-SIGN ,carbohydrates (lipids) ,Biochemistry ,biology.protein ,Schistosoma mansoni ,Cell Adhesion Molecules - Abstract
International audience; Due to their interactions with C-type lectin receptors (CLRs), glycans from the helminth Schistosoma mansoni represent promising leads for treatment of autoimmune diseases, allergies or cancer. We chemo-enzymatically synthesized nine O-glycans based on the two predominant O-glycan cores observed in the infectious stages of schistosomiasis, the mucin core 2 and the S. mansoni core. The O-glycans were fucosylated next to a selection of N-glycans directly on a microarray slide using a recombinant fucosyltransferase and GDP-fucose or GDP-6-azidofucose as donor. Binding assays with fluorescently labelled human CLRs DC-SIGN, DC-SIGNR and MGL revealed the novel O-glycan O8 as the best ligand for MGL from our panel. Significant binding to DC-SIGN was also found for azido-fucosylated glycans. Contrasting binding specificities were observed between the monovalent carbohydrate recognition domain (CRD) and the tetravalent extracellular domain (ECD) of DC-SIGNR.
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- 2020
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18. Structure-Based Design of Glycodendrimer Antagonists for Improved DC-SIGN Targeting
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Michel Thépaut, Franck Fieschi, Anna Bernardi, Giulio Goti, Cinzia Colombo, Corinne Vivès, and Silvia Achilli
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DC-SIGN ,Langerin ,biology ,Chemistry ,biology.protein ,Biophysics ,Structure based ,Avidity ,Biological activity ,Binding site ,Surface plasmon resonance ,Strong binding - Abstract
DC-SIGN multivalent antagonists have emerged as effective antiadhesive agents against various pathogen infections. Recently, our group have shown that high potency can be achieved upon bridging two of the four binding sites displayed by the protein. Here we present our endeavors to accomplish the tetracoordination of DC-SIGN through the synthesis of two cross-shaped glycodendrimers. The choice of a tailored rigid scaffold allowed multivalent presentation of glycomimetics in a spatially defined fashion, while providing good water solubility to the constructs. Evaluation of the biological activity by SPR assay revealed strong binding avidity towards DC-SIGN and increased selectivity over langerin.
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- 2020
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19. TETRALEC, Artificial Tetrameric Lectins: A Tool to Screen Ligand and Pathogen Interactions
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Sabine Mayer-Lambertz, Corinne Vivès, Bernd Lepenies, Franck Fieschi, João T. Monteiro, Christine Ebel, Michel Thépaut, Niels-Christian Reichardt, Aline Le Roy, Sonia Serna, Silvia Achilli, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Immunology Unit and Research Center for Emerging Infections and Zoonoses, University of Veterinary Medicine Hannover, CIC biomaGUNE, Glycotechnology Laboratory, Biomedical Research Networking Center in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Instituto de Salud Carlos III [Madrid] (ISC)-ministerio de ciencia e innovacion, Department of Biomolecular Systems [Potsdam], Max Planck Institute of Colloids and Interfaces, Max-Planck-Gesellschaft-Max-Planck-Gesellschaft, and ANR-17-EURE-0003,CBH-EUR-GS,CBH-EUR-GS(2017)
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0301 basic medicine ,Glycan ,Recombinant Fusion Proteins ,glycan array ,Ligands ,01 natural sciences ,Catalysis ,Article ,Flow cytometry ,law.invention ,lcsh:Chemistry ,Inorganic Chemistry ,03 medical and health sciences ,C-type lectin ,Confocal microscopy ,law ,Candida albicans ,medicine ,pathogen recognition ,Lectins, C-Type ,Physical and Theoretical Chemistry ,Receptor ,lcsh:QH301-705.5 ,Molecular Biology ,Spectroscopy ,biology ,medicine.diagnostic_test ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,010405 organic chemistry ,Chemistry ,Organic Chemistry ,Lectin ,General Medicine ,multivalency ,Ligand (biochemistry) ,Flow Cytometry ,In vitro ,eye diseases ,0104 chemical sciences ,Computer Science Applications ,Immunoglobulin Fc Fragments ,DC-SIGNR ,030104 developmental biology ,lcsh:Biology (General) ,lcsh:QD1-999 ,Biochemistry ,biology.protein - Abstract
International audience; C-type lectin receptor (CLR)/carbohydrate recognition occurs through low affinity interactions. Nature compensates that weakness by multivalent display of the lectin carbohydrate recognition domain (CRD) at the cell surface. Mimicking these low affinity interactions in vitro is essential to better understand CLR/glycan interactions. Here, we present a strategy to create a generic construct with a tetrameric presentation of the CRD for any CLR, termed TETRALEC. We applied our strategy to a naturally occurring tetrameric CRD, DC-SIGNR, and compared the TETRALEC ligand binding capacity by synthetic N- and O-glycans microarray using three different DC-SIGNR constructs i) its natural tetrameric counterpart, ii) the monomeric CRD and iii) a dimeric Fc-CRD fusion. DC-SIGNR TETRALEC construct showed a similar binding profile to that of its natural tetrameric counterpart. However, differences observed in recognition of low affinity ligands underlined the importance of the CRD spatial arrangement. Moreover, we further extended the applications of DC-SIGNR TETRALEC to evaluate CLR/pathogens interactions. This construct was able to recognize heat-killed Candida albicans by flow cytometry and confocal microscopy, a so far unreported specificity of DC-SIGNR. In summary, the newly developed DC-SIGNR TETRALEC tool proved to be useful to unravel novel CLR/glycan interactions, an approach which could be applied to other CLRs.
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- 2020
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20. Development of C-type lectin-oriented surfaces for high avidity glycoconjugates: towards mimicking multivalent interactions on the cell surface
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Corinne Vivès, Vanessa Porkolab, Anna Bernardi, Carlo Pifferi, Ieva Sutkeviciute, Marwa Taouai, Olivier Renaudet, Michel Thépaut, Franck Fieschi, Mohammed Benazza, Stefania Ordanini, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Département de Chimie Moléculaire (DCM), Université Grenoble Alpes (UGA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Università degli Studi di Milano [Milano] (UNIMI), Laboratoire de Glycochimie, des Antimicrobiens et des Agro-ressources - UMR CNRS 7378 (LG2A ), Université de Picardie Jules Verne (UPJV)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), and Università degli Studi di Milano = University of Milan (UNIMI)
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Surface Properties ,Glycoconjugate ,Molecular Conformation ,Receptors, Cell Surface ,010402 general chemistry ,01 natural sciences ,Biochemistry ,03 medical and health sciences ,C-type lectin ,Glycomimetic ,Humans ,Lectins, C-Type ,Avidity ,Physical and Theoretical Chemistry ,Surface plasmon resonance ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Chemistry ,Organic Chemistry ,Lectin ,Surface Plasmon Resonance ,Ligand (biochemistry) ,0104 chemical sciences ,Covalent bond ,Biophysics ,biology.protein ,Cell Adhesion Molecules ,Glycoconjugates - Abstract
International audience; Multivalent interactions between complex carbohydrates and oligomeric C-type lectins govern a wide range of immune responses. Up to date, standard SPR (surface plasmon resonance) competitive assays have largely been to evaluate binding properties from monosaccharide units (low affinity, mM) to multivalent elemental antagonists (moderate affinity, μM). Herein, we report typical case-studies of SPR competitive assays showing that they underestimate the potency of glycoclusters to inhibit the interaction between DC-SIGN and immobilized glycoconjugates. This paper describes the design and implementation of a SPR direct interaction over DC-SIGN oriented surfaces, extendable to other C-type lectin surfaces as such Langerin. This setup provides an overview of intrinsic avidity generation emanating simultaneously from multivalent glycoclusters and from DC-SIGN tetramers organized in nanoclusters at the cell membrane. To do so, covalent biospecific capture of DC-SIGN via StreptagII/StrepTactin interaction preserves tetrameric DC-SIGN, accessibility and topology of its active sites, that would have been dissociated using standard EDC-NHS procedure under acidic conditions. From the tested glycoclusters libraries, we demonstrated that the scaffold architecture, the valency and the glycomimetic-based ligand are crucial to reach nanomolar affinities for DC-SIGN. The glycocluster 3·D illustrates the tightest binding partner in this set for a DC-SIGN surface (KD = 18 nM). Moreover, the selectivity at monovalent scale of glycomimetic D can be easily analyzed at multivalent scale comparing its binding over different C-type lectin immobilized surfaces. This approach may give rise to novel insights into the multivalent binding mechanisms responsible for avidity and make a major contribution to the full characterization of the binding potency of promising specific and multivalent immodulators.
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- 2020
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21. Interdomain Flexibility within NADPH Oxidase Suggested by SANS Using LMNG Stealth Carrier
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Annelise Vermot, Anne L. Martel, Corinne Vivès, Sergei Grudinin, Michael Härtlein, Aline Le Roy, Michel Thépaut, Isabelle Petit-Hartlein, Christine Ebel, Martine Moulin, Susan M.E. Smith, Cécile Breyton, Franck Fieschi, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Sciences et Société, Historicité, Éducation et Pratiques (EA S2HEP), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, Institut Laue-Langevin (ILL), Algorithms for Modeling and Simulating Nanosystems [2018-...] (NANO-D-POST [2018-2020]), Inria Grenoble - Rhône-Alpes, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria), Department of Molecular and Cellular Biology [Kennesaw], Kennesaw State University (KSU), ANR-17-CE11-0013,BANDIT,d'une Nox bactérienne à la conception d'outil de criblage(2017), ANR-16-CE92-0001,FLUOR,Tensioactifs Fluorés pour l'Étude de Protéines Membranaires(2016), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), ILL, Algorithms for Modeling and Simulation of Nanosystems (NANO-D), Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Laboratoire Jean Kuntzmann (LJK), Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP ), Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP ), and Université Grenoble Alpes (UGA)
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0303 health sciences ,Molecular model ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Biophysics ,Membrane Proteins ,NADPH Oxidases ,Dehydrogenase ,Articles ,Neopentyl glycol ,Small-angle neutron scattering ,Transmembrane protein ,03 medical and health sciences ,chemistry.chemical_compound ,Crystallography ,Neutron Diffraction ,0302 clinical medicine ,chemistry ,Docking (molecular) ,Scattering, Small Angle ,Linker ,Integral membrane protein ,Oxidation-Reduction ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
International audience; Small angle neutron scattering (SANS) provides a method to obtain important low-resolution information for integral membrane proteins (IMPs), challenging targets for structural determination. Specific deuteration furnishes a "stealth" carrier for the solubilized IMP. We used SANS to determine a structural envelope of SpNOX, the Streptococcus pneumoniae NADPH oxidase (NOX), a prokaryotic model system for exploring structure and function of eukaryotic NOXes. SpNOX was solubilized in the detergent lauryl maltose neopentyl glycol, which provides optimal SpNOX stability and activity. Using deuterated solvent and protein, the lauryl maltose neopentyl glycol was experimentally undetected in SANS. This affords a cost-effective SANS approach for obtaining novel structural information on IMPs. Combining SANS data with molecular modeling provided a first, to our knowledge, structural characterization of an entire NOX enzyme. It revealed a distinctly less compact structure than that predicted from the docking of homologous crystal structures of the separate transmembrane and dehydrogenase domains, consistent with a flexible linker connecting the two domains.
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- 2020
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22. Second-Generation Dendrimers with Chondroitin Sulfate Type-E Disaccharides as Multivalent Ligands for Langerin
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Pedro M. Nieto, Javier Rojo, José L. de Paz, Corinne Vivès, Pedro Domínguez-Rodríguez, Michel Thépaut, Franck Fieschi, Glycosystems Laboratory, Instituto de Investigaciones Químicas, Centro de Investigaciones Científicas Isla de la Cartuja, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)
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Dendrimers ,Polymers and Plastics ,Langerin ,Disaccharide ,Bioengineering ,02 engineering and technology ,Disaccharides ,Ligands ,010402 general chemistry ,01 natural sciences ,Biomaterials ,chemistry.chemical_compound ,Sulfation ,Polysaccharides ,C-type lectin ,Dendrimer ,Materials Chemistry ,Chondroitin sulfate ,Surface plasmon resonance ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,biology ,Chondroitin Sulfates ,Lectin ,021001 nanoscience & nanotechnology ,Combinatorial chemistry ,0104 chemical sciences ,chemistry ,biology.protein ,0210 nano-technology - Abstract
International audience; Chondroitin sulfate type-E (CS-E) is a sulfated polysaccharide that shows several interesting biological activities, such as modulation of the neuronal growth factor signaling and its interaction with langerin, a C-type lectin with a crucial role in the immunological system. However, applications of CS-E are hampered by the typical heterogeneous structure of the natural polysaccharide. Well-defined, homogeneous CS-E analogues are highly demanded. Here, we report the synthesis of monodispersed, structurally well-defined second-generation glycodendrimers displaying up to 18 CS-E disaccharide units. These complex multivalent systems have a molecular weight and a number of disaccharide repeating units comparable with those of the natural polysaccharides. In addition, surface plasmon resonance experiments revealed a calcium-independent interaction between these glycodendrimers and langerin, in the micromolar range, highlighting the utility of these compounds as CS-E mimetics.
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- 2020
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23. Down-regulation of NOX2 activity in phagocytes mediated by ATM-kinase dependent phosphorylation
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Franck Fieschi, Sylvain Beaumel, Corinne Vivès, Anne-Marie Hesse, Franck Debeurme, Antoine Picciocchi, Marie José Stasia, Didier Grunwald, Myriam Ferro, Susan M.E. Smith, Heather Stieglitz, Pahk Thepchatri, Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Etude de la dynamique des protéomes (EDyP), Laboratoire de Biologie à Grande Échelle (BGE - UMR S1038), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Biologie du Cancer et de l'Infection (BCI ), Department of Molecular and Cellular Biology, Kennesaw State University (KSU), Thérapeutique Recombinante Expérimentale (TIMC-IMAG-TheREx), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-VetAgro Sup (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-VetAgro Sup (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Centre Hospitalier Universitaire [Grenoble] (CHU), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Etude de la dynamique des protéomes (EDyP ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Department of Molecular and Cellular Biology [Kennesaw], Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)
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0301 basic medicine ,Cytochrome ,[SDV]Life Sciences [q-bio] ,Mutant ,Down-Regulation ,Ataxia Telangiectasia Mutated Proteins ,Biochemistry ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Downregulation and upregulation ,Cell Line, Tumor ,Physiology (medical) ,Humans ,Phosphorylation ,Ataxia telangiectasia mutated (ATM) ,ComputingMilieux_MISCELLANEOUS ,Phagocytes ,NADPH oxidase ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,030102 biochemistry & molecular biology ,biology ,urogenital system ,Kinase ,Chemistry ,Superoxide ,Neutrophil ,NOX ,Cell biology ,030104 developmental biology ,Gene Expression Regulation ,NADPH Oxidase 2 ,NOX-specific Insertion Sequence (NIS) ,cardiovascular system ,biology.protein ,Phorbol ,Protein Processing, Post-Translational ,hormones, hormone substitutes, and hormone antagonists ,Signal Transduction ,circulatory and respiratory physiology - Abstract
International audience; NADPH oxidases (NOX) have many biological roles, but their regulation to control production of potentially toxic ROS molecules remains unclear. A previously identified insertion sequence of 21 residues (called NIS) influences NOX activity, and its predicted flexibility makes it a good candidate for providing a dynamic switch controlling the NOX active site. We constructed NOX2 chimeras in which NIS had been deleted or exchanged with those from other NOXs (NIS1, 3 and 4). All contained functional heme and were expressed normally at the plasma membrane of differentiated PLB-985 cells. However, NOX2-ΔNIS and NOX2-NIS1 had neither NADPH-oxidase nor reductase activity and exhibited abnormal translocation of p47(phox) and p67(phox) to the phagosomal membrane. This suggested a functional role of NIS. Interestingly after activation, NOX2-NIS3 cells exhibited superoxide overproduction compared with wild-type cells. Paradoxically, the Vmax of purified unstimulated NOX2-NIS3 was only one-third of that of WT-NOX2. We therefore hypothesized that post-translational events regulate NOX2 activity and differ between NOX2-NIS3 and WT-NOX2. We demonstrated that Ser486, a phosphorylation target of ataxia telangiectasia mutated kinase (ATM kinase) located in the NIS of NOX2 (NOX2-NIS), was phosphorylated in purified cytochrome b558 after stimulation with phorbol 12-myristate-13-acetate (PMA). Moreover, ATM kinase inhibition and a NOX2 Ser486Ala mutation enhanced NOX activity whereas a Ser486Glu mutation inhibited it. Thus, the absence of Ser486 in NIS3 could explain the superoxide overproduction in the NOX2-NIS3 mutant. These results suggest that PMA-stimulated NOX2-NIS phosphorylation by ATM kinase causes a dynamic switch that deactivates NOX2 activity. We hypothesize that this downregulation is defective in NOX2-NIS3 mutant because of the absence of Ser486.
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- 2017
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24. Systematic Dual Targeting of Dendritic Cell C-Type Lectin Receptor DC-SIGN and TLR7 Using a Trifunctional Mannosylated Antigen
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Michel Thépaut, Franck Fieschi, Corinne Vivès, Sandra J. van Vliet, Hans van den Elst, Tim Arnoldus, Sven C. M. Bruijns, Rui Jun Eveline Li, Nico J. Meeuwenoord, Gijs A. van der Marel, Herman S. Overkleeft, Tim P Hogervorst, Dmitri V. Filippov, Jeroen D. C. Codée, Silvia Achilli, Yvette van Kooyk, Chung C. Wong, Molecular cell biology and Immunology, AGEM - Digestive immunity, AGEM - Re-generation and cancer of the digestive system, CCA - Cancer biology and immunology, AII - Cancer immunology, Department of Molecular Cell Biology and Immunology (Academic Medical Center, Amsterdam), VU University Medical Center [Amsterdam], Department of Bio-organic Synthesis, Faculty of Science, Leiden Institute of Chemistry, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
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Antigen presentation ,02 engineering and technology ,010402 general chemistry ,01 natural sciences ,DC-SIGN ,lcsh:Chemistry ,Antigen ,C-type lectin ,mannoside ,vaccine model ,tumor-associated antigens ,Original Research ,TLR7 ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Chemistry ,Pattern recognition receptor ,General Chemistry ,Dendritic cell ,021001 nanoscience & nanotechnology ,Ligand (biochemistry) ,Acquired immune system ,0104 chemical sciences ,3. Good health ,Cell biology ,lcsh:QD1-999 ,glyco-antigen ,biology.protein ,0210 nano-technology ,peptide conjugate - Abstract
International audience; Dendritic cells (DCs) are important initiators of adaptive immunity, and they possess a multitude of Pattern Recognition Receptors (PRR) to generate an adequate T cell mediated immunity against invading pathogens. PRR ligands are frequently conjugated to tumor-associated antigens in a vaccination strategy to enhance the immune response toward such antigens. One of these PPRs, DC-SIGN, a member of the C-type lectin receptor (CLR) family, has been extensively targeted with Lewis structures and mannose glycans, often presented in multivalent fashion. We synthesized a library of well-defined mannosides (mono-, di-, and tri-mannosides), based on known "high mannose" structures, that we presented in a systematically increasing number of copies (n = 1, 2, 3, or 6), allowing us to simultaneously study the effect of mannoside configuration and multivalency on DC-SIGN binding via Surface Plasmon Resonance (SPR) and flow cytometry. Hexavalent presentation of the clusters showed the highest binding affinity, with the hexa-α1,2-di-mannoside being the most potent ligand. The four highest binding hexavalent mannoside structures were conjugated to a model melanoma gp100-peptide antigen and further equipped with a Toll-like receptor 7 (TLR7)-agonist as adjuvant for DC maturation, creating a trifunctional vaccine conjugate. Interestingly, DC-SIGN affinity of the mannoside clusters did not directly correlate with antigen presentation enhancing properties and the α1,2-di-mannoside cluster with the highest binding affinity in our library even hampered T cell activation. Overall, this systematic study has demonstrated that multivalent glycan presentation can improve DC-SIGN binding but enhanced binding cannot be directly translated into enhanced antigen presentation and the sole assessment of binding affinity is thus insufficient to determine further functional biological activity. Furthermore, we show that well-defined antigen conjugates combining two different PRR ligands can be generated in a modular fashion to increase the effectiveness of vaccine constructs.
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- 2019
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25. Development of c-type lectin oriented surfaces for high avidity glycoconjugates: towards mimicking multivalent interactions on the cell surface
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Corinne Vivès, Vanessa Porkolab, Ieva Sutkeviciute, Marwa Taouai, Franck Fieschi, Olivier Renaudet, Mohammed Benazza, Anna Bernardi, Stefania Ordanini, Carlo Pifferi, and Michel Thépaut
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chemistry.chemical_classification ,0303 health sciences ,biology ,Glycoconjugate ,Lectin ,010402 general chemistry ,Ligand (biochemistry) ,01 natural sciences ,0104 chemical sciences ,03 medical and health sciences ,chemistry ,Glycomimetic ,C-type lectin ,Covalent bond ,Biophysics ,biology.protein ,Avidity ,Surface plasmon resonance ,030304 developmental biology - Abstract
Multivalent interactions between complex carbohydrates and oligomeric C-type lectins govern a wide range of immune responses. Up to date, standard SPR (surface plasmon resonance) competitive assays have largely been to evaluate binding properties from monosaccharide units (low affinity, mM) to multivalent elemental antagonists (moderate affinity, µM). Herein, we report typical case-studies of SPR competitive assays showing that they underestimate the potency of glycoclusters to inhibit the interaction between DC-SIGN and immobilized glycoconjugates. This paper describes the design and implementation of a SPR direct interaction over DC-SIGN oriented surfaces, extendable to other C-type lectin surfaces as such Langerin. This setup provides a microscopic overview of intrinsic avidity generation emanating simultaneously from multivalent glycoclusters and from DC-SIGN tetramers that are organized in nanoclusters on the cell membrane. For this purpose, covalent biospecific capture of DC-SIGN via StreptagII /StrepTactin interaction offers the preservation of tetrameric DC-SIGN and the accessibility/functionality of all active sites. From the tested glycoclusters libraries, we demonstrated that the scaffold architecture, the valency and the glycomimetic-based ligand are crucial to reach nanomolar affinities for DC-SIGN. The glycocluster 3.D illustrates the tightest binding partner in this set for a DC-SIGN surface (Kd= 18 nM). Moreover, the selectivity at monovalent scale of glycomimetic D can be easily analyzed at multivalent scale comparing its binding over different C-type lectin immobilized surfaces. This approach may give rise to novel insights into the multivalent binding mechanisms responsible to avidity and make a major contribution to the full characterization of the binding potency of promising specific and multivalent immunomodulators.
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- 2019
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26. Chemoenzymatic Synthesis of N-glycan Positional Isomers and Evidence for Branch Selective Binding by Monoclonal Antibodies and Human C-type Lectin Receptors
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Michel Thépaut, Cornelis H. Hokke, Begoña Echeverria, Niels-Christian Reichardt, Julie Pham, Silvia Achilli, Sonia Serna, Corinne Vivès, Franck Fieschi, CIC biomaGUNE, Glycotechnology Laboratory, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Department of Parasitology, Leiden University Medical Center (LUMC), Universiteit Leiden-Universiteit Leiden, Biomedical Research Networking Center in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Instituto de Salud Carlos III [Madrid] (ISC)-ministerio de ciencia e innovacion, Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)
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0301 basic medicine ,Glycan ,medicine.drug_class ,Receptors, Cell Surface ,Monoclonal antibody ,01 natural sciences ,Biochemistry ,03 medical and health sciences ,Mice ,Isomerism ,C-type lectin ,Polysaccharides ,Structural isomer ,medicine ,Animals ,Humans ,Lectins, C-Type ,Receptor ,Innate immune system ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,010405 organic chemistry ,Chemistry ,Lectin ,Antibodies, Monoclonal ,General Medicine ,0104 chemical sciences ,carbohydrates (lipids) ,030104 developmental biology ,Receptors, Mitogen ,biology.protein ,Biocatalysis ,Molecular Medicine ,Antibody ,Cell Adhesion Molecules - Abstract
Here, we describe a strategy for the rapid preparation of pure positional isomers of complex N-glycans to complement an existing array comprising a larger number of N-glycans and smaller glycan structures. The expanded array was then employed to study context-dependent binding of structural glycan fragments by monoclonal antibodies and C-type lectins. A partial enzymatic elongation of semiprotected core structures was combined with the protecting-group-aided separation of positional isomers by preparative HPLC. This methodology, which avoids the laborious chemical differentiation of antennae, was employed for the preparation of eight biantennary N-glycans with Galβ1,4GlcNAc (LN), GalNAcβ1,4GlcNAc (LDN), and GalNAcβ1,4[Fucα1,3]GlcNAc (LDNF) motifs presented on either one or both antennae. Screening of the binding specificities of three anti-LeX monoclonal IgM antibodies raised against S. mansoni glycans and three C-type lectin receptors of the innate immune system, namely DC-SIGN, DC-SIGNR, and LSECtin, revealed a surprising context-dependent fine specificity for the recognition of the glycan motifs. Moreover, we observed a striking selection of one individual positional isomer over the other by the C-type lectins tested, underscoring the biological relevance of the structural context of glycan elements in molecular recognition.
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- 2018
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27. A Two-component NADPH Oxidase (NOX)-like System in Bacteria Is Involved in the Electron Transfer Chain to the Methionine Sulfoxide Reductase MsrP
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Vincent Nivière, Christelle Caux-Thang, Antoine Picciocchi, Jérôme Dupuy, Jean-Pierre Andrieu, Corinne Vivès, Franck Fieschi, Isabelle Petit-Hartlein, Céline Juillan-Binard, Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie et Biologie des Métaux (LCBM - UMR 5249), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)
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0301 basic medicine ,Protein family ,030106 microbiology ,Green Fluorescent Proteins ,Escherichia coli (E. coli) ,medicine.disease_cause ,Biochemistry ,flavin reductase Fre ,Electron Transport ,YedZ ,03 medical and health sciences ,chemistry.chemical_compound ,Flavin reductase ,medicine ,Escherichia coli ,membrane protein ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Amino Acid Sequence ,heme ,Molecular Biology ,Heme ,Histidine ,Sequence Homology, Amino Acid ,NADPH oxidase ,NADPH Oxidases ,Cell Biology ,Periplasmic space ,Surface Plasmon Resonance ,electron transfer ,[SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology ,030104 developmental biology ,chemistry ,Membrane protein ,flavin mononucleotide (FMN) ,Methionine Sulfoxide Reductases ,Mutagenesis, Site-Directed ,Enzymology ,Methionine sulfoxide reductase ,Spectrophotometry, Ultraviolet ,methionine sulfide reductase MsrPQ - Abstract
International audience; MsrPQ is a newly identified methionine sulfoxide reductase system found in bacteria, which appears to be specifically involved in the repair of periplasmic proteins oxidized by hypochlorous acid. It involves two proteins: a periplasmic one, MsrP, previously named YedY, carrying out the Msr activity, and MsrQ, an integral b-type heme membrane-spanning protein, which acts as the specific electron donor to MsrP. MsrQ, previously named YedZ, was mainly characterized by bioinformatics as a member of the FRD superfamily of heme-containing membrane proteins, which include the NADPH oxidase proteins (NOX/DUOX). Here we report a detailed biochemical characterization of the MsrQ protein from Escherichia coli We optimized conditions for the overexpression and membrane solubilization of an MsrQ-GFP fusion and set up a purification scheme allowing the production of pure MsrQ. Combining UV-visible spectroscopy, heme quantification, and site-directed mutagenesis of histidine residues, we demonstrated that MsrQ is able to bind two b-type hemes through the histidine residues conserved between the MsrQ and NOX protein families. In addition, we identify the E. coli flavin reductase Fre, which is related to the dehydrogenase domain of eukaryotic NOX enzymes, as an efficient cytosolic electron donor to the MsrQ heme moieties. Cross-linking experiments as well as surface Plasmon resonance showed that Fre interacts with MsrQ to form a specific complex. Taken together, these data support the identification of the first prokaryotic two-component protein system related to the eukaryotic NOX family and involved in the reduction of periplasmic oxidized proteins.
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- 2017
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28. Facile access to pseudo-thio - 1,2 - dimannoside, a new glycomimetic DC-SIGN antagonist
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Silvia Achilli, Franck Fieschi, Anna Bernardi, Francesca Vasile, Alice Tamburrini, Cinzia Colombo, Corinne Vivès, Sara Sattin, Dipartimento di Chimica, Università degli Studi di Milano [Milano] (UNIMI), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), MP3, SPR, European Project: Immunoshape, Università degli Studi di Milano = University of Milan (UNIMI), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
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Models, Molecular ,Stereochemistry ,Clinical Biochemistry ,Disaccharide ,Pharmaceutical Science ,Thio ,Epoxide ,Receptors, Cell Surface ,010402 general chemistry ,01 natural sciences ,Biochemistry ,DC-SIGN ,chemistry.chemical_compound ,Glycomimetic ,Enzymatic hydrolysis ,Drug Discovery ,Humans ,Molecule ,Lectins, C-Type ,Molecular Biology ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,010405 organic chemistry ,Chemistry ,Organic Chemistry ,Antagonist ,Surface Plasmon Resonance ,Mannobioside ,0104 chemical sciences ,Thioglycosides ,Drug Design ,Mannosides ,Glycomimetics ,biology.protein ,Molecular Medicine ,Cell Adhesion Molecules ,NOESY - Abstract
International audience; The synthesis and conformational analysis of pseudo-thio-1,2-dimannoside are described. This molecule mimics mannobioside (Manα(1,2)Man) and is an analog of pseudo-1,2-dimannoside, with expected increased stability to enzymatic hydrolysis. A short and efficient synthesis was developed based on an epoxide ring-opening reaction by a mannosyl thiolate, generated in situ from the corresponding thioacetate. NMR-NOESY studies supported by MM3(∗) calculations showed that the pseudo-thio-1,2-dimannoside shares the conformational behavior of the pseudo-1,2-dimannoside and is a structural mimic of the natural disaccharide. Its affinity for DC-SIGN was measured by SPR and found to be comparable to the corresponding O-linked analog, offering good opportunities for further developments.
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- 2017
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29. DC-SIGN Neck Domain Is a pH-sensor Controlling Oligomerization
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Corinne Vivès, Dominique Durand, Michel Thépaut, Patrice Vachette, Christine Ebel, Georges Tabarani, David Stroebel, and Franck Fieschi
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Circular dichroism ,Ligand ,Cell adhesion molecule ,Chemistry ,Cell Biology ,Plasma protein binding ,Biochemistry ,Crystallography ,Tetramer ,Ionic strength ,Biophysics ,Extracellular ,Ultracentrifuge ,Molecular Biology - Abstract
DC-SIGN is a C-type lectin receptor of dendritic cells and is involved in the early stages of numerous infectious diseases. DC-SIGN is organized into a tetramer enabling multivalent interaction with pathogens. Once formed, the DC-SIGN-pathogen complex can be internalized into compartments of increasing acidity. We have studied the pH dependence of the oligomerization state and conformation of the entire extracellular domain and neck region. We present evidence for equilibrium between the monomeric and tetrameric states of the extracellular domain, which exhibits a marked dependence with respect to both pH and ionic strength. Using solution x-ray scattering we have obtained a molecular envelope of the extracellular domain in which a model has been built. Our results highlight the central role of the neck domain in the pH-sensitive control of the oligomerization state, in the extended conformation of the protein, and in carbohydrate recognition domain organization and presentation. This work opens new insight into the molecular mechanism of ligand release and points to new avenues to block the first step of this important infection pathway.
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- 2009
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30. Conformational changes in p47phoxupon activation highlighted by mass spectrometry coupled to hydrogen/deuterium exchange and limited proteolysis
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Franck Fieschi, Corinne Vivès, Mathieu Castellan, Petr Man, Julien Marcoux, Eric Forest, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Institute of Microbiology, Czech Academy of Sciences [Prague] (CAS), Institut de Biosciences et de Biotechnologies de Grenoble (ex-IRTSV) (BIG), Institut National de la Santé et de la Recherche Médicale (INSERM)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de pharmacologie et de biologie structurale (IPBS), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Acad. of Sci. of the Czech Republic, Prague, Institut National de la Santé et de la Recherche Médicale (INSERM)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Université Grenoble Alpes (UGA), Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])
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H/D exchange ,Conformational change ,Neutrophils ,Protein Conformation ,Proteolysis ,Electrospray ionization ,Biophysics ,Mass spectrometry ,Models, Biological ,Biochemistry ,Mass Spectrometry ,03 medical and health sciences ,Limited proteolysis ,Structural Biology ,Genetics ,medicine ,Humans ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Molecular Biology ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,NADPH oxidase ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,biology ,medicine.diagnostic_test ,Hydrolysis ,030302 biochemistry & molecular biology ,Deuterium Exchange Measurement ,NADPH Oxidases ,p47phox ,Cell Biology ,Kinetics ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM] ,Enzyme ,chemistry ,biology.protein ,Phosphorylation ,Hydrogen–deuterium exchange - Abstract
The neutrophil NADPH oxidase is an enzymatic complex involved in innate immunity. Phosphorylation of p47(phox) promotes its translocation with p67(phox) and p40(phox), followed by membrane interaction and assembly with flavocytochrome b(558) into a functional complex. To characterise p47(phox) conformational changes during activation, we used wild-type and the S303/304/328E triple mutant mimicking the phosphorylated state. Hydrogen/deuterium exchange and limited proteolysis coupled to mass spectrometry were used to discriminate between the various structural models. An increase in deuteration confirmed that p47(phox) adopts an open and more flexible conformation after activation. Limited proteolysis correlated this change with increased auto-inhibitory region (AIR) accessibility. These results establish a structural link between the AIR release and the exposure of the Phox homology (PX) domain.
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- 2009
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31. Overproduction, purification and preliminary crystallographic analysis of the carbohydrate-recognition domain of human langerin
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Richard Kahn, Michel Thépaut, Corinne Vivès, Franck Fieschi, and Guillaume Pompidor
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Langerin ,Glycoconjugate ,Birbeck granules ,media_common.quotation_subject ,Carbohydrates ,Biophysics ,macromolecular substances ,Crystallography, X-Ray ,Biochemistry ,Antigens, CD ,Structural Biology ,Genetics ,Humans ,Lectins, C-Type ,Avidity ,Internalization ,Overproduction ,DNA Primers ,Mannan-binding lectin ,media_common ,chemistry.chemical_classification ,Base Sequence ,integumentary system ,biology ,Lectin ,equipment and supplies ,Condensed Matter Physics ,eye diseases ,Crystallography ,Mannose-Binding Lectins ,chemistry ,Crystallization Communications ,biological sciences ,biology.protein ,bacteria ,Crystallization - Abstract
Langerin, a lectin that is specific to Langerhans cells, interacts with glycoconjugates through its carbohydrate-recognition domain (CRD). This carbohydrate binding occurs by an avidity-based mechanism that is enabled by the neck domain responsible for trimerization. Langerin binds HIV through its CRD and thus plays a protective role against its propagation by the internalization of virions in Birbeck granules. Here, the overproduction, purification and crystallization of the langerin CRD is reported. Crystals obtained by the hanging-drop vapour-diffusion method allowed the collection of a complete data set to 1.5 A resolution and belonged to the tetragonal space group P4(2), with unit-cell parameters a = b = 79.55, c = 90.14 A.
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- 2008
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32. Alteration of the langerin oligomerization state affects Birbeck granule formation
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Michel Thépaut, Franck Fieschi, Patrice Vachette, Eric Chabrol, Colette Dezutter-Dambuyant, Corinne Vivès, Richard Kahn, Dominique Durand, Jenny Valladeau-Guilemond, Julien Marcoux, Institut de biologie structurale ( IBS - UMR 5075 ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Centre de Recherche en Cancérologie de Lyon ( CRCL ), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de biochimie et biophysique moléculaire et cellulaire ( IBBMC ), Université Paris-Sud - Paris 11 ( UP11 ) -Centre National de la Recherche Scientifique ( CNRS ), SPR, Spectro de masse, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de biochimie et biophysique moléculaire et cellulaire (IBBMC), Université Paris-Sud - Paris 11 (UP11)-Centre National de la Recherche Scientifique (CNRS), Thomas, Frank, Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)
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Models, Molecular ,Langerin ,[SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Birbeck granules ,Mutant ,Biophysics ,Context (language use) ,Plasma protein binding ,HIV Envelope Protein gp120 ,medicine.disease_cause ,Crystallography, X-Ray ,Cytoplasmic Granules ,Transfection ,Cell Line ,Mannans ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Virion binding ,Antigens, CD ,Scattering, Small Angle ,medicine ,Animals ,Humans ,Lectins, C-Type ,Chromatography, High Pressure Liquid ,030304 developmental biology ,Mannan-binding lectin ,0303 health sciences ,Mutation ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Fibroblasts ,3. Good health ,Cell biology ,Protein Structure, Tertiary ,Cross-Linking Reagents ,Mannose-Binding Lectins ,Biochemistry ,Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ,biology.protein ,Mutant Proteins ,Protein Multimerization ,Proteins and Nucleic Acids ,030215 immunology ,Protein Binding ,[ SDV.BBM.BS ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM] - Abstract
International audience; Langerin, a trimeric C-type lectin specifically expressed in Langerhans cells, has been reported to be a pathogen receptor through the recognition of glycan motifs by its three carbohydrate recognition domains (CRD). In the context of HIV-1 (human immunodeficiency virus-1) transmission, Langerhans cells of genital mucosa play a protective role by internalizing virions in Birbeck Granules (BG) for elimination. Langerin (Lg) is directly involved in virion binding and BG formation through its CRDs. However, nothing is known regarding the mechanism of langerin assembly underlying BG formation. We investigated at the molecular level the impact of two CRD mutations, W264R and F241L, on langerin structure, function, and BG assembly using a combination of biochemical and biophysical approaches. Although the W264R mutation causes CRD global unfolding, the F241L mutation does not affect the overall structure and gp120 (surface HIV-1 glycoprotein of 120 kDa) binding capacities of isolated Lg-CRD. In contrast, this mutation induces major functional and structural alterations of the whole trimeric langerin extracellular domain (Lg-ECD). As demonstrated by small-angle x-ray scattering comparative analysis of wild-type and mutant forms, the F241L mutation perturbs the oligomerization state and the global architecture of Lg-ECD. Correlatively, despite conserved intrinsic lectin activity of the CRD, avidity property of Lg-ECD is affected as shown by a marked decrease of gp120 binding. Beyond the change of residue itself, the F241L mutation induces relocation of the K200 side chain also located within the interface between protomers of trimeric Lg-ECD, thereby explaining the defective oligomerization of mutant Lg. We conclude that not only functional CRDs but also their correct spatial presentation are critical for BG formation as well as gp120 binding.
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- 2015
33. p47 phox Molecular Activation for Assembly of the Neutrophil NADPH Oxidase Complex
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Franck Fieschi, Corinne Vivès, Petr Man, Julien Marcoux, Eric Forest, Isabelle Petit-Haertlein, Laboratoire des Protéines Membranaires (LPM), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institute of Microbiology, Czech Academy of Sciences [Prague] (CAS), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
Stereochemistry ,Neutrophils ,Mutation, Missense ,Biochemistry ,SH3 domain ,Protein Structure, Secondary ,src Homology Domains ,03 medical and health sciences ,Enzyme activator ,NADPH oxidase complex ,Animals ,Humans ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,NADPH oxidase ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Chemistry ,030302 biochemistry & molecular biology ,NADPH Oxidases ,Cell Biology ,PX domain ,Enzyme Activation ,Cytosol ,Amino Acid Substitution ,Protein Structure and Folding ,Biophysics ,biology.protein ,Hydrogen–deuterium exchange ,Proto-oncogene tyrosine-protein kinase Src - Abstract
International audience; The p47phox cytosolic factor from neutrophilic NADPH oxidase has always been resistant to crystallogenesis trials due to its modular organization leading to relative flexibility. Hydrogen/deuterium exchange coupled to mass spectrometry was used to obtain structural information on the conformational mechanism that underlies p47phox activation. We confirmed a relative opening of the protein with exposure of the SH3 Src loops thatare known to bind p22phox upon activation. A new surface was shown to be unmasked after activation, representing a potential autoinhibitory surface that may block the interaction of the PX domain with the membrane in the resting state. Within this surface, we identified 2 residues involved in the interaction with the PX domain. The double mutant R162A/D166A showed a higher affinity for specific phospholipids but none for the C-terminalpart of p22phox, reflecting an intermediate conformation between the autoinhibited and activated forms.
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- 2010
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34. Leukotriene BLT2 receptor monomers activate the G(i2) GTP-binding protein more efficiently than dimers.: G protein activation by monomeric and dimeric GPCRs
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Corinne Vivès, Jean-Louis Banères, Gérald Gaibelet, Emilie Detouillon, Thierry Durroux, Christiane Mendre, Laure Arcemisbéhère, Sébastien Granier, Marie-Noëlle Balestre, Tuhinadri Sen, Hélène Orcel, Franck Fieschi, Laure Boudier, Bernard Mouillac, Marjorie Damian, Rita Rahmeh, Institut de Génomique Fonctionnelle (IGF), Université Montpellier 1 (UM1)-Université Montpellier 2 - Sciences et Techniques (UM2)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Institut des Biomolécules Max Mousseron [Pôle Chimie Balard] (IBMM), 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), This work was granted by INSERM, CNRS, the European community (#LSHB-CT-2003-503337), the ANRS foundation (#AC14), the French Ministry of Research (ACI BCMS #328) and the National Agency of Research (ANR 06-BLAN-0087). L.A. was the recipient of a PhD fellowship from the French Ligue contre le Cancer. The strategy described here has been patented by INSERM/CNRS (patent 03/07411 and WO 2004/113539 A3), ANR-06-BLAN-0087,GPCR dimers,Molecular and dynamic aspects of G-protein coupled receptor dimers functioning(2006), Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université de Montpellier (UM)-Ecole Nationale Supérieure de Chimie de Montpellier (ENSCM)
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MESH: Signal Transduction ,MESH: Fluorescence Resonance Energy Transfer ,G protein ,Receptors, Leukotriene B4 ,GTP-Binding Protein beta Subunits ,GTP-Binding Protein alpha Subunits, Gi-Go ,Biochemistry ,MESH: GTP-Binding Protein beta Subunits ,03 medical and health sciences ,0302 clinical medicine ,GTP-Binding Protein gamma Subunits ,Protein A/G ,Fluorescence Resonance Energy Transfer ,Humans ,MESH: Protein Binding ,5-HT5A receptor ,Molecular Biology ,030304 developmental biology ,G alpha subunit ,G protein-coupled receptor ,0303 health sciences ,MESH: Humans ,biology ,MESH: Integrin alphaV ,Chemistry ,MESH: Protein Multimerization ,Binding protein ,MESH: GTP-Binding Protein alpha Subunits, Gi-Go ,Cell Biology ,Integrin alphaV ,MESH: Receptors, Leukotriene B4 ,[SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences ,biology.protein ,Protein G ,Protein Multimerization ,030217 neurology & neurosurgery ,MESH: GTP-Binding Protein gamma Subunits ,Protein Binding ,Signal Transduction - Abstract
International audience; Accumulating evidence indicates that G protein-coupled receptors can assemble as dimers/oligomers but the role of this phenomenon in G protein coupling and signaling is not yet clear. We have used the purified leukotriene B(4) receptor BLT2 as a model to investigate the capacity of receptor monomers and dimers to activate the adenylyl cyclase inhibitory G(i2) protein. For this, we overexpressed the recombinant receptor as inclusion bodies in the Escherichia coli prokaryotic system, using a human alpha(5) integrin as a fusion partner. This strategy allowed the BLT2 as well as several other G protein-coupled receptors from different families to be produced and purified in large amounts. The BLT2 receptor was then successfully refolded to its native state, as measured by high-affinity LTB(4) binding in the presence of the purified G protein G alpha(i2). The receptor dimer, in which the two protomers displayed a well defined parallel orientation as assessed by fluorescence resonance energy transfer, was then separated from the monomer. Using two methods of receptor-catalyzed guanosine 5'-3-O-(thio)triphosphate binding assay, we clearly demonstrated that monomeric BLT2 stimulates the purified G alpha(i2) beta(1) gamma(2) protein more efficiently than the dimer. These data suggest that assembly of two BLT2 protomers into a dimer results in the reduced ability to signal.
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- 2010
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35. Investigating alternative acidic proteases for H/D exchange coupled to mass spectrometry: Plasmepsin 2 but not plasmepsin 4 is active under quenching conditions
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Corinne Vivès, Eric Thierry, Luca Signor, Eric Forest, Julien Marcoux, Franck Fieschi, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'Informatique, Signaux, et Systèmes de Sophia Antipolis (I3S), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Université Nice Sophia Antipolis (1965 - 2019) (UNS)
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Proteases ,Molecular Sequence Data ,Plasmodium falciparum ,Plasmepsin ,Proteomics ,Mass spectrometry ,01 natural sciences ,Mass Spectrometry ,Substrate Specificity ,03 medical and health sciences ,Hemoglobins ,Pepsin ,Structural Biology ,[CHIM.ANAL]Chemical Sciences/Analytical chemistry ,Animals ,Aspartic Acid Endopeptidases ,Humans ,Amino Acid Sequence ,Horses ,Spectroscopy ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Chromatography ,Quenching (fluorescence) ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Myoglobin ,010401 analytical chemistry ,Deuterium Exchange Measurement ,Proteins ,Pepsin A ,Recombinant Proteins ,0104 chemical sciences ,Enzyme ,chemistry ,Biochemistry ,biology.protein ,Hydrogen–deuterium exchange ,Cattle ,Chromatography, Liquid - Abstract
Structural studies of proteins by hydrogen/deuterium exchange coupled to mass spectrometry (DXMS) require the use of proteases working at acidic pH and low temperatures. The spatial resolution of this technique can be improved by combining several acidic proteases, each generating a set of different peptides. Three commercial aspartic proteases are used, namely, pepsin, and proteases XIII and XVIII. However, given their low purity, high enzyme/protein ratios have to be used with proteases XIII and XVIII. In the present work, we investigate the activity of two alternative acidic proteases from Plasmodium falciparum under different pH and temperature conditions. Peptide mapping of four different proteins after digestion with pepsin, plasmepsin 2 (PSM2), and plasmepsin 4 (PSM4) were compared. PSM4 is inactive at pH 2.2 and 0 degrees C, making it unusable for DXMS studies. However, PSM2 showed low but reproducible activity under DXMS conditions. It displayed no substrate specificity and, like pepsin, no strict sequence specificity. Altogether, these results show that PSM2 but not PSM4 is a potential new tool for DXMS studies.
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- 2010
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36. NADPH oxidase activator p67(phox) behaves in solution as a multidomain protein with semi-flexible linkers
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Dominique Durand, Javier Pérez, Dominique Cannella, Eva Pebay-Peyroula, Franck Fieschi, Patrice Vachette, and Corinne Vivès
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NADPH oxidase ,Modular structure ,biology ,Activator (genetics) ,Chemistry ,Computational Biology ,RAC1 ,Phosphoproteins ,Flavocytochrome b ,Protein Structure, Tertiary ,Cytosol ,Protein structure ,Biochemistry ,X-Ray Diffraction ,Structural Biology ,NADPH oxidase complex ,Scattering, Small Angle ,biology.protein ,Biophysics - Abstract
The NADPH oxidase complex is involved in the destruction of phagocytosed pathogens through the production of reactive oxygen species. This activatable complex consists of a membranous heterodimeric flavocytochrome b, a small G-protein Rac1/Rac2 and cytosolic factors, p47(phox), p67(phox) and p40(phox). p67(phox), due to its modular structure, is the NADPH oxidase component for which global structure information is most scarce despite its mandatory role in activation and its central position in the whole complex organization. Indeed, p67(phox) is the only factor establishing interaction with all others. In this study, we report the SAXS analysis of p67(phox). Our data reveals that p67(phox) behaves as a multidomain protein with semi-flexible linkers. On the one hand, it appears to be a very elongated molecule with its various domains organized as beads on a string. Linkers are predicted to be partially or mainly unstructured and features of our experimental data do point towards inter-domain flexibility. On the other hand, our work also suggests that the protein is not as extended as unstructured linkers could allow, thereby implying the existence of intra-molecular interactions within p67(phox). We suggest that the dual character of p67(phox) conformation in solution is central to ensure the numerous interactions to be accommodated.
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- 2009
37. DC-SIGN neck domain is a pH-sensor controlling oligomerization: SAXS and hydrodynamic studies of extracellular domain
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Georges, Tabarani, Michel, Thépaut, David, Stroebel, Christine, Ebel, Corinne, Vivès, Patrice, Vachette, Dominique, Durand, and Franck, Fieschi
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Sequence Homology, Amino Acid ,Circular Dichroism ,X-Rays ,Molecular Sequence Data ,Molecular Conformation ,Receptors, Cell Surface ,Hydrogen-Ion Concentration ,Ligands ,Extracellular Matrix ,ddc:570 ,Protein Structure and Folding ,Humans ,Scattering, Radiation ,Lectins, C-Type ,Amino Acid Sequence ,Cloning, Molecular ,Cell Adhesion Molecules ,Ultracentrifugation ,Protein Binding - Abstract
The journal of biological chemistry 284, 21229-21240 (2009). doi:10.1074/jbc.M109.021204, Published by Soc., Bethesda, Md.
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- 2009
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38. Structural studies of langerin and birbeck granule: a macromolecular organization model
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Jenny Valladeau, Carine Monnier, Colette Dezutter-Dambuyant, Corinne Vivès, Franck Fieschi, Sem Saeland, Michel Thépaut, Richard Kahn, Christine Ebel, Bertrand Arnou, Anne Imberty, Alessandra Nurisso, inconnu, Inconnu, Centre de Recherches sur les Macromolécules Végétales (CERMAV), and Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)
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Models, Molecular ,Langerhans cell ,Langerin ,Protein Conformation ,Birbeck granules ,media_common.quotation_subject ,Molecular Sequence Data ,Cytoplasmic Granules ,Transfection ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Cell Line ,03 medical and health sciences ,Immune system ,Oligosaccharide binding ,Antigens, CD ,Organelle ,Extracellular ,medicine ,Animals ,Humans ,Lectins, C-Type ,Amino Acid Sequence ,Internalization ,ComputingMilieux_MISCELLANEOUS ,030304 developmental biology ,media_common ,0303 health sciences ,integumentary system ,biology ,Dendritic Cells ,3. Good health ,0104 chemical sciences ,Cell biology ,Mannose-Binding Lectins ,medicine.anatomical_structure ,Langerhans Cells ,biology.protein - Abstract
Dendritic cells, a sentinel immunity cell lineage, include different cell subsets that express various C-type lectins. For example, epidermal Langerhans cells express langerin, and some dermal dendritic cells express DC-SIGN. Langerin is a crucial component of Birbeck granules, the Langerhans cell hallmark organelle, and may have a preventive role toward HIV, by its internalization into Birbeck granules. Since langerin carbohydrate recognition domain (CRD) is crucial for HIV interaction and Birbeck granule formation, we produced the CRD of human langerin and solved its structure at 1.5 A resolution. On this basis gp120 high-mannose oligosaccharide binding has been evaluated by molecular modeling. Hydrodynamic studies reveal a very elongated shape of recombinant langerin extracellular domain (ECD). A molecular model of the langerin ECD, integrating the CRD structure, has been generated and validated by comparison with hydrodynamic parameters. In parallel, Langerhans cells were isolated from human skin. From their analysis by electron microscopy and the langerin ECD model, an ultrastructural organization is proposed for Birbeck granules. To delineate the role of the different langerin domains in Birbeck granule formation, we generated truncated and mutated langerin constructs. After transfection into a fibroblastic cell line, we highlighted, in accordance with our model, the role of the CRD in the membrane zipping occurring in BG formation as well as some contribution of the cytoplasmic domain. Finally, we have shown that langerin ECD triggering with a specific mAb promotes global rearrangements of LC morphology. Our results open the way to the definition of a new membrane deformation mechanism.
- Published
- 2009
39. Mannose hyperbranched dendritic polymers interact with clustered organization of DC-SIGN and inhibit gp120 binding
- Author
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Franck Fieschi, Hugues Lortat-Jacob, Javier Rojo, Georges Tabarani, Corinne Vivès, Christine Ebel, José J. Reina, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Glycosystems Laboratory, Instituto de Investigaciones Químicas (IIQ), Universidad de Sevilla / University of Sevilla-Consejo Superior de Investigaciones Científicas [Madrid] (CSIC)-Universidad de Sevilla / University of Sevilla-Consejo Superior de Investigaciones Científicas [Madrid] (CSIC), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Consejo Superior de Investigaciones Científicas [Madrid] (CSIC)-Universidad de Sevilla-Consejo Superior de Investigaciones Científicas [Madrid] (CSIC)-Universidad de Sevilla
- Subjects
Mannose ,HIV Envelope Protein gp120 ,01 natural sciences ,Biochemistry ,chemistry.chemical_compound ,MESH: HIV Envelope Protein gp120 ,Biopolymers ,Structural Biology ,Surface plasmon resonance ,Receptor ,MESH: Receptors, Cell Surface ,chemistry.chemical_classification ,0303 health sciences ,biology ,Molecular Structure ,Antiviral compound ,3. Good health ,DC-SIGN ,Solutions ,MESH: Biopolymers ,MESH: Mannose ,Multivalency ,MESH: Cell Adhesion Molecules ,Integrin ,MESH: Molecular Structure ,Biophysics ,Carbohydrates ,Receptors, Cell Surface ,MESH: Solutions ,010402 general chemistry ,03 medical and health sciences ,Genetics ,Lectins, C-Type ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Molecular Biology ,030304 developmental biology ,Lipid microdomain ,Lectin ,Cell Biology ,0104 chemical sciences ,gp120 ,chemistry ,biology.protein ,Antiviral compounds ,Glycoprotein ,Cell Adhesion Molecules ,MESH: Lectins, C-Type - Abstract
7 páginas, 5 figuras. Edited by Michael R. Bubb, DC-SIGN (dendritic cell-specific ICAM-3 grabbing non-integrin) is a C-type lectin receptor of dendritic cells and is involved in the initial steps of numerous infectious diseases. Surface plasmon resonance has been used to study the affinity of a glycodendritic polymer with 32 mannoses, to DC-SIGN. This glycodendrimer binds to DC-SIGN surfaces in the submicromolar range. This binding depends on a clustered organization of DC-SIGN mimicking its natural organization as microdomain in the dendritic cells plasma membrane. Moreover, this compound inhibits DC-SIGN binding to the HIV glycoprotein gp120 with an IC50 in the micromolar range and therefore can be considered as a potential antiviral drug., We thank CSIC – 2004FR0027 – (J.R.); FIS – PI030093 – (J.R.) and Ensemble contre le SIDA-Sidaction (F.F.) for funding support. We acknowledge Dr Francisco Véas and Alain Méchulam for the kind gift of gp120 HXB2.
- Published
- 2006
- Full Text
- View/download PDF
40. Intriguing conformation changes associated with the trans/cis isomerization of a prolyl residue in the active site of the DsbA C33A mutant
- Author
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Etienne Ondo-Mbele, Amadou Koné, Laurence Serre, and Corinne Vivès
- Subjects
Models, Molecular ,Proline ,Stereochemistry ,Protein Conformation ,Molecular Sequence Data ,Protein Disulfide-Isomerases ,Peptide binding ,Crystallography, X-Ray ,Isomerism ,Structural Biology ,Oxidoreductase ,Molecular Biology ,chemistry.chemical_classification ,Binding Sites ,biology ,Molecular Structure ,Chemistry ,Escherichia coli Proteins ,Active site ,Periplasmic space ,DsbA ,Catalytic cycle ,Mutation ,biology.protein ,Thioredoxin ,Cysteine - Abstract
Escherichia coli DsbA belongs to the thioredoxin family and catalyzes the formation of disulfide bonds during the folding of proteins in the bacterial periplasm. It active site (C30–P31–H32–C33) consists of a disulfide bridge that is transferred to newly translocated proteins. The work reported here refers to the DsbA mutant termed C33A that retains, towards reduced unfolded thrombin inhibitor, an activity comparable with the wild-type enzyme. Besides, C33A is also able to form a stable covalent complex with DsbB, the membrane protein responsible for maintaining DsbA in its active form. We have determined the crystal structure of C33A at 2.0 A resolution. Although the general architecture of wt DsbA is conserved, we observe the trans / cis isomerization of P31 in the active site and further conformational changes in the so-called “peptide binding groove” region. Interestingly, these modifications involve residues that are specific to DsbA but not to the thioredoxin family fold. The C33A crystal structure exhibits as well a hydrophobic ligand bound close to the active site of the enzyme. The structural analysis of C33A may actually explain the peculiar behavior of this mutant in regards with its interaction with DsbB and thus provides new insights for understanding the catalytic cycle of DsbA.
- Published
- 2004
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