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115 results on '"David Pignol"'

1. Reduction of Protein Bound Methionine Sulfoxide by a Periplasmic Dimethyl Sulfoxide Reductase

Author

Lionel Tarrago, Sandrine Grosse, David Lemaire, Laetitia Faure, Mathilde Tribout, Marina I. Siponen, Mila Kojadinovic-Sirinelli, David Pignol, Pascal Arnoux, and Monique Sabaty

Subjects
dimethyl sulfoxide reductase, enzyme kinetics, methionine sulfoxide, oxidative stress, protein oxidation, protein quality control, Therapeutics. Pharmacology, RM1-950
Abstract

In proteins, methionine (Met) can be oxidized into Met sulfoxide (MetO). The ubiquitous methionine sulfoxide reductases (Msr) A and B are thiol-oxidoreductases reducing MetO. Reversible Met oxidation has a wide range of consequences, from protection against oxidative stress to fine-tuned regulation of protein functions. Bacteria distinguish themselves by the production of molybdenum-containing enzymes reducing MetO, such as the periplasmic MsrP which protects proteins during acute oxidative stress. The versatile dimethyl sulfoxide (DMSO) reductases were shown to reduce the free amino acid MetO, but their ability to reduce MetO within proteins was never evaluated. Here, using model oxidized proteins and peptides, enzymatic and mass spectrometry approaches, we showed that the Rhodobacter sphaeroides periplasmic DorA-type DMSO reductase reduces protein bound MetO as efficiently as the free amino acid L-MetO and with catalytic values in the range of those described for the canonical Msrs. The identification of this fourth type of enzyme able to reduce MetO in proteins, conserved across proteobacteria and actinobacteria, suggests that organisms employ enzymatic systems yet undiscovered to regulate protein oxidation states.

Published
2020
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2. Heterologous expression of membrane proteins: choosing the appropriate host.

Author

Florent Bernaudat, Annie Frelet-Barrand, Nathalie Pochon, Sébastien Dementin, Patrick Hivin, Sylvain Boutigny, Jean-Baptiste Rioux, Daniel Salvi, Daphné Seigneurin-Berny, Pierre Richaud, Jacques Joyard, David Pignol, Monique Sabaty, Thierry Desnos, Eva Pebay-Peyroula, Elisabeth Darrouzet, Thierry Vernet, and Norbert Rolland

Subjects
Medicine, Science
Abstract

BackgroundMembrane proteins are the targets of 50% of drugs, although they only represent 1% of total cellular proteins. The first major bottleneck on the route to their functional and structural characterisation is their overexpression; and simply choosing the right system can involve many months of trial and error. This work is intended as a guide to where to start when faced with heterologous expression of a membrane protein.Methodology/principal findingsThe expression of 20 membrane proteins, both peripheral and integral, in three prokaryotic (E. coli, L. lactis, R. sphaeroides) and three eukaryotic (A. thaliana, N. benthamiana, Sf9 insect cells) hosts was tested. The proteins tested were of various origins (bacteria, plants and mammals), functions (transporters, receptors, enzymes) and topologies (between 0 and 13 transmembrane segments). The Gateway system was used to clone all 20 genes into appropriate vectors for the hosts to be tested. Culture conditions were optimised for each host, and specific strategies were tested, such as the use of Mistic fusions in E. coli. 17 of the 20 proteins were produced at adequate yields for functional and, in some cases, structural studies. We have formulated general recommendations to assist with choosing an appropriate system based on our observations of protein behaviour in the different hosts.Conclusions/significanceMost of the methods presented here can be quite easily implemented in other laboratories. The results highlight certain factors that should be considered when selecting an expression host. The decision aide provided should help both newcomers and old-hands to select the best system for their favourite membrane protein.

Published
2011
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3. Single-step production of a recyclable nanobiocatalyst for organophosphate pesticides biodegradation using functionalized bacterial magnetosomes.

Author

Nicolas Ginet, Romain Pardoux, Géraldine Adryanczyk, Daniel Garcia, Catherine Brutesco, and David Pignol

Subjects
Medicine, Science
Abstract

Enzymes are versatile catalysts in laboratories and on an industrial scale; improving their immobilization would be beneficial to broadening their applicability and ensuring their (re)use. Lipid-coated nano-magnets produced by magnetotactic bacteria are suitable for a universally applicable single-step method of enzyme immobilization. By genetically functionalizing the membrane surrounding these magnetite particles with a phosphohydrolase, we engineered an easy-to-purify, robust and recyclable biocatalyst to degrade ethyl-paraoxon, a commonly used pesticide. For this, we genetically fused the opd gene from Flavobacterium sp. ATCC 27551 encoding a paraoxonase to mamC, an abundant protein of the magnetosome membrane in Magnetospirillum magneticum AMB-1. The MamC protein acts as an anchor for the paraoxonase to the magnetosome surface, thus producing magnetic nanoparticles displaying phosphohydrolase activity. Magnetosomes functionalized with Opd were easily recovered from genetically modified AMB-1 cells: after cellular disruption with a French press, the magnetic nanoparticles are purified using a commercially available magnetic separation system. The catalytic properties of the immobilized Opd were measured on ethyl-paraoxon hydrolysis: they are comparable with the purified enzyme, with K(m) (and k(cat)) values of 58 µM (and 178 s(-1)) and 43 µM (and 314 s(-1)) for the immobilized and purified enzyme respectively. The Opd, a metalloenzyme requiring a zinc cofactor, is thus properly matured in AMB-1. The recycling of the functionalized magnetosomes was investigated and their catalytic activity proved to be stable over repeated use for pesticide degradation. In this study, we demonstrate the easy production of functionalized magnetic nanoparticles with suitably genetically modified magnetotactic bacteria that are efficient as a reusable nanobiocatalyst for pesticides bioremediation in contaminated effluents.

Published
2011
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4. A second actin-like MamK protein in Magnetospirillum magneticum AMB-1 encoded outside the genomic magnetosome island.

Author

Jean-Baptiste Rioux, Nadège Philippe, Sandrine Pereira, David Pignol, Long-Fei Wu, and Nicolas Ginet

Subjects
Medicine, Science
Abstract

Magnetotactic bacteria are able to swim navigating along geomagnetic field lines. They synthesize ferromagnetic nanocrystals that are embedded in cytoplasmic membrane invaginations forming magnetosomes. Regularly aligned in the cytoplasm along cytoskeleton filaments, the magnetosome chain effectively forms a compass needle bestowing on bacteria their magnetotactic behaviour. A large genomic island, conserved among magnetotactic bacteria, contains the genes potentially involved in magnetosome formation. One of the genes, mamK has been described as encoding a prokaryotic actin-like protein which when it polymerizes forms in the cytoplasm filamentous structures that provide the scaffold for magnetosome alignment. Here, we have identified a series of genes highly similar to the mam genes in the genome of Magnetospirillum magneticum AMB-1. The newly annotated genes are clustered in a genomic islet distinct and distant from the known magnetosome genomic island and most probably acquired by lateral gene transfer rather than duplication. We focused on a mamK-like gene whose product shares 54.5% identity with the actin-like MamK. Filament bundles of polymerized MamK-like protein were observed in vitro with electron microscopy and in vivo in E. coli cells expressing MamK-like-Venus fusions by fluorescence microscopy. In addition, we demonstrate that mamK-like is transcribed in AMB-1 wild-type and DeltamamK mutant cells and that the actin-like filamentous structures observed in the DeltamamK strain are probably MamK-like polymers. Thus MamK-like is a new member of the prokaryotic actin-like family. This is the first evidence of a functional mam gene encoded outside the magnetosome genomic island.

Published
2010
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6. Repeated horizontal gene transfers triggered parallel evolution of magnetotaxis in two evolutionary divergent lineages of magnetotactic bacteria

Author

Christopher T. Lefèvre, Caroline L. Monteil, Béatrice Alonso, Denis S. Grouzdev, Zoé Rouy, Stéphane Cruveiller, David Pignol, Nicolas Ginet, Guy Perrière, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), ANR-18-CE31-0003,SIGMAG,SIGNATURE DES MAGNETITES PRODUITES PAR LES BACTERIES MAGNETOTACTIQUES : PERSPECTIVES CHIMIQUES ET ISOTOPIQUES(2018), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Gene Transfer, Horizontal, Magnetotactic bacteria, [SDV.BID.SPT]Life Sciences [q-bio]/Biodiversity/Systematics, Phylogenetics and taxonomy, Microbiology, Article, 03 medical and health sciences, Phylogenetics, Gram-Negative Bacteria, Magnetotaxis, Magnetospirillum, Ecology, Evolution, Behavior and Systematics, ComputingMilieux_MISCELLANEOUS, Alphaproteobacteria, 030304 developmental biology, 0303 health sciences, Experimental evolution, Bacteria, biology, 030306 microbiology, Clonal interference, [SDV.BID.EVO]Life Sciences [q-bio]/Biodiversity/Populations and Evolution [q-bio.PE], biology.organism_classification, Evolutionary biology, Parallel evolution, Adaptation
Abstract

Under the same selection pressures, two genetically divergent populations may evolve in parallel toward the same adaptive solutions. Here, we hypothesized that magnetotaxis (i.e., magnetically guided chemotaxis) represents a key adaptation to micro-oxic habitats in aquatic sediments and that its parallel evolution homogenized the phenotypes of two evolutionary divergent clusters of freshwater spirilla. All magnetotactic bacteria affiliated to the Magnetospirillum genus (Alphaproteobacteria class) biomineralize the same magnetic particle chains and share highly similar physiological and ultrastructural features. We looked for the processes that could have contributed at shaping such an evolutionary pattern by reconciling species and gene trees using newly sequenced genomes of Magnetospirillum related bacteria. We showed that repeated horizontal gene transfers and homologous recombination of entire operons contributed to the parallel evolution of magnetotaxis. We propose that such processes could represent a more parsimonious and rapid solution for adaptation compared with independent and repeated de novo mutations, especially in the case of traits as complex as magnetotaxis involving tens of interacting proteins. Besides strengthening the idea about the importance of such a function in micro-oxic habitats, these results reinforce previous observations in experimental evolution suggesting that gene flow could alleviate clonal interference and speed up adaptation under some circumstances.

Published
2020

7. Transformation Cycle of Magnetosomes in Human Stem Cells: From Degradation to Biosynthesis of Magnetic Nanoparticles Anew

Author

Christine Péchoux, Claire Wilhelm, Sandra Prévéral, Christopher T. Lefèvre, Aurore Van de Walle, David Pignol, Nicolas Menguy, Aida Serrano, Alberto Curcio, Ana Espinosa, Matière et Systèmes Complexes (MSC), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), CSIC, Inst Ciencia Mat Madrid, Cantoblanco, E-28049 Madrid, Spain, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Unité de recherche génomique et physiologie de la lactation (GPL), Institut National de la Recherche Agronomique (INRA), Génétique Animale et Biologie Intégrative (GABI), AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Université Paris-Saclay, Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de recherche pour le développement [IRD] : UR206-Centre National de la Recherche Scientifique (CNRS), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Comunidad de Madrid 2018-T1/1ND-100S, European Project: 648779,H2020,ERC-2014-CoG,MaTissE(2015), Matière et Systèmes Complexes (MSC (UMR_7057)), Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Université Paris-Saclay-AgroParisTech-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de recherche pour le développement [IRD] : UR206-Muséum national d'Histoire naturelle (MNHN)-Centre National de la Recherche Scientifique (CNRS)

Subjects
magnetite, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, Magnetotactic bacteria, Surface Properties, Magnetism, Magnetosome, General Physics and Astronomy, Nanoparticle, 02 engineering and technology, 010402 general chemistry, 01 natural sciences, magnetosomes, biodegradation, ferrihydrite, chemistry.chemical_compound, Ferrihydrite, Humans, General Materials Science, Particle Size, Magnetite Nanoparticles, Lipid bilayer, Cells, Cultured, Magnetite, Chemistry, General Engineering, Mesenchymal Stem Cells, 021001 nanoscience & nanotechnology, equipment and supplies, 0104 chemical sciences, Biophysics, Magnetic nanoparticles, nanoparticles, biosynthesis, 0210 nano-technology
Abstract

International audience; The nanoparticles produced by magnetotactic bacteria, called magnetosomes, are made of a magnetite core with high levels of crystallinity surrounded by a lipid bilayer. This organized structure has been developed during the course of evolution of these organisms to adapt to their specific habitat and is assumed to resist degradation and to be able to withstand the demanding biological environment. Herein, we investigated magnetosomes' structural fate upon internalization in human stem cells using magnetic and photothermal measurements, electron microscopy, and X-ray absorption spectroscopy. All measurements first converge to the demonstration that intracellular magnetosomes can experience an important biodegradation, with up to 70% of their initial content degraded, which is associated with the progressive storage of the released iron in the ferritin protein. It correlates with an extensive magnetite to ferrihydrite phase transition. The ionic species delivered by this degradation could then be used by the cells to biosynthesize magnetic nanoparticles anew. In this case, cell magnetism first decreased with magnetosomes being dissolved, but then cells remagnetized entirely, evidencing the neo-synthesis of biogenic magnetic nanoparticles. Bacteria-made biogenic magnetosomes can thus be totally remodeled by human stem cells, into human cells-made magnetic nanoparticles.

Published
2020

8. RGD-functionalized magnetosomes are efficient tumor radioenhancers for X-rays and protons

Author

Hervé Michel, David Pignol, Géraldine Adryanczyk Perrier, Joel Herault, Olivier Humbert, Juliette Thariat, Christopher T. Lefèvre, Jérôme Doyen, Béatrice Cambien, Christopher Hoog, Sandra Prévéral, Thierry Pourcher, Maha Hafsi, Transporteurs et Imagerie, Radiothérapie en Oncologie et Mécanismes biologiques des Altérations du Tissu Osseux (TIRO-MATOs - UMR E4320), UMR E4320 (TIRO-MATOs), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Côte d'Azur (UCA)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Côte d'Azur (UCA)-Service Hospitalier Frédéric Joliot (SHFJ), 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 Paris-Saclay-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 Paris-Saclay, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre de Lutte contre le Cancer Antoine Lacassagne [Nice] (UNICANCER/CAL), UNICANCER-Université Côte d'Azur (UCA), Institut de Chimie de Nice (ICN), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Métropole Nice Côte d'Azur (NCA), Centre Régional de Lutte contre le Cancer François Baclesse [Caen] (UNICANCER/CRLC), UNICANCER-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU), Service Hospitalier Frédéric Joliot (SHFJ), Université Paris-Saclay-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)-UMR E4320 (TIRO-MATOs), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Côte d'Azur (UCA)-Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Côte d'Azur (UCA), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), and Normandie Université (NU)-UNICANCER-Tumorothèque de Caen Basse-Normandie (TCBN)

Subjects
Radiation-Sensitizing Agents, media_common.quotation_subject, medicine.medical_treatment, [SDV]Life Sciences [q-bio], Magnetosome, Biomedical Engineering, Mice, Nude, Pharmaceutical Science, Medicine (miscellaneous), Bioengineering, 02 engineering and technology, X-Ray Therapy, Mice, 03 medical and health sciences, Cell Line, Tumor, Proton Therapy, medicine, Animals, Humans, General Materials Science, Magnetospirillum, Internalization, Proton therapy, 030304 developmental biology, media_common, Mice, Inbred BALB C, 0303 health sciences, Chemistry, Cancer, Neoplasms, Experimental, 021001 nanoscience & nanotechnology, medicine.disease, 3. Good health, Radiation therapy, Cancer cell, Biophysics, Molecular Medicine, Magnetic nanoparticles, Nanomedicine, Female, Magnetosomes, 0210 nano-technology, Oligopeptides
Abstract

International audience; Although chemically synthesized ferro/ferrimagnetic nanoparticles have attracted great attention in cancer theranostics, they lack radio-enhancement efficacy due to low targeting and internalization ability. Herein, we investigated the potential of RGD-tagged magnetosomes, bacterial biogenic magnetic nanoparticles naturally coated with a biological membrane and genetically engineered to express an RGD peptide, as tumor radioenhancers for conventional radiotherapy and proton therapy. Although native and RGD-magnetosomes similarly enhanced radiation-induced damage to plasmid DNA, RGD-magnetoprobes were able to boost the efficacy of radiotherapy to a much larger extent than native magnetosomes both on cancer cells and in tumors. Combined to magnetosomes@RGD, proton therapy exceeded the efficacy of X-rays at equivalent doses. Also, increased secondary emissions were measured after irradiation of magnetosomes with protons versus photons. Our results indicate the therapeutic advantage of using functionalized magnetoparticles to sensitize tumors to both X-rays and protons and strengthen the case for developing biogenic magnetoparticles for multimodal nanomedicine in cancer therapy.

Published
2020

9. Crystal structure of the transcriptional repressor DdrO: insight into the metalloprotease/repressor-controlled radiation response in Deinococcus

Author

Philippe Roche, Fabrice Confalonieri, Pascale Servant, Romaric Magerand, David Lemaire, David Pignol, François Parcy, Pascal Arnoux, Géraldine Brandelet, Raquel Martin-Arevalillo, Laurence Blanchard, Jade Doloy, Nicolas Eugénie, Arjan de Groot, Marina I. Siponen, Renaud Dumas, Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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 de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Radiorésistance des bactéries et des archées (RBA), Département Biologie des Génomes (DBG), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Régulateurs du développement de la fleur (Flo_RE), Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-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 National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Interactions Protéine Métal (IPM), Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC), ANR-19-CE12-0010,NOVOREP,Réparation des dommages ou mort cellulaire chez les bactéries exposées aux stress, Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-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)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), ANR-19-CE12-0010,NOVOREP,Réparation des dommages ou mort cellulaire chez les bactéries exposées aux stress(2019), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-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)-Institut National de la Recherche Agronomique (INRA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay, Université Paris-Saclay, 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 Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-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 National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche en Cancérologie de Marseille (CRCM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), MARAVAL, Alexandra, and Réparation des dommages ou mort cellulaire chez les bactéries exposées aux stress - - NOVOREP2019 - ANR-19-CE12-0010 - AAPG2019 - VALID

Subjects
Models, Molecular, DNA damage, [SDV]Life Sciences [q-bio], Repressor, [SDV.CAN]Life Sciences [q-bio]/Cancer, Cleavage (embryo), Crystallography, X-Ray, DNA-binding protein, Protein Structure, Secondary, 03 medical and health sciences, Protein structure, Structural Biology, Stress, Physiological, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Genetics, Deinococcus, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Peptide sequence, 030304 developmental biology, 0303 health sciences, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, biology, 030306 microbiology, Gene Expression Regulation, Bacterial, biology.organism_classification, Protein Structure, Tertiary, [SDV] Life Sciences [q-bio], Repressor Proteins, Biophysics, Metalloproteases, Alpha helix, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, DNA Damage, Transcription Factors
Abstract

Exposure to harmful conditions such as radiation and desiccation induce oxidative stress and DNA damage. In radiation-resistant Deinococcus bacteria, the radiation/desiccation response is controlled by two proteins: the XRE family transcriptional repressor DdrO and the COG2856 metalloprotease IrrE. The latter cleaves and inactivates DdrO. Here, we report the biochemical characterization and crystal structure of DdrO, which is the first structure of a XRE protein targeted by a COG2856 protein. DdrO is composed of two domains that fold independently and are separated by a flexible linker. The N-terminal domain corresponds to the DNA-binding domain. The C-terminal domain, containing three alpha helices arranged in a novel fold, is required for DdrO dimerization. Cleavage by IrrE occurs in the loop between the last two helices of DdrO and abolishes dimerization and DNA binding. The cleavage site is hidden in the DdrO dimer structure, indicating that IrrE cleaves DdrO monomers or that the interaction with IrrE induces a structural change rendering accessible the cleavage site. Predicted COG2856/XRE regulatory protein pairs are found in many bacteria, and available data suggest two different molecular mechanisms for stress-induced gene expression: COG2856 protein-mediated cleavage or inhibition of oligomerization without cleavage of the XRE repressor.

Published
2019

10. Ectosymbiotic bacteria at the origin of magnetoreception in a marine protist

Author

David Vallenet, Eric Viollier, Damien Faivre, Nathalie Leonhardt, Stéphanie Fouteau, Géraldine Adryanczyk, Corinne Cruaud, Christopher T. Lefèvre, Nicolas Menguy, Purificación López-García, David Pignol, Richard J. Weld, Karim Benzerara, Valérie Barbe, Caroline L. Monteil, Magali Floriani, Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Génomique métabolique (UMR 8030), Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-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 Paris-Saclay-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 d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Institut de recherche pour le développement [IRD] : UR206-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'écotoxicologie des radionucléides (IRSN/PSE-ENV/SRTE/LECO), Service de recherche sur les transferts et les effets des radionucléides sur les écosystèmes (IRSN/PSE-ENV/SRTE), Institut de Radioprotection et de Sûreté Nucléaire (IRSN)-Institut de Radioprotection et de Sûreté Nucléaire (IRSN), Institut de Physique du Globe de Paris (IPGP), Institut national des sciences de l'Univers (INSU - CNRS)-Université Paris Diderot - Paris 7 (UPD7)-Université de La Réunion (UR)-Institut de Physique du Globe de Paris (IPG Paris)-Centre National de la Recherche Scientifique (CNRS), Plant Environmental Physiology and Stress Signaling (PEPSS), Department of Biomaterials [Potsdam], Max Planck Institute of Colloids and Interfaces, Max-Planck-Gesellschaft-Max-Planck-Gesellschaft, Ecologie Systématique et Evolution (ESE), Université Paris-Sud - Paris 11 (UP11)-AgroParisTech-Centre National de la Recherche Scientifique (CNRS), Lincoln University, New Zealand, Dumont d’Urville Science and Technology Programme (grant no. DDU-LVL1501), New Zealand Ministry for Business, Innovation and Employment (grant no. LVLX1703), ANR-16-TERC-0025,BIOMAGNET,Exploration de la biodiversité des bactéries magnétotactiques(2016), ANR-14-CE35-0018,GroMa,Greigite or magnetite: Environmental and genetic determinants controlling biomineralization in magnetotactic bacteria(2014), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), 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 d'Évry-Val-d'Essonne (UEVE), PSE-ENV/SRTE/LECO, Institut de Radioprotection et de Sûreté Nucléaire (IRSN), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut national des sciences de l'Univers (INSU - CNRS)-IPG PARIS-Université Paris Diderot - Paris 7 (UPD7)-Université de La Réunion (UR)-Centre National de la Recherche Scientifique (CNRS), Signalisation de l'Adaptation des Végétaux à l'Environnement (SAVE), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-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)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), and Laboratoire de Radioécologie et d'Ecotoxicologie (DEI/SECRE/LRE)

Subjects
Microbiology (medical), [SDV]Life Sciences [q-bio], Immunology, Magnetosome, Biology, medicine.disease_cause, Deltaproteobacteria, Applied Microbiology and Biotechnology, Microbiology, 03 medical and health sciences, Symbiosis, Genetics, medicine, Euglenozoa, 14. Life underwater, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, Mutualism (biology), 0303 health sciences, 030306 microbiology, Protist, Magnetoreception, Cell Biology, biology.organism_classification, Anoxic waters, Evolutionary biology
Abstract

Mutualistic symbioses are often a source of evolutionary innovation and drivers of biological diversification1. Widely distributed in the microbial world, particularly in anoxic settings2,3, they often rely on metabolic exchanges and syntrophy2,4. Here, we report a mutualistic symbiosis observed in marine anoxic sediments between excavate protists (Symbiontida, Euglenozoa)5 and ectosymbiotic Deltaproteobacteria biomineralizing ferrimagnetic nanoparticles. Light and electron microscopy observations as well as genomic data support a multi-layered mutualism based on collective magnetotactic motility with division of labour and interspecies hydrogen-transfer-based syntrophy6. The guided motility of the consortia along the geomagnetic field is allowed by the magnetic moment of the non-motile ectosymbiotic bacteria combined with the protist motor activity, which is a unique example of eukaryotic magnetoreception7 acquired by symbiosis. The nearly complete deltaproteobacterial genome assembled from a single consortium contains a full magnetosome gene set8, but shows signs of reduction, with the probable loss of flagellar genes. Based on the metabolic gene content, the ectosymbiotic bacteria are anaerobic sulfate-reducing chemolithoautotrophs that likely reduce sulfate with hydrogen produced by hydrogenosome-like organelles6 underlying the plasma membrane of the protist. In addition to being necessary hydrogen sinks, ectosymbionts may provide organics to the protist by diffusion and predation, as shown by magnetosome-containing digestive vacuoles. Phylogenetic analyses of 16S and 18S ribosomal RNA genes from magnetotactic consortia in marine sediments across the Northern and Southern hemispheres indicate a host–ectosymbiont specificity and co-evolution. This suggests a historical acquisition of magnetoreception by a euglenozoan ancestor from Deltaproteobacteria followed by subsequent diversification. It also supports the cosmopolitan nature of this type of symbiosis in marine anoxic sediments. Here, the authors identify a mutualistic symbiosis between a non-motile, magnetic deltaproteobacterium and a protist, resulting in eukaryotic magnetoreception in marine anoxic sediments.

Published
2019

11. Tuning the redox properties of a [4Fe-4S] center to modulate the activity of Mo-bisPGD periplasmic nitrate reductase

Author

Bruno Guigliarelli, David Pignol, Kamal Zeamari, Guillaume Gerbaud, Florence Chaspoul, Pascal Arnoux, Bénédicte Burlat, Monique Sabaty, Sandrine Grosse, Vincent Fourmond, Frédéric Biaso, Bioénergétique et Ingénierie des Protéines (BIP ), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Institut méditerranéen de biodiversité et d'écologie marine et continentale (IMBE), Avignon Université (AU)-Aix Marseille Université (AMU)-Institut de recherche pour le développement [IRD] : UMR237-Centre National de la Recherche Scientifique (CNRS), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), and ANR-16-CE29-0010,MOLYERE,Comprendre et Contrôler la Réactivité du Cofacteur à Molybdène dans les Enzymes et les Protéines Maquettes(2016)

Subjects
Iron-Sulfur Proteins, 0301 basic medicine, Stereochemistry, [SDV]Life Sciences [q-bio], Mutation, Missense, Biophysics, Rhodobacter sphaeroides, 010402 general chemistry, Nitrate reductase, Nitrate Reductase, 01 natural sciences, Biochemistry, Redox, Electron Transport, 03 medical and health sciences, Electron transfer, Catalytic Domain, Redox titration, [CHIM]Chemical Sciences, ComputingMilieux_MISCELLANEOUS, biology, Chemistry, Active site, Cell Biology, Periplasmic space, biology.organism_classification, Electron transport chain, 0104 chemical sciences, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, 030104 developmental biology, Amino Acid Substitution, biology.protein, Periplasmic Proteins, Oxidation-Reduction
Abstract

Molybdoenzymes are ubiquitous in living organisms and catalyze, for most of them, oxidation-reduction reactions using a large range of substrates. Periplasmic nitrate reductase (NapAB) from Rhodobacter sphaeroides catalyzes the 2-electron reduction of nitrate into nitrite. Its active site is a Mo bis-(pyranopterin guanine dinucleotide), or Mo-bisPGD, found in most prokaryotic molybdoenzymes. A [4Fe-4S] cluster and two c-type hemes form an intramolecular electron transfer chain that deliver electrons to the active site. Lysine 56 is a highly conserved amino acid which connects, through hydrogen-bonds, the [4Fe-4S] center to one of the pyranopterin ligands of the Mo-cofactor. This residue was proposed to be involved in the intramolecular electron transfer, either defining an electron transfer pathway between the two redox cofactors, and/or modulating their redox properties. In this work, we investigated the role of this lysine by combining site-directed mutagenesis, activity assays, redox titrations, EPR and HYSCORE spectroscopies. Removal of a positively-charged residue at position 56 strongly decreased the redox potential of the [4Fe-4S] cluster at pH 8 by 230 mV to 400 mV in the K56H and K56M mutants, respectively, thus affecting the kinetics of electron transfer from the hemes to the [4Fe-4S] center up to 5 orders of magnitude. This effect was partly reversed at acidic pH in the K56H mutant likely due to protonation of the imidazole ring of the histidine. Overall, our study demonstrates the critical role of a charged residue from the second coordination sphere in tuning the reduction potential of the [4Fe-4S] cluster in RsNapAB and related molybdoenzymes.

Published
2019

12. Magnetosomes: biogenic iron nanoparticles produced by environmental bacteria

Author

Sandra Prévéral, Anissa Dieudonné, David Pignol, Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)

Subjects
0303 health sciences, Magnetotactic bacteria, biology, Bacteria, 030306 microbiology, Chemistry, Iron, [SDV]Life Sciences [q-bio], Magnetosome, Nanotechnology, General Medicine, biology.organism_classification, Applied Microbiology and Biotechnology, Cancer treatment, 03 medical and health sciences, Environmental Microbiology, Nanoparticles, Magnetosomes, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, Biotechnology, Biomineralization
Abstract

The scientific community's interest in magnetotactic bacteria has increased substantially in recent decades. These prokaryotes have the particularity of synthesizing nanomagnets, called magnetosomes. The majority of research is based on several scientific questions. Where do magnetotactic bacteria live, what are their characteristics, and why are they magnetic? What are the molecular phenomena of magnetosome biomineralization and what are the physical characteristics of magnetosomes? In addition to scientific curiosity to better understand these stunning organisms, there are biotechnological opportunities to consider. Magnetotactic bacteria, as well as magnetosomes, are used in medical applications, for example cancer treatment, or in environmental ones, for example bioremediation. In this mini-review, we investigated all the aspects mentioned above and summarized the currently available knowledge.

Published
2019

13. Control by Metals of Staphylopine Dehydrogenase Activity during Metallophore Biosynthesis

Author

Christine Hajjar, Roberto Fanelli, Clémentine Laffont, Catherine Brutesco, Gregorio Cullia, Mathilde Tribout, Didier Nurizzo, Elise Borezée-Durant, Romé Voulhoux, David Pignol, Jérôme Lavergne, Florine Cavelier, Pascal Arnoux, Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), European Synchrotron Radiation Facility (ESRF), MICrobiologie de l'ALImentation au Service de la Santé (MICALIS), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Association Vaincre la Mucoviscidose VLM, financement RFI20160501495, ANR-14-CE09-0007,ANIBAL,Un nouveau metallophore dérivé de la nicotianamine chez des bactéries pathogènes(2014), 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), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-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)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA))

Subjects
Models, Molecular, 0303 health sciences, Staphylococcus aureus, pseudomonas-aeruginosa, 030306 microbiology, [SDV]Life Sciences [q-bio], Imidazoles, Molecular Conformation, crystal-structure, General Chemistry, nutritional immunity, Biochemistry, Catalysis, 03 medical and health sciences, Colloid and Surface Chemistry, iron, Metals, Heavy, transport, escherichia-coli, ions, Oxidoreductases, roles, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology
Abstract

International audience; Enzymatic regulations are central processes for the adaptation to changing environments. In the particular case of metallophore-dependent metal uptake, there is a need to quickly adjust the production of these metallophores to the metal level outside the cell, to avoid metal shortage or overload, as well as waste of metallophores. In Staphylococcus aureus, CntM catalyzes the last biosynthetic step in the production of staphylopine, a broad-spectrum metallophore, through the reductive condensation of a pathway intermediate (xNA) with pyruvate. Here, we describe the chemical synthesis of this intermediate, which was instrumental in the structural and functional characterization of CntM and confirmed its opine synthase properties. The three-dimensional structure of CntM was obtained in an "open" form, in the apo state or as a complex with substrate or product. The xNA substrate appears mainly stabilized by its imidazole ring through a pi-pi interaction with the side chain of Tyr240. Intriguingly, we found that metals exerted various and sometime antagonistic effects on the reaction catalyzed by CntM: zinc and copper are moderate activators at low concentration and then total inhibitors at higher concentration, whereas manganese is only an activator and cobalt and nickel are only inhibitors. We propose a model in which the relative affinity of a metal toward xNA and an inhibitory binding site on the enzyme controls activation, inhibition, or both as a function of metal concentration. This metal-dependent regulation of a metallophore-producing enzyme might also take place in vivo, which could contribute to the adjustment of metallophore production to the internal metal level.

Published
2019

14. Ectosymbiotic bacteria at the origin of magnetoreception in a marine protist

Author

Caroline L, Monteil, David, Vallenet, Nicolas, Menguy, Karim, Benzerara, Valérie, Barbe, Stéphanie, Fouteau, Corinne, Cruaud, Magali, Floriani, Eric, Viollier, Géraldine, Adryanczyk, Nathalie, Leonhardt, Damien, Faivre, David, Pignol, Purificación, López-García, Richard J, Weld, and Christopher T, Lefevre

Subjects
Deltaproteobacteria, Geologic Sediments, Oceans and Seas, Euglenozoa, Eukaryota, Ferrosoferric Oxide, Biological Coevolution, Magnetic Fields, Species Specificity, RNA, Ribosomal, Anaerobiosis, Magnetosomes, Symbiosis, Genome, Bacterial, Locomotion, Phylogeny, Hydrogen
Abstract

Mutualistic symbioses are often a source of evolutionary innovation and drivers of biological diversification

Published
2018

15. Controlled Biomineralization of Magnetite in Bacteria

Author

Elodie C. T. Descamps, Christopher T. Lefèvre, Jean-Baptiste Abbe, and David Pignol

Subjects
0301 basic medicine, biology, Magnetotactic bacteria, 030106 microbiology, Magnetosome, biology.organism_classification, 03 medical and health sciences, chemistry.chemical_compound, 030104 developmental biology, chemistry, Biochemistry, Organelle, Magnetotaxis, Bacteria, Magnetite, Biomineralization
Published
2016

16. $Rhodobacter\ sphaeroides$ methionine sulfoxide reductase P reduces $R$ ‑ and $S$ ‑diastereomers of methionine sulfoxide from a broad‑spectrum of protein substrates

Author

Sandrine Grosse, Guylaine Miotello, Pascal Arnoux, David Lemaire, Lionel Tarrago, Marina I. Siponen, Jean Armengaud, David Pignol, Béatrice Alonso, Monique Sabaty, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Interactions Protéine Métal (IPM), Laboratoire Innovations technologiques pour la Détection et le Diagnostic (LI2D), Service de Pharmacologie et Immunoanalyse (SPI), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-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 National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-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 National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Médicaments et Technologies pour la Santé (MTS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Commissariat a l'Energie Atomique et aux Energies Alternatives (CEA) [ANR 16-CE11-0012], ANR-16-CE11-0012,METOXIC,Enzyme de réparation des protéines oxydées: un nouveau système de contrôle qualité dans le périplasme(2016), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, proteomics 15, [SDV]Life Sciences [q-bio], Protein oxidation, Biochemistry, 03 medical and health sciences, chemistry.chemical_compound, Rhodobacter sphaeroides, protein oxidation, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology, periplasm, Methionine, biology, Methionine sulfoxide, microbiology, Enzyme kinetics, molybdoenzyme, Cell Biology, Periplasmic space, biology.organism_classification, oxidative 14 stress, 030104 developmental biology, chemistry, methionine sulfoxide reductase, Methionine sulfoxide reductase, Photosynthetic bacteria, MSRA
Abstract

International audience; Methionine (Met) is prone to oxidation and can be converted to Met sulfoxide (MetO), which exists as $R$- and $S$-diastereomers. MetO can be reduced back to Met by the ubiquitous methionine sulfoxide reductase (Msr) enzymes. Canonical MsrA and MsrB were shown to be absolutely stereospecific for the reduction of S ‑ and R‑ diastereomer, respectively. Recently, a new enzymatic system, MsrQ/MsrP which is conserved in all gram‑negative bacteria, was identified as a key actor in the reduction of oxidized periplasmic proteins. The haem‑binding membrane protein MsrQ transmits reducing power from the electron transport chains to the molybdoenzyme MsrP, which acts as a protein‑MetO reductase. The MsrQ/MsrP function was well established genetically, but the identity and biochemical properties of MsrP substrates remain unknown. In this work, using the purified MsrP enzyme from the photosynthetic bacteria $Rhodobacter\ sphaeroides$ as a model, we show that it can reduce a broad spectrum of protein substrates. The most efficiently reduced MetO are found in clusters of amino acid sequences devoid of threonine and proline on the C‑terminal side. Moreover, $R.\ sphaeroides$ MsrP lacks stereospecificity as it can reduce both R‑ and S‑ diastereomers of MetO, similarly to its $Escherichia\ coli$ homolog, and preferentially acts on unfolded oxidized proteins. Overall, these results provide important insights into the function of a bacterial envelop protecting system, which should help understand how bacteria cope in harmful environments.

Published
2018

17. Genomic study of a novel magnetotactic Alphaproteobacteria uncovers the multiple ancestry of magnetotaxis

Author

Béatrice Alonso, Christopher T. Lefèvre, Nicolas Menguy, Caroline L. Monteil, Nicolas Ginet, Stéphane Cruveiller, Guy Perrière, David Pignol, Nicolas Waisbord, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Bioinformatique, phylogénie et génomique évolutive (BPGE), Département PEGASE [LBBE] (PEGASE), Laboratoire de Biométrie et Biologie Evolutive - UMR 5558 (LBBE), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-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 Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-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)-Laboratoire de Biométrie et Biologie Evolutive - UMR 5558 (LBBE), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-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), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Institut de recherche pour le développement [IRD] : UR206-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Tufts University [Medford], Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Génomique métabolique (UMR 8030), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-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 d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), ANR-14-CE35-0018,GroMa,Greigite or magnetite: Environmental and genetic determinants controlling biomineralization in magnetotactic bacteria(2014), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), 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)-Université d'Évry-Val-d'Essonne (UEVE)

Subjects
0301 basic medicine, Gene Transfer, Horizontal, Magnetotactic bacteria, [SDV]Life Sciences [q-bio], 030106 microbiology, Magnetosome, Microbiology, Evolution, Molecular, Magnetics, 03 medical and health sciences, Mediterranean Sea, Magnetotaxis, 14. Life underwater, Magnetospirillum, Ecology, Evolution, Behavior and Systematics, Alphaproteobacteria, Comparative genomics, biology, Phylum, biology.organism_classification, Evolutionary biology, France, Magnetosomes, Proteobacteria, Water Microbiology, Genome, Bacterial
Abstract

Ecological and evolutionary processes involved in magnetotactic bacteria (MTB) adaptation to their environment have been a matter of debate for many years. Ongoing efforts for their characterization are progressively contributing to understand these processes, including the genetic and molecular mechanisms responsible for biomineralization. Despite numerous culture-independent MTB characterizations, essentially within the Proteobacteria phylum, only few species have been isolated in culture because of their complex growth conditions. Here, we report a newly cultivated magnetotactic, microaerophilic and chemoorganoheterotrophic bacterium isolated from the Mediterranean Sea in Marseille, France: Candidatus Terasakiella magnetica strain PR-1 that belongs to an Alphaproteobacteria genus with no magnetotactic relative. By comparing the morphology and the whole genome shotgun sequence of this MTB with those of closer relatives, we brought further evidence that the apparent vertical ancestry of magnetosome genes suggested by previous studies within Alphaproteobacteria hides a more complex evolutionary history involving horizontal gene transfers and/or duplication events before and after the emergence of Magnetospirillum, Magnetovibrio and Magnetospira genera. A genome-scale comparative genomics analysis identified several additional candidate functions and genes that could be specifically associated to MTB lifestyle in this class of bacteria.

Published
2018

18. Targeted thermal therapy with genetically engineered magnetite magnetosomes@RGD: Photothermia is far more efficient than magnetic hyperthermia

Author

Anouchka Plan Sangnier, Yoann Lalatonne, Sandra Prévéral, Chistopher T. Lefèvre, David Pignol, Alberto Curcio, Claire Wilhelm, Amanda K. A. Silva, CNRS UMR 7057 - Laboratoire Matières et Systèmes Complexes (MSC) (MSC), Centre National de la Recherche Scientifique (CNRS), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-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), Microbiologie Environnementale et Moléculaire (MEM), 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)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Matière et Systèmes Complexes (MSC (UMR_7057)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Recherche Vasculaire Translationnelle (LVTS (UMR_S_1148 / U1148)), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Matière et Systèmes Complexes (MSC), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7)

Subjects
Male, Hot Temperature, Magnetotactic bacteria, [SDV]Life Sciences [q-bio], Magnetosome, Pharmaceutical Science, Mice, Nude, Uterine Cervical Neoplasms, Antineoplastic Agents, 02 engineering and technology, 010402 general chemistry, 01 natural sciences, HeLa, Mice, Animals, Humans, Magnetite Nanoparticles, ComputingMilieux_MISCELLANEOUS, biology, Chemistry, Prostatic Neoplasms, 021001 nanoscience & nanotechnology, biology.organism_classification, 0104 chemical sciences, 3. Good health, Magnetic hyperthermia, Tumor progression, Heat generation, Cancer cell, PC-3 Cells, Biophysics, Magnetic nanoparticles, Female, Magnetosomes, 0210 nano-technology, Genetic Engineering, Oligopeptides, HeLa Cells
Abstract

Providing appropriate means for heat generation by low intratumoral nanoparticle concentrations is a major challenge for cancer nanotherapy. Here we propose RGD-tagged magnetosomes (magnetosomes@RGD) as a biogenic, genetically engineered, inorganic platform for multivalent thermal cancer treatment. Magnetosomes@RGD are biomagnetite nanoparticles synthesized by genetically modified magnetotactic bacteria thanks to a translational fusion of the RGD peptide with the magnetosomal protein MamC. Magnetosomes@RGD thus combine the high crystallinity of their magnetite core with efficient surface functionalization. The specific affinity of RGD was first quantified by single-cell magnetophoresis with a variety of cell types, including immune, muscle, endothelial, stem and cancer cells. The highest affinity and cellular uptake was observed with PC3 prostatic and HeLa uterine cancer cells. The efficiency of photothermia and magnetic hyperthermia was then compared on PC3 cells. Unexpectedly, photothermia was far more efficient than magnetic hyperthermia, which was almost totally inhibited by the cellular environment. RGD targeting was then assessed in vivo at tumor site, in mice bearing PC3 tumors. As a result, we demonstrate that targeted magnetic nanoparticles could generate heat on a therapeutic level after systemic administration, but only under laser excitation, and successfully inhibit tumor progression.

Published
2018

19. Accumulation and Dissolution of Magnetite Crystals in a Magnetically Responsive Ciliate

Author

David Pignol, Caroline L. Monteil, Christopher T. Lefèvre, Nicolas Menguy, Sandra Prévéral, Alan Warren, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Institut de recherche pour le développement [IRD] : UR206-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), National History Museum of London, ANR-14-CE35-0018,GroMa,Greigite or magnetite: Environmental and genetic determinants controlling biomineralization in magnetotactic bacteria(2014), ANR-16-TERC-0025,BIOMAGNET,Exploration de la biodiversité des bactéries magnétotactiques(2016), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, iron cycling, Food Chain, magnetite, Magnetotactic bacteria, Microorganism, [SDV]Life Sciences [q-bio], Magnetosome, Applied Microbiology and Biotechnology, 03 medical and health sciences, Environmental Microbiology, grazing, 14. Life underwater, Ciliate, Ecology, biology, Bacteria, Chemistry, Aquatic ecosystem, magnetotactic, 15. Life on land, equipment and supplies, biology.organism_classification, biomineralization, Ferrosoferric Oxide, 030104 developmental biology, Solubility, 13. Climate action, Environmental chemistry, Oligohymenophorea, Protozoa, France, Magnetosomes, Food Science, Biotechnology, Biomineralization
Abstract

Magnetotactic bacteria (MTB) represent a group of microorganisms that are widespread in aquatic habitats and thrive at oxic-anoxic interfaces. They are able to scavenge high concentrations of iron thanks to the biomineralization of magnetic crystals in their unique organelles, the so-called magnetosome chains. Although their biodiversity has been intensively studied, their ecology and impact on iron cycling remain largely unexplored. Predation by protozoa was suggested as one of the ecological processes that could be involved in the release of iron back into the ecosystem. Magnetic protozoa were previously observed in aquatic environments, but their diversity and the fate of particulate iron during grazing are poorly documented. In this study, we report the morphological and molecular characterizations of a magnetically responsive MTB-grazing protozoan able to ingest high quantities of MTB. This protozoan is tentatively identified as Uronema marinum , a ciliate known to be a predator of bacteria. Using light and electron microscopy, we investigated in detail the vacuoles in which the lysis of phagocytized prokaryotes occurs. We carried out high-resolution observations of aligned magnetosome chains and ongoing dissolution of crystals. Particulate iron in the ciliate represented approximately 0.01% of its total volume. We show the ubiquity of this interaction in other types of environments and describe different grazing strategies. These data contribute to the mounting evidence that the interactions between MTB and protozoa might play a significant role in iron turnover in microaerophilic habitats. IMPORTANCE Identifying participants of each biogeochemical cycle is a prerequisite to our understanding of ecosystem functioning. Magnetotactic bacteria (MTB) participate in iron cycling by concentrating large amounts of biomineralized iron minerals in their cells, which impacts their chemical environment at, or below, the oxic-anoxic transition zone in aquatic habitats. It was shown that some protozoa inhabiting this niche could become magnetic by the ingestion of magnetic crystals biomineralized by grazed MTB. In this study, we show that magnetic MTB grazers are commonly observed in marine and freshwater sediments and can sometimes accumulate very large amounts of particulate iron. We describe here different phagocytosis strategies, determined using magnetic particles from MTB as tracers after their ingestion by the protozoa. This study paves the way for potential scientific or medical applications using MTB grazers as magnetosome hyperaccumulators.

Published
2018

20. The Rhodobacter sphaeroides methionine sulfoxide reductase MsrP can reduce R- and S-diastereomers of methionine sulfoxide from a broad-spectrum of protein substrates

Author

David Lemaire, Monique Sabaty, Jean Armengaud, David Pignol, Lionel Tarrago, Pascal Arnoux, Sandrine Grosse, Guylaine Miotello, Marina I. Siponen, and Béatrice Alonso

Subjects
Methionine, biology, Chemistry, Methionine sulfoxide, Sulfoxide, Periplasmic space, medicine.disease_cause, biology.organism_classification, chemistry.chemical_compound, Rhodobacter sphaeroides, Biochemistry, medicine, Methionine sulfoxide reductase, Escherichia coli, MSRA
Abstract

SummaryMethionine (Met) is prone to oxidation and can be converted to Met sulfoxide (MetO), which exists as R- and S-diastereomers. MetO can be reduced back to Met by the ubiquitous methionine sulfoxide reductase (Msr) enzymes. Canonical MsrA and MsrB were shown as absolutely stereospecific for the reduction of S- and R-diastereomer, respectively. Recently, the molybdenum-containing protein MsrP, conserved in all gram-negative bacteria, was shown to be able to reduce MetO of periplasmic proteins without apparent stereospecificity inEscherichia coli.Here, we describe the substrate specificity of theRhodobacter sphaeroidesMsrP. Proteomics analysis coupled to enzymology approaches indicate that it reduces a broad spectrum of periplasmic oxidized proteins. Moreover, using model proteins, we demonstrated that RsMsrP preferentially reduces unfolded oxidized proteins and we confirmed that this enzyme, like itsE. colihomolog, can reduce bothR-andS-diastereomers of MetO with similar efficiency.

Published
2018
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21. Pushing the limits of nickel detection to nanomolar range using a set of engineered bioluminescent Escherichia coli

Author

Camille Escoffier, Agnès Rodrigue, Marie-Andrée Mandrand-Berthelot, Daniel Garcia, David Pignol, Julien Cayron, Erwan Gueguen, Elsa Prudent, Microbiologie, adaptation et pathogénie (MAP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Trafic et signalisation membranaires chez les bactéries (MTSB), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Microbiologie Environnementale et Moléculaire (MEM), École des Mines de Saint-Étienne (Mines Saint-Étienne MSE), Institut Mines-Télécom [Paris] (IMT), ANR-11-ECOT-0009,COMBITOX,COnception d'un instrument pour la Mesure BIologique multiparamètrique en continu de TOXiques(2011), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), and Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL)

Subjects
0301 basic medicine, Health, Toxicology and Mutagenesis, chemistry.chemical_element, Biosensing Techniques, macromolecular substances, Biology, medicine.disease_cause, Sensitivity and Specificity, 03 medical and health sciences, Genes, Reporter, Nickel, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Escherichia coli, medicine, Environmental Chemistry, Bioluminescence, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Promoter Regions, Genetic, Detection limit, Reporter gene, Escherichia coli Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Gene Expression Regulation, Bacterial, General Medicine, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Pollution, Molecular biology, Repressor Proteins, Luminescent Proteins, 030104 developmental biology, Biochemistry, chemistry, Regulatory circuit, Efflux, Biosensor
Abstract

International audience; The detection of nickel in water is of great importance due to its harmfulness for living organism. A way to detect Ni is the use of whole-cell biosensors. The aim of the present work was to build a light-emitting bacterial biosensor for the detection of Ni with high specificity and low detection limit properties. For that purpose, the regulatory circuit implemented relied on the RcnR Ni/Co metallo-regulator and its rcnA natural target promoter fused to the lux reporter genes. To convert RcnR to specifically detect Ni, several mutations were tested and the C35A retained. Deleting the Ni efflux pump rcnA and introducing genes encoding several Ni-uptake systems lowered the detection thresholds. When these constructs were assayed in several Escherichia coli strains, it appeared that the detection thresholds were highly variable. The TD2158 wild-type E. coli gave rise to a biosensor ten times more active and sensitive than its W3110 E. coli K12 equivalent. This biosensor was able to confidently detect Ni concentrations as little as 80 nM (4.7 μg l −1), which makes its use compatible with the norms governing the drinking water quality.

Published
2015

22. An algal photoenzyme converts fatty acids to hydrocarbons

Author

Bertrand Légeret, Pavel Müller, Yonghua Li-Beisson, Yohann Couté, David Pignol, Fred Beisson, Gilles Peltier, Damien Sorigué, Emmanuelle Billon, Stéphanie Blangy, Didier Nurizzo, Pascal Arnoux, Pierre Richaud, Solène L. Y. Moulin, Klaus Brettel, Stéphan Cuiné, Sabine Brugière, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Synthèse et étude de systèmes à intêret biologique (SEESIB), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Centre National de la Recherche Scientifique (CNRS), Troubles cognitifs dégénératifs et vasculaires (DN2M), Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-INSERM, Bioénergie et Microalgues (EBM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Etude de la dynamique des protéomes (EDyP), Laboratoire de Biologie à Grande Échelle (BGE - UMR S1038), 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 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), European Synchrotron Radiation Facility (ESRF), Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay, Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), 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), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), ANR-13-JSV5-0005,MUsCA,Ingénierie métabolique d'une microalgue verte en vue de la production d'alcanes à chaine moyenne(2013), ANR-11-IDEX-0001,Amidex,INITIATIVE D'EXCELLENCE AIX MARSEILLE UNIVERSITE(2011), ANR-10-INBS-0005,FRISBI,Infrastructure Française pour la Biologie Structurale Intégrée(2010), ANR-10-INBS-0008,ProFI,Infrastructure Française de Protéomique(2010), Biologie végétale et microbiologie environnementale - UMR7265 ( BVME ), Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Aix Marseille Université ( AMU ), Laboratoire de Biologie à Grande Échelle ( BGE - UMR S1038 ), Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Grenoble Alpes [Saint Martin d'Hères]-Institut National de la Santé et de la Recherche Médicale ( INSERM ), European Synchrotron Radiation Facility ( ESRF ), Biologie et Biotechnologie des Cyanobactéries ( B2CYA ), Département Microbiologie ( Dpt Microbio ), Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ), Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ), and Environnement, Bioénergie, Microalgues et Plantes (EBMP)

Subjects
0301 basic medicine, Photosynthetic reaction centre, Light, Carboxy-Lyases, Decarboxylation, [SDV]Life Sciences [q-bio], Chlorella, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Alkenes, 010402 general chemistry, 01 natural sciences, Cofactor, 03 medical and health sciences, chemistry.chemical_compound, Oxidoreductase, Alkanes, Phylogeny, ComputingMilieux_MISCELLANEOUS, Plant Proteins, chemistry.chemical_classification, Flavin adenine dinucleotide, Multidisciplinary, biology, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, Fatty Acids, Fatty acid, Lipid metabolism, Lipid Metabolism, Photochemical Processes, 0104 chemical sciences, 030104 developmental biology, Enzyme, chemistry, Biochemistry, Biocatalysis, Flavin-Adenine Dinucleotide, biology.protein, Oxidoreductases
Abstract

Algal enzyme driven by blue light Microalgae make hydrocarbons. In searching for the enzyme responsible, Sorigué et al. found a glucose-methanolcholine oxidoreductase (see the Perspective by Scrutton). Expression of the enzyme in Escherichia coli showed that hydrocarbon production requires visible light. In fact, the enzyme requires a constant input of blue photons to carry out its catalytic reaction. A long hydrophobic tunnel in the enzyme stabilizes the fatty acid substrates in proximity to the flavin adenine dinucleotide cofactor. Science , this issue p. 903 ; see also p. 872

Published
2017

23. Pseudopaline, a staphylopine-like metallophore involved in zinc and nickel uptake in Pseudomonas aeruginosa

Author

Ryszard Lobinski, Ahmed Liratni, Ghassan Ghssein, Nicolas Gomez, Pierre Richaud, Shuanglong Wang, Elise Borezée-Durant, Geneviève Ball, Catherine Brutesco, Christine Hajjar, Romé Voulhoux, Sophie Bleves, David Pignol, Laurent Ouerdane, Pascal Arnoux, and Sébastien Lhospice

Subjects
0303 health sciences, 030306 microbiology, Operon, Repressor, chemistry.chemical_element, Zinc, Biology, 03 medical and health sciences, chemistry.chemical_compound, Alpha ketoglutarate, chemistry, Biosynthesis, Biochemistry, Moiety, Nicotianamine, Histidine, 030304 developmental biology
Abstract

Metal uptake is vital for all living organisms. In metal scarce conditions, a common bacterial strategy consists in the biosynthesis of metallophores, their export in the extracellular medium and the recovery of a metal-metallophore complex through dedicated membrane transporters. Staphylopine is a recently described metallophore distantly related to plant nicotianamine that contributes to the broad-spectrum metal uptake capabilities of Staphylococcus aureus. Here, we characterize a four genes operon (PA4837–PA4834) in Pseudomonas aeruginosa involved in the biosynthesis and trafficking of a staphylopine-like metallophore named pseudopaline. Pseudopaline differs from staphylopine with regard to the stereochemistry of its histidine moiety associated to an alpha ketoglutarate moiety instead of pyruvate. In vivo, the pseudopaline operon is regulated by zinc through the Zur repressor. The metal-uptake property of the pseudopaline system appears different from that of staphylopine with a predominant effect on nickel uptake, and on zinc uptake in metal scarce conditions mimicking a chelating environment, thus reconciling the regulation of the cnt operon by zinc with its function as a zinc importer under metal scarce conditions.AUTHOR SUMMARYZinc is an essential micronutrients for bacteria, particularly important at the host-pathogen interface where the host tends to sequester metals in a so called nutritional immunity framework, and the pathogenic bacterium increases its metal uptake efforts in order to keep up with its metal requirements. Here we reveal a novel metallophore, named pseudopaline, which is synthesized and exported by Pseudomonas aeruginosa and serves for the uptake of nickel in metal poor media, and for the uptake of zinc in metal scarce conditions that mimic the chelating environment that presumably prevails within a host.

Published
2017
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24. Desulfamplus magnetovallimortis gen. nov., sp nov., a magnetotactic bacterium from a brackish desert spring able to biomineralize greigite and magnetite, that represents a novel lineage in the Desulfobacteraceae

Author

Caroline L. Monteil, Christopher T. Lefèvre, David Pignol, Nicolas Menguy, Nicolas Ginet, Elodie C. T. Descamps, Dennis A. Bazylinski, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Institut de recherche pour le développement [IRD] : UR206-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), University of Nevada [Reno], U.S. National Science Foundation grant EAR-1423939, ANR-14-CE35-0018,GroMa,Greigite or magnetite: Environmental and genetic determinants controlling biomineralization in magnetotactic bacteria(2014), ANR-12-NANO-0013,MEFISTO,Nano-aimants biologiques fonctionnalisés pour la théranostique(2012), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Greigite, DNA, Bacterial, Deltaproteobacteria, Magnetotactic bacteria, Iron, Magnetosome, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Sulfides, Applied Microbiology and Biotechnology, Microbiology, California, 03 medical and health sciences, Fumarates, Botany, Magnetotaxis, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Ecology, Evolution, Behavior and Systematics, Base Composition, [SDV.GEN]Life Sciences [q-bio]/Genetics, biology, Strain (chemistry), Obligate anaerobe, Sequence Analysis, DNA, biology.organism_classification, Ferrosoferric Oxide, Bacterial Typing Techniques, 030104 developmental biology, [SDE]Environmental Sciences, Desulfobacteraceae, Magnetosomes, Genome, Bacterial
Abstract

International audience; A magnetotactic bacterium, designated strain BW-1(T), was isolated from a brackish spring in Death Valley National Park (California, USA) and cultivated in axenic culture. The Gram-negative cells of strain BW-1(T) are relatively large and rod-shaped and possess a single polar flagellum (monotrichous). This strain is the first magnetotactic bacterium isolated in axenic culture capable of producing greigite and/or magnetite nanocrystals aligned in one or more chains per cell. Strain BW-1(T) is an obligate anaerobe that grows chemoorganoheterotrophically while reducing sulfate as a terminal electron acceptor. Optimal growth occurred at pH 7.0 and 28 degrees C with fumarate as electron donor and carbon source. Based on its genome sequence, the G + C content is 40.72 mol %. Phylogenomic and phylogenetic analyses indicate that strain BW-1(T) belongs to the Desulfobacteraceae family within the Deltaproteobacteria class. Based on average amino acid identity, strain BW-1(T) can be considered as a novel species of a new genus, for which the name Desulfamplus magnetovallimortis is proposed. The type strain of D. magnetovallimortis is BW-1(T) (JCM 18010(T)-DSM 103535(T)). (C) 2017 Elsevier GmbH. All rights reserved.

Published
2017

25. Genetically tailored magnetosomes used as MRI probe for molecular imaging of brain tumor

Author

Sandra Prévéral, Erwan Selingue, Sébastien Mériaux, Marianne Boucher, David Pignol, Christopher T. Lefèvre, Michel Pean, Géraldine Adryanczyk-Perrier, Françoise Geffroy, Nicolas Ginet, L. Bellanger, HYBIS, Laboratoire d'étude des transferts en hydrologie et environnement (LTHE), Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique de Grenoble (INPG)-Institut national des sciences de l'Univers (INSU - CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-Observatoire des Sciences de l'Univers de Grenoble (OSUG), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut national des sciences de l'Univers (INSU - CNRS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique de Grenoble (INPG)-Institut national des sciences de l'Univers (INSU - CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-Observatoire des Sciences de l'Univers de Grenoble (OSUG), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut national des sciences de l'Univers (INSU - CNRS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Department of Applied Geophysics, Technische Universität Berlin (TUB)-Technische Universität Berlin (TUB), Unité d'imagerie par résonance magnétique à très haut champ et de spectroscopie (UNIRS), Service NEUROSPIN (NEUROSPIN), 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 Paris-Saclay-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 Paris-Saclay, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-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), Microbiologie Environnementale et Moléculaire (MEM), 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)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Service de Pharmacologie et Immunoanalyse (SPI), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Observatoire des Sciences de l'Univers de Grenoble (OSUG ), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Institut national des sciences de l'Univers (INSU - CNRS)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Service de Pharmacologie et d'Immunoanalyse (SPI), Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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 National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-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)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)

Subjects
0301 basic medicine, Materials science, media_common.quotation_subject, Magnetosome, Biophysics, Brain tumor, Contrast Media, Molecular Probe Techniques, Bioengineering, Single step, Nanoconjugates, 02 engineering and technology, Article, Biomaterials, Mice, 03 medical and health sciences, Nuclear magnetic resonance, Cell Line, Tumor, Biomarkers, Tumor, medicine, Animals, Tissue Distribution, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Internalization, media_common, medicine.diagnostic_test, Brain Neoplasms, RGD peptide, Magnetic resonance imaging, 021001 nanoscience & nanotechnology, medicine.disease, Magnetic Resonance Imaging, Molecular Imaging, Genetic Enhancement, 030104 developmental biology, Mechanics of Materials, Molecular Probes, Ceramics and Composites, Magnetosomes, Lipid vesicle, Molecular imaging, 0210 nano-technology, Oligopeptides
Abstract

International audience; We investigate here the potential of single step production of genetically engineeredmagnetosomes, bacterial biogenic iron-oxide nanoparticles embedded in a lipid vesicle, as a new tailorable magnetic resonance molecular imaging probe. We demonstrate in vitro the specific binding and the significant internalization into U87 cells of magnetosomes decorated with RGD peptide. After injection at the tail vein of glioblastoma-bearing mice, we evidence in the first 2 h the rapid accumulation of both unlabeled and functionalized magnetosomes inside the tumor by Enhanced Permeability and Retention effects. 24 h after the injection, a specific enhancement of the tumor contrast is observed on MR images only for RGD-labeled magnetosomes. Post mortem acquisition of histological data confirms MRI results with more magnetosomes found into the tumor treated with functionalized magnetosomes. This work establishes the first proof-of-concept of a successful bio-integrated production of molecular imaging probe for MRI.

Published
2017

26. Conservation and diversity of the IrrE/DdrO-controlled radiation response in radiation-resistant Deinococcus bacteria

Author

Laurence, Blanchard, Philippe, Guérin, David, Roche, Stéphane, Cruveiller, David, Pignol, David, Vallenet, Jean, Armengaud, and Arjan, de Groot

Subjects
Genetic Variation, Gene Expression Regulation, Bacterial, zinc peptidase, Regulon, transcriptional repressor, Repressor Proteins, transcriptomics, proteomics, Bacterial Proteins, Stress, Physiological, Metalloproteases, population characteristics, bacteria, Deinococcus, gene regulation, Original Research
Abstract

The extreme radiation resistance of Deinococcus bacteria requires the radiation‐stimulated cleavage of protein DdrO by a specific metalloprotease called IrrE. DdrO is the repressor of a predicted radiation/desiccation response (RDR) regulon, composed of radiation‐induced genes having a conserved DNA motif (RDRM) in their promoter regions. Here, we showed that addition of zinc ions to purified apo‐IrrE, and short exposure of Deinococcus cells to zinc ions, resulted in cleavage of DdrO in vitro and in vivo, respectively. Binding of IrrE to RDRM‐containing DNA or interaction of IrrE with DNA‐bound DdrO was not observed. The data are in line with IrrE being a zinc peptidase, and indicate that increased zinc availability, caused by oxidative stress, triggers the in vivo cleavage of DdrO unbound to DNA. Transcriptomics and proteomics of Deinococcus deserti confirmed the IrrE‐dependent regulation of predicted RDR regulon genes and also revealed additional members of this regulon. Comparative analysis showed that the RDR regulon is largely well conserved in Deinococcus species, but also showed diversity in the regulon composition. Notably, several RDR genes with an important role in radiation resistance in Deinococcus radiodurans, for example pprA, are not conserved in some other radiation‐resistant Deinococcus species.

Published
2017

27. Toward inline multiplex biodetection of metals, bacteria, and toxins in water networks: the COMBITOX project

Author

Pierre Cholat, Agnès Rodrigue, Mireille Ansaldi, David Pignol, Ingrid Bazin, Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut de Microbiologie de la Méditerranée (IMM), Laboratoire de Génie de l'Environnement Industriel (LGEI), IMT - MINES ALES (IMT - MINES ALES), Institut Mines-Télécom [Paris] (IMT)-Institut Mines-Télécom [Paris] (IMT), Trafic et signalisation membranaires chez les bactéries (MTSB), Microbiologie, adaptation et pathogénie (MAP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), ANR-11-ECOT-0009,COMBITOX,COnception d'un instrument pour la Mesure BIologique multiparamètrique en continu de TOXiques(2011), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon, 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Engineering, Health, Toxicology and Mutagenesis, 02 engineering and technology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 010501 environmental sciences, Bioinformatics, 01 natural sciences, Human health, Time response, Environmental Chemistry, Multiplex, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 0105 earth and related environmental sciences, Light detector, [SDV.GEN]Life Sciences [q-bio]/Genetics, Physical conditioning, business.industry, General Medicine, 021001 nanoscience & nanotechnology, Pollution, 6. Clean water, 13. Climate action, [SDE]Environmental Sciences, Biochemical engineering, 0210 nano-technology, business, Biosensor, Conceptual level, Field conditions
Abstract

This special issue of Environmental Science and Pollution Research highlights selected papers whose results have been obtained in the course of the COMBITOX project. COMBITIOX is an interdisciplinary research project funded by the French National Research Agency (ANR) aiming at conceiving an inline multiparametric device for the surveillance of water networks using biosensors. This device is not intended to fully replace chemical methods, but when compared to analytical chromatographic methodologies, biological sensors can offer rapid and on-site monitoring of even trace levels of targeted compounds (Sun et al. 2015) and can quickly raise the alarm in the event of an accidental or intentional pollution. Numerous developments have been published to improve the sensitivity, specificity, and time response of various biosensors in laboratory conditions (Xiong et al. 2012) (der Meer et al. 2010), but their actual transfer into technological devices for the surveillance of water networks remains at a conceptual level. Thus, the challenge here is to go a step beyond and validate biosensors under real-life field conditions by incorporating them in a single inline detector. During the course of COMBITOX, we could define the interface between the biosensors and a common light detector as well as the physical conditioning of the bioreagents and usage protocol. Our resulting prototype allow the detection of bioavailable toxic compounds as well as microorganisms, impacting human health through the drinking water network or interfering with the biological process of modern wastewater treatment plants. We also plan to propose this system to meet the emerging threats such as bioterrorism. COMBITOX focuses on three families of Bobjects^ to detect: metals (cadmium, mercury, arsenic, nickel, etc.), environmental and/or food toxins, and pathogenic microorganisms. Whole-cell biosensors based on reporter gene under the control of an inducible promoter are used to detect various metals (Hynninen and Virta 2010), the antibody/antigen interaction for toxins (Makaraviciute and Ramanaviciene 2013), and the specific infection of bacteria by bacteriophages for pathogenic microorganisms (Smartt et al. 2012) (Vinay et al. 2015). In all cases, the signal measured is photochemical (fluorescence, bioluminescence, or chemo-luminescence): such a method to transduce the biological recognition is very sensitive and a single photodetector can be used for all biosensors included in the device. The challenge here rather lies in the design and the optimization of the different biological compounds for their use in the field while maintaining a high sensibility and robustness. As a consequence, the different articles presented in this special issue focus on original strategies for the optimization and the adaptation of the three types of biosensors for their use in a semi-autonomous inline water analyzer. In the case of whole-cell biosensors, improvement of the dose-responses Responsible editor: Philippe Garrigues

Published
2017

28. Semi-autonomous inline water analyzer: design of a common light detector for bacterial, phage, and immunological biosensors

Author

Nicolas Ginet, Mireille Ansaldi, Agnès Rodrigue, Sandra Prévéral, Damien Meunier, Bertrand Miclot, Camille Escoffier, Jean-Raphaël Fantino, Nathalie Franche, Philippe Larosa, Ingrid Bazin, Elodie C. T. Descamps, David Pignol, Pierre Cholat, Catherine Brutesco, Daniel Garcia, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut de Microbiologie de la Méditerranée (IMM), Laboratoire de Génie de l'Environnement Industriel (LGEI), IMT - MINES ALES (IMT - MINES ALES), Institut Mines-Télécom [Paris] (IMT)-Institut Mines-Télécom [Paris] (IMT), Trafic et signalisation membranaires chez les bactéries (MTSB), Microbiologie, adaptation et pathogénie (MAP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), and Université de Lyon

Subjects
0301 basic medicine, Spectrum analyzer, Engineering, Light, Health, Toxicology and Mutagenesis, Environmental pollution, Nanotechnology, Biosensing Techniques, Technology readiness level, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 010501 environmental sciences, 01 natural sciences, 03 medical and health sciences, Water Quality, Environmental Chemistry, Bacteriophages, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Reaction chamber, 0105 earth and related environmental sciences, Light detector, [SDV.GEN]Life Sciences [q-bio]/Genetics, Bacteria, business.industry, Water, General Medicine, Pollution, 030104 developmental biology, [SDE]Environmental Sciences, Water Microbiology, business, Biosensor, Food Analysis, Computer hardware, Environmental Monitoring
Abstract

International audience; The use of biosensors as sensitive and rapid alert systems is a promising perspective to monitor accidental or intentional environmental pollution, but their implementation in the field is limited by the lack of adapted inline water monitoring devices. We describe here the design and initial qualification of an analyzer prototype able to accommodate three types of biosensors based on entirely different methodologies (immunological, whole-cell, and bacteriophage biosensors), but whose responses rely on the emission of light. We developed a custom light detector and a reaction chamber compatible with the specificities of the three systems and resulting in statutory detection limits. The water analyzer prototype resulting from the COMBITOX project can be situated at level 4 on the Technology Readiness Level (TRL) scale and this technical advance paves the way to the use of biosensors on-site.

Published
2017

29. Bacterial host and reporter gene optimization for genetically encoded whole cell biosensors

Author

Nicolas Ginet, Elodie C. T. Descamps, Géraldine Adryanczyk-Perrier, Louis Dumas, Agnès Rodrigue, David Pignol, Arjan de Groot, Julien Cayron, Elsa Prudent, Sandra Prévéral, Camille Escoffier, Manon Ricquebourg, Catherine Brutesco, Daniel Garcia, Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Microbiologie Environnementale et Moléculaire (MEM), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Microbiologie, adaptation et pathogénie (MAP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Bioénergie et Microalgues (EBM), Trafic et signalisation membranaires chez les bactéries (MTSB), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), ANR-11-ECOT-0009,COMBITOX,COnception d'un instrument pour la Mesure BIologique multiparamètrique en continu de TOXiques(2011), Biologie végétale et microbiologie environnementale - UMR7265 ( BVME ), Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Aix Marseille Université ( AMU ), Ecologie microbienne ( EM ), Centre National de la Recherche Scientifique ( CNRS ) -Ecole Nationale Vétérinaire de Lyon ( ENVL ) -Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Recherche Agronomique ( INRA ) -VetAgro Sup ( VAS ), Centre Sciences des Processus Industriels et Naturels ( SPIN-ENSMSE ), École des Mines de Saint-Étienne ( Mines Saint-Étienne MSE ), Institut Mines-Télécom [Paris]-Institut Mines-Télécom [Paris], Département GéoSciences et Environnement ( GSE-ENSMSE ), Institut Mines-Télécom [Paris]-Institut Mines-Télécom [Paris]-SPIN, Environnement Ville Société ( EVS ), École normale supérieure - Lyon ( ENS Lyon ) -Université Lumière - Lyon 2 ( UL2 ) -Université Jean Moulin - Lyon III-Institut National des Sciences Appliquées de Lyon ( INSA Lyon ), Université de Lyon-Institut National des Sciences Appliquées ( INSA ) -Université de Lyon-Institut National des Sciences Appliquées ( INSA ) -Université Jean Monnet [Saint-Étienne] ( UJM ) -École Nationale des Travaux Publics de l'État ( ENTPE ) -Ecole Nationale Supérieure des Mines de Saint-Etienne-École nationale supérieure d'architecture de Lyon ( ENSAL ) -Centre National de la Recherche Scientifique ( CNRS ), Microbiologie, adaptation et pathogénie ( MAP ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon ( INSA Lyon ), Université de Lyon-Institut National des Sciences Appliquées ( INSA ) -Institut National des Sciences Appliquées ( INSA ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire de chimie bactérienne ( LCB ), Aix Marseille Université ( AMU ) -Centre National de la Recherche Scientifique ( CNRS ), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Environnement, Bioénergie, Microalgues et Plantes (EBMP), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon, and Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées de Lyon (INSA Lyon)

Subjects
0301 basic medicine, Arsenites, Health, Toxicology and Mutagenesis, Biosensing Techniques, macromolecular substances, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 010501 environmental sciences, medicine.disease_cause, 01 natural sciences, Gene Expression Regulation, Enzymologic, Microbiology, [ SDE ] Environmental Sciences, 03 medical and health sciences, chemistry.chemical_compound, Genes, Reporter, Metals, Heavy, Escherichia coli, medicine, Environmental Chemistry, Bioluminescence, Deinococcus, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Promoter Regions, Genetic, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 0105 earth and related environmental sciences, Arsenite, Reporter gene, [SDV.GEN]Life Sciences [q-bio]/Genetics, biology, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, technology, industry, and agriculture, Water, Gene Expression Regulation, Bacterial, General Medicine, biology.organism_classification, Deinococcus deserti, Pollution, Luciferases, Bacterial, 030104 developmental biology, chemistry, Biochemistry, [SDE]Environmental Sciences, [ SDV.GEN ] Life Sciences [q-bio]/Genetics, Biosensor, Water Pollutants, Chemical, Bacteria, Environmental Monitoring
Abstract

International audience; Whole-cell biosensors based on reporter genes allow detection of toxic metals in water with high selectivity and sensitivity under laboratory conditions; nevertheless, their transfer to a commercial inline water analyzer requires specific adaptation and optimization to field conditions as well as economical considerations. We focused here on both the influence of the bacterial host and the choice of the reporter gene by following the responses of global toxicity biosensors based on constitutive bacterial promoters as well as arsenite biosensors based on the arsenite-inducible P-ars promoter. We observed important variations of the bioluminescence emission levels in five different Escherichia coli strains harboring two different lux-based biosensors, suggesting that the best host strain has to be empirically selected for each new biosensor under construction. We also investigated the bioluminescence reporter gene system transferred into Deinococcus deserti, an environmental, desiccation- and radiation-tolerant bacterium that would reduce the manufacturing costs of bacterial biosensors for commercial water analyzers and open the field of biodetection in radioactive environments. We thus successfully obtained a cell survival biosensor and a metal biosensor able to detect a concentration as low as 100 nM of arsenite in D. deserti. We demonstrated that the arsenite biosensor resisted desiccation and remained functional after 7 days stored in air-dried D. deserti cells. We also report here the use of a new near-infrared (NIR) fluorescent reporter candidate, a bacteriophytochrome from the magnetotactic bacterium Magnetospirillum magneticum AMB-1, which showed a NIR fluorescent signal that remained optimal despite increasing sample turbidity, while in similar conditions, a drastic loss of the lux-based biosensors signal was observed.

Published
2017

30. Kinetics of substrate inhibition of periplasmic nitrate reductase

Author

Bénédicte Burlat, Pascal Arnoux, Christophe Léger, Bruno Guigliarelli, Julien Jacques, David Pignol, Monique Sabaty, Vincent Fourmond, Bioénergétique et Ingénierie des Protéines (BIP ), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Stereochemistry, Biophysics, Reductase, Nitrate reductase, Photochemistry, Biochemistry, Substrate Specificity, chemistry.chemical_compound, Nitrate, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Electrochemistry, Enzyme kinetics, Nitrites, DMSO reductase, [PHYS.PHYS]Physics [physics]/Physics [physics], Substrate (chemistry), Cell Biology, Periplasmic space, Kinetics, chemistry, Periplasm, Thermodynamics, Molybdenum cofactor, Oxidation-Reduction, EPR spectroscopy
Abstract

International audience; Periplasmic nitrate reductase catalyzes the reduction of nitrate into nitrite using a mononuclear molybdenum cofactor that has nearly the same structure in all enzymes of the DMSO reductase family. In previous electrochemical investigations, we found that the enzyme exists in several inactive states, some of which may have been previously isolated and mistaken for catalytic intermediates. In particular, the enzyme slowly and reversibly inactivates when exposed to high concentrations of nitrate. Here, we study the kinetics of substrate inhibition and its dependence on electrode potential and substrate concentration to learn about the properties of the active and inactive forms of the enzyme. We conclude that the substrate-inhibited enzyme never significantly accumulates in the EPR-active Mo(+ V) state. This conclusion is relevant to spectroscopic investigations where attempts are made to trap a Mo(+ V) catalytic intermediate using high concentrations of nitrate.

Published
2014

31. Radiation response inDeinococcus deserti: IrrE is a metalloprotease that cleaves repressor protein DdrO

Author

David Lemaire, David Pignol, Laurent Bellanger, Arjan de Groot, Géraldine Brandelet, Laurence Blanchard, Monika Ludanyi, and Rémi Dulermo

Subjects
0303 health sciences, Metalloproteinase, biology, 030306 microbiology, DNA repair, Repressor, Deinococcus deserti, medicine.disease_cause, biology.organism_classification, Microbiology, 03 medical and health sciences, Biochemistry, parasitic diseases, medicine, bacteria, population characteristics, Deinococcus, Molecular Biology, Psychological repression, Gene, Escherichia coli, 030304 developmental biology
Abstract

Deinococcus bacteria are famous for their extreme radiation tolerance. The IrrE protein was shown to be essential for radiation tolerance and, in an unelucidated manner, for induction of a number of genes in response to radiation, including recA and other DNA repair genes. Earlier studies indicated that IrrE could be a zinc peptidase, but proteolytic activity was not demonstrated. Here, using several in vivo and in vitro experiments, IrrE from Deinococcus deserti was found to interact with DdrO, a predicted regulator encoded by a radiation-induced gene that is, like irrE, highly conserved in Deinococcus. Moreover, IrrE was found to cleave DdrO in vitro and when the proteins were coexpressed in Escherichia coli. This cleavage was not observed in the presence of metal chelator EDTA or when IrrE contains a mutation in the conserved active-site motif of metallopeptidases. In D. deserti, IrrE-dependent cleavage of DdrO was observed after exposure to radiation. Furthermore, DdrO-dependent repression of the promoter of a radiation-induced gene was shown. These results demonstrate that IrrE is a metalloprotease and we propose that IrrE-mediated cleavage inactivates repressor protein DdrO, leading to transcriptional induction of various genes required for repair and survival after exposure of Deinococcus to radiation.

Published
2014

32. Comparative genomic analysis provides insights into the evolution and niche adaptation of marineMagnetospirasp. QH-2 strain

Author

Hongmiao Pan, Sophie Mangenot, Long-Fei Wu, Ying Li, Jing Yang, Zoé Rouy, Claudine Médigue, Emmanuel Talla, Valérie Barbe, Jean-Baptiste Rioux, Pedro M. Coutinho, Nadège Philippe, Tian Xiao, François Alberto, Wei-Jia Zhang, Jiesheng Tian, Nicolas Ginet, Bernard Henrissat, Sheng-Da Zhang, Boyang Ji, Lichen Zhang, Wenyan Zhang, David Pignol, Dorothée Murat, Pascal Arnoux, Monique Sabaty, and Nathalie Pradel

Subjects
0303 health sciences, Magnetotactic bacteria, biology, 030306 microbiology, Magnetosome, Spirillum, biology.organism_classification, Microbiology, Genome, 03 medical and health sciences, Evolutionary biology, Gene cluster, Botany, 14. Life underwater, Niche adaptation, Magnetospirillum, Ecology, Evolution, Behavior and Systematics, 030304 developmental biology, Synteny
Abstract

Magnetotactic bacteria (MTB) are capable of synthesizing intracellular organelles, the magnetosomes, that are membrane-bounded magnetite or greigite crystals arranged in chains. Although MTB are widely spread in various ecosystems, few axenic cultures are available, and only freshwater Magnetospirillum spp. have been genetically analysed. Here, we present the complete genome sequence of a marine magnetotactic spirillum, Magnetospira sp. QH-2. The high number of repeats and transposable elements account for the differences in QH-2 genome structure compared with other relatives. Gene cluster synteny and gene correlation analyses indicate that the insertion of the magnetosome island in the QH-2 genome occurred after divergence between freshwater and marine magnetospirilla. The presence of a sodium-quinone reductase, sodium transporters and other functional genes are evidence of the adaptive evolution of Magnetospira sp. QH-2 to the marine ecosystem. Genes well conserved among freshwater magnetospirilla for nitrogen fixation and assimilatory nitrate respiration are absent from the QH-2 genome. Unlike freshwater Magnetospirillum spp., marine Magnetospira sp. QH-2 neither has TonB and TonB-dependent receptors nor does it grow on trace amounts of iron. Taken together, our results show a distinct, adaptive evolution of Magnetospira sp. QH-2 to marine sediments in comparison with its closely related freshwater counterparts.

Published
2013

33. Monophyletic origin of magnetotaxis and the first magnetosomes

Author

Sebastian Kolinko, Christopher T. Lefèvre, Dennis A. Bazylinski, Fernanda Abreu, Dirk Schüler, Ulysses Lins, Luiz Gonzaga Paula de Almeida, Denis Trubitsyn, Ana Tereza Ribeiro de Vasconcelos, Nicolas Ginet, and David Pignol

Subjects
Genetics, biology, Magnetotactic bacteria, Phylogenetic tree, Genomic island, Magnetosome, Horizontal gene transfer, Magnetotaxis, Proteobacteria, biology.organism_classification, Microbiology, Gene, Ecology, Evolution, Behavior and Systematics
Abstract

Summary Horizontal gene transfer (HGT), the transfer of genetic material other than by descent, is thought to have played significant roles in the evolution and distribution of genes in prokaryotes. These include those responsible for the ability of motile, aquatic magnetotactic bacteria (MTB) to align and swim along magnetic field lines and the biomineralization of magnetosomes that are responsible for this behaviour. There is some genomic evidence that HGT might be responsible for the distribution of magnetosome genes in different phylogenetic groups of bacteria. For example, in the genomes of a number of MTB, magnetosome genes are present as clusters within a larger structure known as the magnetosome genomic island surrounded by mobile elements such as insertion sequences and transposases as well as tRNA genes. Despite this, there is no strong direct proof of HGT between these organisms. Here we show that a phylogenetic tree based on magnetosome protein amino acid sequences from a number of MTB was congruent with the tree based on the organisms' 16S rRNA gene sequences. This shows that evolution and divergence of these proteins and the 16S rRNA gene occurred similarly. This suggests that magnetotaxis originated monophyletically in the Proteobacteria phylum and implies that the common ancestor of all Proteobacteria was magnetotactic.

Published
2013

34. Biosynthesis of a broad-spectrum nicotianamine-like metallophore in Staphylococcus aureus

Author

Ryszard Lobinski, Elise Borezée-Durant, Felipe Cava, David Lemaire, Ghassan Ghssein, Christine Hajjar, Catherine Brutesco, Amélie Izaute, David Pignol, Romé Voulhoux, Pierre Richaud, Pascal Arnoux, Akbar Espaillat, Laurent Ouerdane, Shuanglong Wang, Clémentine Fojcik, Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Microbiologie Environnementale et Moléculaire (MEM), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-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)-Aix Marseille Université (AMU)-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 sciences analytiques et de physico-chimie pour l'environnement et les materiaux (IPREM), Université de Pau et des Pays de l'Adour (UPPA)-Centre National de la Recherche Scientifique (CNRS), MICrobiologie de l'ALImentation au Service de la Santé (MICALIS), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Interactions Protéine Métal (IPM), Bioénergie et Microalgues (EBM), Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Umeå University, CEA program ToxNuc-E, HélioBiotec platform, ANR-14-CE09-0007,ANIBAL,Un nouveau metallophore dérivé de la nicotianamine chez des bactéries pathogènes(2014), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie Analytique Bio-Inorganique et Environnement (LCABIE), Laboratory for Molecular Infection Medicine Sweden, Department of Molecular Biology, Umeå Centre of Microbial Research, Umeå, Sweden, French Atomic Energy CommissionCEA program ToxNuc-E HelioBiotec platform - European Regional Development Fund Région Provence-Alpes-Côte d'AzurHelioBiotec platform - French Ministry of Research HelioBiotec platform - 'Commissariat a l'Energie Atomique et aux Energies Alternatives' Région Provence-Alpes-Côte d'AzurLaboratory for Molecular Infection Medicine Sweden Knut & Alice Wallenberg FoundationSwedish Research CouncilEuropean CommissionKempe foundation China Scholarship Council, Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Lobinski, Ryszard, and Appel à projets générique - Un nouveau metallophore dérivé de la nicotianamine chez des bactéries pathogènes - - ANIBAL2014 - ANR-14-CE09-0007 - Appel à projets générique - VALID

Subjects
0301 basic medicine, Operon, [SDV]Life Sciences [q-bio], transferase, Dehydrogenase, Nicotianamine synthase, chemistry.chemical_compound, iron, Transferase, pathogenicity, Nicotianamine, oxidoreductase, azetidine 2 carboxylic acid, Amino Acid Isomerases, chemistry.chemical_classification, Multidisciplinary, metallophore, nicotianamine synthase, zinc, Imidazoles, Cobalt, [SDV] Life Sciences [q-bio], Biochemistry, Metabolome, Oxidoreductases, Azetidinecarboxylic Acid, Staphylococcus aureus, enzymology, 030106 microbiology, Biology, Biosynthesis, microbial activity, nicotine derivative, 03 medical and health sciences, nickel, Oxidoreductase, [CHIM]Chemical Sciences, Histidine, Alkyl and Aryl Transferases, nicotianamine, mutagenicity, Gene Expression Regulation, Bacterial, histidine, heavy metal, operon, Biosynthetic Pathways, staphylopine, 030104 developmental biology, chemistry, Gene Expression Regulation, copper, biology.protein, phytochemistry, imidazole derivative
Abstract

A new metal scavenger for bacteria All cells must find a way to acquire trace metals. Bacteria and plants scavenge iron, for instance, by synthesizing and releasing iron-chelating compounds called siderophores. Ghssein et al. describe three enzymes in Staphylococcus aureus that are responsible for the biosynthesis of another type of metallophore (see the Perspective by Nolan). Metabolomics and a range of biochemical assays show that this compound, named staphylopine, is involved in the uptake of a range of metals, depending on the growth environment. The genes required for staphylopine biosynthesis are conserved across a number of pathogenic bacteria and are similar to those for a broad-spectrum metallophore produced by plants. Science , this issue p. 1105 ; see also p. 1055

Published
2016

35. Magnetochrome: a c-type cytochrome domain specific to magnetotatic bacteria

Author

David Pignol, Marina I. Siponen, Géraldine Adryanczyk, Nicolas Ginet, and Pascal Arnoux

Subjects
Biocrystallization, biology, Magnetotactic bacteria, Cytochrome, Cytochrome c, Molecular Sequence Data, Magnetosome, Cytochrome c Group, biology.organism_classification, Biochemistry, Protein Structure, Tertiary, Electron Transport, Bacterial Proteins, Organelle, Magnetotaxis, biology.protein, Amino Acid Sequence, Magnetosomes, Magnetospirillum, Crystallization, Oxidation-Reduction
Abstract

Magnetotactic bacteria consist of a group of taxonomically, physiologically and morphologically diverse prokaryotes, with the singular ability to align with geomagnetic field lines, a phenomenon referred to as magnetotaxis. This magnetotactic property is due to the presence of iron-rich crystals embedded in lipidic vesicles forming an organelle called the magnetosome. Magnetosomes are composed of single-magnetic-domain nanocrystals of magnetite (Fe 3 O 4 ) or greigite (Fe 3 S 4 ) embedded in biological membranes, thereby forming a prokaryotic organelle. Four specific steps are described in this organelle formation: (i) membrane specialization, (ii) iron acquisition, (iii) magnetite (or greigite) biocrystallization, and (iv) magnetosome alignment. The formation of these magnetic crystals is a genetically controlled process, which is governed by enzyme-catalysed processes. On the basis of protein sequence analysis of genes known to be involved in magnetosome formation in Magnetospirillum magneticum AMB-1, we have identified a subset of three membrane-associated or periplasmic proteins containing a double cytochrome c signature motif CXXCH: MamE, MamP and MamT. The presence of these proteins suggests the existence of an electron-transport chain inside the magnetosome, contributing to the process of biocrystallization. We have performed heterologous expression in E. coli of the cytochrome c motif-containing domains of MamE, MamP and MamT. Initial biophysical characterization has confirmed that MamE, MamP and MamT are indeed c -type cytochromes. Furthermore, determination of redox potentials for this new family of c -type cytochromes reveals midpoint potentials of −76 and −32 mV for MamP and MamE respectively.

Published
2012

36. Coproporphyrin Excretion and Low Thiol Levels Caused by Point Mutation in the Rhodobacter sphaeroides S -Adenosylmethionine Synthetase Gene

Author

Christine Lamouroux, Chloë Roustan, Géraldine Adryanczyk, David Pignol, Monique Sabaty, and Stéphan Cuiné

Subjects
DNA, Bacterial, Coproporphyrins, S-Adenosylmethionine, Physiology and Metabolism, DNA Mutational Analysis, Molecular Sequence Data, Mutant, Mutation, Missense, Rhodobacter sphaeroides, Cysteine synthase, Nitrate reductase, Models, Biological, Microbiology, chemistry.chemical_compound, Point Mutation, Amino Acid Sequence, Cysteine, Sulfhydryl Compounds, Molecular Biology, biology, Genetic Complementation Test, Methionine Adenosyltransferase, Sequence Analysis, DNA, Glutathione, biology.organism_classification, Molecular biology, Amino Acid Substitution, chemistry, Biochemistry, Uroporphyrinogen III, biology.protein, Molybdenum cofactor, Sequence Alignment
Abstract

A spontaneous mutant of Rhodobacter sphaeroides f. sp. denitrificans IL-106 was found to excrete a large amount of a red compound identified as coproporphyrin III, an intermediate in bacteriochlorophyll and heme synthesis. The mutant, named PORF, is able to grow under phototrophic conditions but has low levels of intracellular cysteine and glutathione and overexpresses the cysteine synthase CysK. The expression of molybdoenzymes such as dimethyl sulfoxide (DMSO) and nitrate reductases is also affected under certain growth conditions. Excretion of coproporphyrin and overexpression of CysK are not directly related but were both found to be consequences of a diminished synthesis of the key metabolite S -adenosylmethionine (SAM). The wild-type phenotype is restored when the gene metK encoding SAM synthetase is supplied in trans . The metK gene in the mutant strain has a mutation leading to a single amino acid change (H145Y) in the encoded protein. This point mutation is responsible for a 70% decrease in intracellular SAM content which probably affects the activities of numerous SAM-dependent enzymes such as coproporphyrinogen oxidase (HemN); uroporphyrinogen III methyltransferase (CobA), which is involved in siroheme synthesis; and molybdenum cofactor biosynthesis protein A (MoaA). We propose a model showing that the attenuation of the activities of SAM-dependent enzymes in the mutant could be responsible for the coproporphyrin excretion, the low cysteine and glutathione contents, and the decrease in DMSO and nitrate reductase activities.

Published
2010

37. Magnetosomes, Biogenic Magnetic Nanomaterials for Brain Molecular Imaging with 17.2 T MRI Scanner

Author

Laurence Motte, Benjamin Marty, Franck Lethimonnier, Sandra Prévéral, Christopher T. Lefèvre, Nicolas Ginet, Yoann Lalatonne, Géraldine Adryanczyk-Perrier, Françoise Geffroy, David Pignol, Michel Pean, Marianne Boucher, Sébastien Mériaux, Daniel Garcia, Service NEUROSPIN (NEUROSPIN), Université Paris-Saclay-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), Chimie, Structures et Propriétés de Biomatériaux et d'Agents Thérapeutiques (CSPBAT), Université Paris 13 (UP13)-Institut Galilée-Université Sorbonne Paris Cité (USPC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (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 Paris-Saclay, and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Materials science, Brain vasculature, Magnetotactic bacteria, Magnetosome, Biomedical Engineering, Pharmaceutical Science, Contrast Media, 02 engineering and technology, 010402 general chemistry, 01 natural sciences, Nanomaterials, Biomaterials, chemistry.chemical_compound, Magnetics, Mice, Nuclear magnetic resonance, In vivo, medicine, Animals, [CHIM]Chemical Sciences, Magnetospirillum, Magnetite Nanoparticles, ComputingMilieux_MISCELLANEOUS, Magnetite, medicine.diagnostic_test, Brain, Magnetic resonance imaging, Dextrans, 021001 nanoscience & nanotechnology, Magnetic Resonance Imaging, Ferrosoferric Oxide, 0104 chemical sciences, 3. Good health, Molecular Imaging, Nanostructures, chemistry, Nanoparticles, Magnetosomes, Molecular imaging, 0210 nano-technology, Biomedical engineering
Abstract

The fast development of sensitive molecular diagnostic tools is currently paving the way for a personalized medicine. A new class of ultrasensitive magnetic resonance imaging (MRI) T₂-contrast agents based on magnetosomes, magnetite nanocrystals biomineralized by magnetotactic bacteria, is proposed here. The contrast agents can be injected into the blood circulation and detected in the picomolar range. Purified magnetosomes are water-dispersible and stable within physiological conditions and exhibit at 17.2 T a transverse relaxivity r₂ four times higher than commercial ferumoxide. The subsequent gain in sensitivity by T₂(*) -weighted imaging at 17.2 T of the mouse brain vasculature is evidenced in vivo after tail vein injection of magnetosomes representing a low dose of iron (20 μmoliron kg(-1)), whereas no such phenomenon with the same dose of ferumoxide is observed. Preclinical studies of human pathologies in animal models will benefit from the combination of high magnetic field MRI with sensitive, low dose, easy-to-produce biocompatible contrast agents derived from bacterial magnetosomes.

Published
2015

38. Genetic and Biochemical Evidence for the Involvement of a Molybdenum-Dependent Enzyme in One of the Selenite Reduction Pathways ofRhodobacter sphaeroidesf. sp.denitrificansIL106

Author

Monique Sabaty, Bénédicte Pierru, Sandrine Grosse, and David Pignol

Subjects
Coenzymes, chemistry.chemical_element, Rhodobacter sphaeroides, Nitrate reductase, Applied Microbiology and Biotechnology, Cofactor, chemistry.chemical_compound, Sodium Selenite, Metalloproteins, Dimethyl Sulfoxide, Oxidoreductases Acting on Sulfur Group Donors, chemistry.chemical_classification, DMSO reductase, Nitrates, Ecology, biology, Dimethyl sulfoxide, Pteridines, Tungsten Compounds, Physiology and Biotechnology, biology.organism_classification, Culture Media, Enzyme, chemistry, Biochemistry, biology.protein, Oxidoreductases, Molybdenum cofactor, Molybdenum Cofactors, Oxidation-Reduction, Selenium, Food Science, Biotechnology
Abstract

Selenite reduction inRhodobacter sphaeroidesf. sp.denitrificanswas observed under photosynthetic conditions, following a 100-h lag period. This adaptation period was suppressed if the medium was inoculated with a culture previously grown in the presence of selenite, suggesting that selenite reduction involves an inducible enzymatic pathway. A transposon library was screened to isolate mutants affected in selenite reduction. Of the eight mutants isolated, two were affected in molybdenum cofactor synthesis. ThesemoaAandmogAmutants showed an increased duration of the lag phase and a decreased rate of selenite reduction. When grown in the presence of tungstate, a well-known molybdenum-dependent enzyme (molybdoenzyme) inhibitor, the wild-type strain displayed the same phenotype. The addition of tungstate in the medium or the inactivation of the molybdocofactor synthesis induced a decrease of 40% in the rate of selenite reduction. These results suggest that several pathways are involved and that one of them involves a molybdoenzyme. Although addition of nitrate or dimethyl sulfoxide (DMSO) to the medium increased the selenite reduction activity of the culture, neither the periplasmic nitrate reductase NAP nor the DMSO reductase is the implicated molybdoenzyme, since thenapAanddmsAmutants, with expression of nitrate reductase and DMSO reductase, respectively, eliminated, were not affected by selenite reduction. A role for the biotine sulfoxide reductase, another characterized molybdoenzyme, is unlikely, since its overexpression in a defective strain did not restore the selenite reduction activity.

Published
2006

39. A papain-like enzyme at work: Native and acyl–enzyme intermediate structures in phytochelatin synthesis

Author

Denis Vivares, Pascal Arnoux, and David Pignol

Subjects
Anions, Models, Molecular, Time Factors, Protein Conformation, Stereochemistry, Molecular Sequence Data, Arabidopsis, Crystallography, X-Ray, Catalysis, Xenobiotics, chemistry.chemical_compound, Protein structure, Catalytic Domain, Metals, Heavy, Papain, Hydrolase, Escherichia coli, Phytochelatins, Amino Acid Sequence, Cysteine, Binding site, Nostoc, Glutathione Transferase, chemistry.chemical_classification, Binding Sites, Multidisciplinary, Sequence Homology, Amino Acid, ATP synthase, biology, Temperature, Hydrogen Bonding, Glutathione, Biological Sciences, Hydrogen-Ion Concentration, Aminoacyltransferases, Protein Structure, Tertiary, Enzyme, chemistry, Biochemistry, Commentary, biology.protein, Dimerization, Software, Protein Binding
Abstract

Phytochelatin synthase (PCS) is a key enzyme for heavy-metal detoxification in plants. PCS catalyzes the production of glutathione (GSH)-derived peptides (called phytochelatins or PCs) that bind heavy-metal ions before vacuolar sequestration. The enzyme can also hydrolyze GSH and GS-conjugated xenobiotics. In the cyanobacterium Nostoc , the enzyme (NsPCS) contains only the catalytic domain of the eukaryotic synthase and can act as a GSH hydrolase and weakly as a peptide ligase. The crystal structure of NsPCS in its native form solved at a 2.0-Å resolution shows that NsPCS is a dimer that belongs to the papain superfamily of cysteine proteases, with a conserved catalytic machinery. Moreover, the structure of the protein solved as a complex with GSH at a 1.4-Å resolution reveals a γ-glutamyl cysteine acyl–enzyme intermediate stabilized in a cavity of the protein adjacent to a second putative GSH binding site. GSH hydrolase and PCS activities of the enzyme are discussed in the light of both structures.

Published
2005

40. PpsR: a multifaceted regulator of photosynthesis gene expression in purple bacteria

Author

Marianne Jaubert, Sylvie Elsen, David Pignol, and Eric Giraud

Subjects
Regulation of gene expression, 0303 health sciences, Rhodobacter, biology, 030306 microbiology, Regulator, food and beverages, Photosynthesis, biology.organism_classification, Microbiology, Purple bacteria, Cell biology, 03 medical and health sciences, Rhodobacter sphaeroides, Molecular Biology, 030304 developmental biology, Photosystem, Regulator gene
Abstract

Purple bacteria control the level of expression and the composition of their photosystem according to light and redox conditions. This control involves several regulatory systems that have been now well characterized. Among them, the PpsR regulator plays a central role, because it directly or indirectly controls the synthesis of all of the different components of the photosystem. In this review, we report our knowledge of the PpsR protein, highlighting the diversity of its mode of action and focusing on the proteins identified in four model purple bacteria (Rhodobacter capsulatus, Rhodobacter sphaeroides, Rubrivivax gelatinosus, Bradyrhizobium ORS278). This regulator exhibits unique regulatory features in each bacterium: it can activate and/or repress the expression of photosynthesis genes, its activity can be modulated or not by the redox conditions, it can interact with other specific regulators and therefore be involved differently in light and/or redox regulatory circuits.

Published
2005

41. Light and Redox Control of Photosynthesis Gene Expression in Bradyrhizobium

Author

Joël Fardoux, David Pignol, Marianne Jaubert, Jérôme Lavergne, Sébastien Zappa, Jean-Marc Adriano, Sylvie Elsen, André Verméglio, Laure Hannibal, and Eric Giraud

Subjects
Regulation of gene expression, biology, Repressor, Cell Biology, Plasma protein binding, biology.organism_classification, Biochemistry, Bradyrhizobium, Biophysics, Histone octamer, Rhodopseudomonas palustris, Molecular Biology, Gene, Photosystem
Abstract

The two closely related bacteria Bradyrhizobium and Rhodopseudomonas palustris show an unusual mechanism of regulation of photosystem formation by light thanks to a bacteriophytochrome that antirepresses the regulator PpsR. In these two bacteria, we found out, unexpectedly, that two ppsR genes are present. We show that the two Bradyrhizobium PpsR proteins exert antagonistic effects in the regulation of photosystem formation with a classical repressor role for PpsR2 and an unexpected activator role for PpsR1. DNase I footprint analysis show that both PpsR bind to the same DNA TGTN12ACA motif that is present in tandem in the bchC promoter and the crtED intergenic region. Interestingly, the cycA and aerR promoter regions that contain only one conserved palindrome are recognized by PpsR2, but not PpsR1. Further biochemical analyses indicate that PpsR1 only is redox sensitive through the formation of an intermolecular disulfide bond, which changes its oligomerization state from a tetramer to an octamer under oxidizing conditions. Moreover, PpsR1 presents a higher DNA affinity under its reduced form in contrast to what has been previously found for PpsR or its homolog CrtJ from the Rhodobacter species. These results suggest that regulation of photosystem synthesis in Bradyrhizobium involves two PpsR competing for the binding to the same photosynthesis genes and this competition might be modulated by two factors: light via the antagonistic action of a bacteriophytochrome on PpsR2 and redox potential via the switch of PpsR1 oligomerization state.

Published
2004

42. Bacteriophytochrome and regulation of the synthesis of the photosynthetic apparatus in Rhodopseudomonas palustris: pitfalls of using laboratory strains

Author

Sébastien Zappa, Joël Fardoux, Eric Giraud, Laure Hannibal, André Verméglio, Jean-Marc Adriano, Bernard Genty, David Pignol, Pierre Bouyer, and Marianne Jaubert

Subjects
DNA, Bacterial, Molecular Sequence Data, Restriction Mapping, Repressor, Biology, medicine.disease_cause, Photosynthesis, Polymerase Chain Reaction, Bacterial Proteins, Sequence Homology, Nucleic Acid, Botany, medicine, Amino Acid Sequence, Bradyrhizobium, Cloning, Molecular, Physical and Theoretical Chemistry, DNA Primers, Photosystem, Mutation, Base Sequence, Sequence Homology, Amino Acid, Strain (chemistry), biology.organism_classification, Recombinant Proteins, Rhodopseudomonas, Spectrophotometry, Biophysics, Phytochrome, Rhodopseudomonas palustris, Sequence Alignment, Function (biology), Bacteria
Abstract

The synthesis of the photosynthetic apparatus of different strains of Rhodopseudomonas palustris has been studied as a function of the oxygen concentration and far-red light. For strain CEA001, only a small amount of photosynthetic apparatus is synthesized in the dark for oxygen concentration higher than 8% whereas synthesis is strongly enhanced by far-red light illumination. This enhancement is due to the action of a bacteriophytochrome (ORF2127/ORF2128), which antagonizes the repressor PpsR. On the contrary, a large fraction of photosystem is synthesized in the dark and far-red illumination induces no enhancement in strain CGA009. This difference in phenotype of strain CGA009 is explained by a single point-mutation R428C in the helix-turn-helix DNA binding motif of PpsR, rendering it inactive. In addition, a frame-shift mutation had occurred in the gene encoding bacteriophytochrome (ORF2127/ORF2128), conducting to a truncated inactive sensor. We propose that these mutations occurred in culture. Bacteria have developed a sophisticated regulatory process to synthesize their photosynthetic apparatus when light is available. This process is a critical advantage for the bacteria under natural conditions since they optimize their development depending on the available energy resources. On the contrary, under laboratory growth conditions where there is no substrate limitation, there is no crucial need for such a regulation and deleterious mutations affecting this process are of no importance.

Published
2004

43. The Crystal Structure of the Globular Head of Complement Protein C1q Provides a Basis for Its Versatile Recognition Properties

Author

Juan C. Fontecilla-Camps, Arnaud Gruez, David Pignol, Monique Lacroix, Denis Verger, Claudine Darnault, Jordi Juanhuix, Gérard J. Arlaud, and Christine Gaboriaud

Subjects
Models, Molecular, Stereochemistry, chemical and pharmacologic phenomena, Plasma protein binding, Crystallography, X-Ray, Ligands, Biochemistry, Domain (software engineering), Classical complement pathway, Protein structure, immune system diseases, Heterotrimeric G protein, Humans, skin and connective tissue diseases, Molecular Biology, Molecular Structure, Chemistry, Complement C1q, Cell Biology, Protein Structure, Tertiary, Complement (complexity), Complement system, Protein Subunits, C-Reactive Protein, Immunoglobulin G, C1q domain, Calcium, Crystallization, Protein Binding
Abstract

C1q is a versatile recognition protein that binds to an amazing variety of immune and non-immune ligands and triggers activation of the classical pathway of complement. The crystal structure of the C1q globular domain responsible for its recognition properties has now been solved and refined to 1.9 A of resolution. The structure reveals a compact, almost spherical heterotrimeric assembly held together mainly by non-polar interactions, with a Ca2+ ion bound at the top. The heterotrimeric assembly of the C1q globular domain appears to be a key factor of the versatile recognition properties of this protein. Plausible three-dimensional models of the C1q globular domain in complex with two of its physiological ligands, C-reactive protein and IgG, are proposed, highlighting two of the possible recognition modes of C1q. The C1q/human IgG1 model suggests a critical role for the hinge region of IgG and for the relative orientation of its Fab domain in C1q binding.

Published
2003

44. Structural and redox plasticity in the heterodimeric periplasmic nitrate reductase

Author

Pascal Arnoux, Monique Sabaty, David Pignol, Bruno Guigliarelli, Jean Alric, Jean-Marc Adriano, and Bettina Frangioni

Subjects
NAPA, chemistry.chemical_classification, biology, Stereochemistry, Rhodobacter sphaeroides, Periplasmic space, biology.organism_classification, Nitrate reductase, Nitrate Reductase, Protein Structure, Tertiary, Kinetics, chemistry.chemical_compound, Electron transfer, chemistry, Nitrate Reductases, Structural Biology, Oxidoreductase, Respiratory nitrate reductase, Periplasmic Proteins, Protein Structure, Quaternary, Dimerization, Oxidation-Reduction, Molecular Biology, Heme
Abstract

The structure of the respiratory nitrate reductase (NapAB) from Rhodobacter sphaeroides, the periplasmic heterodimeric enzyme responsible for the first step in the denitrification process, has been determined at a resolution of 3.2 A. The di-heme electron transfer small subunit NapB binds to the large subunit with heme II in close proximity to the [4Fe-4S] cluster of NapA. A total of 57 residues at the N- and C-terminal extremities of NapB adopt an extended conformation, embracing the NapA subunit and largely contributing to the total area of 5,900 A(2) buried in the complex. Complex formation was studied further by measuring the variation of the redox potentials of all the cofactors upon binding. The marked effects observed are interpreted in light of the three-dimensional structure and depict a plasticity that contributes to an efficient electron transfer in the complex from the heme I of NapB to the molybdenum catalytic site of NapA.

Published
2003

45. Environmental issues facing chemical, biological, radiological, and nuclear risks

Author

David Pignol, Marie-Thérèse Menager, Eric Ansoborlo, CEA/DSV, Commission des méthodes d'analyse du CEA (CETAMA), Département de recherche sur les procédés pour la mine et le recyclage du combustible (DMRC), CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Microbiologie Environnementale et Moléculaire (MEM), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Chemical Warfare Agents, 010405 organic chemistry, Health, Toxicology and Mutagenesis, media_common.quotation_subject, General Medicine, Scientific literature, 010402 general chemistry, 01 natural sciences, Pollution, 0104 chemical sciences, Procurement, 13. Climate action, Hazardous waste, Originality, Radiological weapon, [SDE]Environmental Sciences, Terrorism, Agency (sociology), Environmental Chemistry, Engineering ethics, ComputingMilieux_MISCELLANEOUS, media_common
Abstract

This special issue of Environmental Science and Pollution Research highlights selected papers presented at the first international conference entitled BCBRN, research and innovation^ (CBRN-RI in the following) which was held on March 16–18, 2015 at Antibes-Juan-les-Pins in France. This conference was co-organized by the French Alternative Energies and Atomic Energy Commission (CEA), the Armament procurement Agency (DGA), the Army Health Services, the Alpes-Maritimes Fire and Rescue Services (SDIS06), and Nice Conference Association. This special issue covers the topics of detection, remediation, and prevention of environments exposed to highly hazardous Chemical Biological Radiological and Nuclear (CBRN) agents following conflicts, natural catastrophes, industrial incidents, or terrorist acts. Worldwide, research, and industrial capacities have been undertaken to provide adapted answers to these new challenges. Besides existing conferences which give the opportunity for industrials to present their newly developed equipment, the aim of the CBRN-RI conference was to create a specific and unique communication space dedicated to research and innovation. It emerged as one of the best platforms for academic exchange and synergy with industrials of the field and provided a forum for researchers to address recent research results and to discuss various technical issues in the following themes: (i) pollution detection, (ii) human and environmental decontamination, (iii) protection, and (iv) therapeutics. Although investment of considerable financial, human, and scientific resources to develop medical countermeasures are developed to rapidly and effectively respond to a deliberate chemical, biological, radiological, or nuclear attack, tools for environment survey and protection are also now of growing interest in the field of NRBC-risks. This special issue contains articles that illustrate enzymatic decontamination processes with the case of organophosphates and environmental detection of toxic compounds as exemplified for toxic heavy metals. Studies focused on dedicated developments for efficient air sampling and detection of chemical warfare agents are also proposed. Original approaches used for the rapid identification of pathogenic bacteria or bacterial toxins in the field are also addressed. This Special Issue cannot hope to encompass the diversity and the originality of the research presented at CBRN-RI 2015; however, as editors, we hope that the Research Articles collected here will encourage environmental scientists to consider participating in the second CBRN-RI edition which is to be held in Lyon (France) from March 20 to 23, 2017. Finally, we sincerely thank all of the sponsors of the 2015 CBRN-RI conference. We also thank all the participants in this stimulating symposium, and in particular, the authors of the papers included in this Special Issue, which reflects the current international activities in the field. The guest editors of this Special Issue hope these papers are a valuable contribution to the scientific literature and useful to researchers and industrials worldwide involved in CBNR concerns. Dr. Philippe Garrigues, Editor-in-Chief of Environmental Science and Pollution Research, and his Editorial team are warmly thanked for their interest on this topic and handling of the review process. Responsible editor: Philippe Garrigues

Published
2015

46. Sulphur shuttling across a chaperone during molybdenum cofactor maturation

Author

Jérôme Lavergne, René Toci, Pascal Arnoux, Marina I. Siponen, Florian Bittner, Flore Oudouhou, Axel Magalon, Christian Ruppelt, David Pignol, Anne Walburger, Ralf R. Mendel, Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Microbiologie Environnementale et Moléculaire (MEM), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut de Microbiologie de la Méditerranée (IMM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)

Subjects
Models, Molecular, Formates, Dimer, Coenzymes, General Physics and Astronomy, Gene Expression, Crystallography, X-Ray, Protein Structure, Secondary, chemistry.chemical_compound, Cloning, Molecular, 0303 health sciences, Multidisciplinary, biology, 3. Good health, Carbon-Sulfur Lyases, Biochemistry, Molybdenum, [SDE]Environmental Sciences, Molybdenum cofactor, Oxidation-Reduction, Plasmids, Protein Binding, inorganic chemicals, Hydrogenase, Stereochemistry, chemistry.chemical_element, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Guanosine Diphosphate, General Biochemistry, Genetics and Molecular Biology, Cofactor, Carbon Cycle, 03 medical and health sciences, Multienzyme Complexes, Escherichia coli, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Binding site, 030304 developmental biology, [SDV.GEN]Life Sciences [q-bio]/Genetics, Binding Sites, 030306 microbiology, General Chemistry, Carbon Dioxide, Formate Dehydrogenases, Protein Structure, Tertiary, chemistry, Chaperone (protein), biology.protein, Biocatalysis, Protein Multimerization, Sulfur, Cysteine, Molecular Chaperones
Abstract

International audience; Formate dehydrogenases (FDHs) are of interest as they are natural catalysts that sequester atmospheric CO2, generating reduced carbon compounds with possible uses as fuel. FDHs activity in Escherichia coli strictly requires the sulphurtransferase EcFdhD, which likely transfers sulphur from IscS to the molybdenum cofactor (Mo-bisPGD) of FDHs. Here we show that EcFdhD binds Mo-bisPGD in vivo and has submicromolar affinity for GDP-used as a surrogate of the molybdenum cofactor's nucleotide moieties. The crystal structure of EcFdhD in complex with GDP shows two symmetrical binding sites located on the same face of the dimer. These binding sites are connected via a tunnel-like cavity to the opposite face of the dimer where two dynamic loops, each harbouring two functionally important cysteine residues, are present. On the basis of structure-guided mutagenesis, we propose a model for the sulphuration mechanism of Mo-bisPGD where the sulphur atom shuttles across the chaperone dimer.

Published
2014

47. Interplay between two bacterial actin homologs, MamK and MamK-Like, is required for the alignment of magnetosome organelles in Magnetospirillum magneticum AMB-1

Author

Ertan Ozyamak, Nicolas Ginet, Nicole Abreu, Soumaya Mannoubi, David Pignol, and Arash Komeili

Subjects
Magnetotactic bacteria, 1.1 Normal biological development and functioning, Magnetosome, Molecular Sequence Data, medicine.disease_cause, Medical and Health Sciences, Microbiology, Bacterial Proteins, Underpinning research, Organelle, medicine, Genetics, Amino Acid Sequence, Magnetospirillum, Molecular Biology, Peptide sequence, Actin, Adenosine Triphosphatases, Mutation, biology, Agricultural and Veterinary Sciences, Bacterial, Articles, Gene Expression Regulation, Bacterial, Biological Sciences, biology.organism_classification, Actins, Cell biology, Gene Expression Regulation, Magnetosomes, Function (biology), Biotechnology
Abstract

Many bacterial species contain multiple actin-like proteins tasked with the execution of crucial cell biological functions. MamK, an actin-like protein found in magnetotactic bacteria, is important in organizing magnetosome organelles into chains that are used for navigation along geomagnetic fields. MamK and numerous other magnetosome formation factors are encoded by a genetic island termed the magnetosome island. Unlike most magnetotactic bacteria, Magnetospirillum magneticum AMB-1 (AMB-1) contains a second island of magnetosome-related genes that was named the magnetosome islet. A homologous copy of mamK , mamK-like , resides within this islet and encodes a protein capable of filament formation in vitro . Previous work had shown that mamK-like is expressed in vivo , but its function, if any, had remained unknown. Though MamK-like is highly similar to MamK, it contains a mutation that in MamK and other actins blocks ATPase activity in vitro and filament dynamics in vivo . Here, using genetic analysis, we demonstrate that mamK-like has an in vivo role in assisting organelle alignment. In addition, MamK-like forms filaments in vivo in a manner that is dependent on the presence of MamK and the two proteins interact in a yeast two-hybrid assay. Surprisingly, despite the ATPase active-site mutation, MamK-like is capable of ATP hydrolysis in vitro and promotes MamK filament turnover in vivo . Taken together, these experiments suggest that direct interactions between MamK and MamK-like contribute to magnetosome alignment in AMB-1.

Published
2014

48. Radiation response in Deinococcus deserti: IrrE is a metalloprotease that cleaves repressor protein DdrO

Author

Monika, Ludanyi, Laurence, Blanchard, Rémi, Dulermo, Géraldine, Brandelet, Laurent, Bellanger, David, Pignol, David, Lemaire, and Arjan, de Groot

Subjects
Repressor Proteins, Stress, Physiological, Molecular Sequence Data, Proteolysis, Escherichia coli, Metalloproteases, Gene Expression, Amino Acid Sequence, Deinococcus, Gene Expression Regulation, Bacterial, Sequence Alignment
Abstract

Deinococcus bacteria are famous for their extreme radiation tolerance. The IrrE protein was shown to be essential for radiation tolerance and, in an unelucidated manner, for induction of a number of genes in response to radiation, including recA and other DNA repair genes. Earlier studies indicated that IrrE could be a zinc peptidase, but proteolytic activity was not demonstrated. Here, using several in vivo and in vitro experiments, IrrE from Deinococcus deserti was found to interact with DdrO, a predicted regulator encoded by a radiation-induced gene that is, like irrE, highly conserved in Deinococcus. Moreover, IrrE was found to cleave DdrO in vitro and when the proteins were coexpressed in Escherichia coli. This cleavage was not observed in the presence of metal chelator EDTA or when IrrE contains a mutation in the conserved active-site motif of metallopeptidases. In D. deserti, IrrE-dependent cleavage of DdrO was observed after exposure to radiation. Furthermore, DdrO-dependent repression of the promoter of a radiation-induced gene was shown. These results demonstrate that IrrE is a metalloprotease and we propose that IrrE-mediated cleavage inactivates repressor protein DdrO, leading to transcriptional induction of various genes required for repair and survival after exposure of Deinococcus to radiation.

Published
2014

49. RNA Sequencing and Proteogenomics Reveal the Importance of Leaderless mRNAs in the Radiation-Tolerant Bacterium Deinococcus deserti

Author

Jean Armengaud, Bernard Fernandez, Stéphane Cruveiller, David Roche, David Pignol, Monika Ludanyi, Laurence Blanchard, David Vallenet, Arjan de Groot, Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-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), Microbiologie Environnementale et Moléculaire (MEM), Génomique métabolique (UMR 8030), Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-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 Paris-Saclay-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 d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biochimie des Systèmes Perturbés (LBSP), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoire Innovations technologiques pour la Détection et le Diagnostic (LI2D), Service de Pharmacologie et Immunoanalyse (SPI), Médicaments et Technologies pour la Santé (MTS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-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 National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Médicaments et Technologies pour la Santé (MTS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), ANR-07-BLAN-0106,DEINOCOCCUS,Structure of the nucleoïd and radioresistance of Deinococcus radiodurans and Deinococcus deserti(2007), 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)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), 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)-Université d'Évry-Val-d'Essonne (UEVE)

Subjects
Proteomics, desiccation tolerance, protein protection, [SDV]Life Sciences [q-bio], Molecular Sequence Data, Biology, genome evolution, medicine.disease_cause, Radiation Tolerance, transcription start sites, Transcriptome, Evolution, Molecular, Eukaryotic translation, Start codon, Bacterial Proteins, Transcription (biology), Genetics, medicine, Deinococcus, small peptides, RNA, Messenger, Gene, Ecology, Evolution, Behavior and Systematics, protein translation initiation, ComputingMilieux_MISCELLANEOUS, Base Sequence, Sequence Analysis, RNA, Deinococcus deserti, Proteogenomics, biology.organism_classification, Adaptation, Physiological, RNA, Bacterial, Research Article
Abstract

Deinococcus deserti is a desiccation- and radiation-tolerant desert bacterium. Differential RNA sequencing (RNA-seq) was performed to explore the specificities of its transcriptome. Strikingly, for 1,174 (60%) mRNAs, the transcription start site was found exactly at (916 cases, 47%) or very close to the translation initiation codon AUG or GUG. Such proportion of leaderless mRNAs, which may resemble ancestral mRNAs, is unprecedented for a bacterial species. Proteomics showed that leaderless mRNAs are efficiently translated in D. deserti. Interestingly, we also found 173 additional transcripts with a 5'-AUG or 5'-GUG that would make them competent for ribosome binding and translation into novel small polypeptides. Fourteen of these are predicted to be leader peptides involved in transcription attenuation. Another 30 correlated with new gene predictions and/or showed conservation with annotated and nonannotated genes in other Deinococcus species, and five of these novel polypeptides were indeed detected by mass spectrometry. The data also allowed reannotation of the start codon position of 257 genes, including several DNA repair genes. Moreover, several novel highly radiation-induced genes were found, and their potential roles are discussed. On the basis of our RNA-seq and proteogenomics data, we propose that translation of many of the novel leaderless transcripts, which may have resulted from single-nucleotide changes and maintained by selective pressure, provides a new explanation for the generation of a cellular pool of small peptides important for protection of proteins against oxidation and thus for radiation/desiccation tolerance and adaptation to harsh environmental conditions.

Published
2014

50. The oxygen sensor MgFnr controls magnetite biomineralization by regulation of denitrification in Magnetospirillum gryphiswaldense

Author

Yingjie, Li, Monique, Sabaty, Sarah, Borg, Karen T, Silva, David, Pignol, and Dirk, Schüler

Subjects
Magnetite biomineralization, Denitrification, Oxygen regulation, Anaerobiosis, Gene Expression Regulation, Bacterial, Magnetospirillum, Magnetospirillum gryphiswaldense, Aerobiosis, Ferrosoferric Oxide, Transcription Factors, Research Article, Oxygen sensor Fnr
Abstract

Background Magnetotactic bacteria are capable of synthesizing magnetosomes only under oxygen-limited conditions. However, the mechanism of the aerobic repression on magnetite biomineralization has remained unknown. In Escherichia coli and other bacteria, Fnr (fumarate and nitrate reduction regulator) proteins are known to be involved in controlling the switch between microaerobic and aerobic metabolism. Here, we report on an Fnr-like protein (MgFnr) and its role in growth metabolism and magnetite biomineralization in the alphaproteobacterium Magnetospirillum gryphiswaldense. Results Deletion of Mgfnr not only resulted in decreased N2 production due to reduced N2O reductase activity, but also impaired magnetite biomineralization under microaerobic conditions in the presence of nitrate. Overexpression of MgFnr in the WT also caused the synthesis of smaller magnetite particles under anaerobic and microaerobic conditions in the presence of nitrate. These data suggest that proper expression of MgFnr is required for WT-like magnetosome synthesis, which is regulated by oxygen. Analyses of transcriptional gusA reporter fusions revealed that besides showing similar properties to Fnr proteins reported in other bacteria, MgFnr is involved in the repression of the expression of denitrification genes nor and nosZ under aerobic conditions, possibly owing to several unique amino acid residues specific to MTB-Fnr. Conclusions We have identified and thoroughly characterized the first regulatory protein mediating denitrification growth and magnetite biomineralization in response to different oxygen conditions in a magnetotactic bacterium. Our findings reveal that the global oxygen regulator MgFnr is a genuine O2 sensor. It is involved in controlling expression of denitrification genes and thereby plays an indirect role in maintaining proper redox conditions required for magnetite biomineralization.

Published
2014

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