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Start Over Author "Benoît Arragain" Topic cryoelectron microscopy
3 results on '"Benoît Arragain"'

1. Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes

Author

Stephen Cusack, Benoît Arragain, Guy Schoehn, Piotr Gerlach, Grégory Effantin, Hélène Malet, Juan Reguera, Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), European Molecular Biology Laboratory [Heidelberg] (EMBL), European Molecular Biology Laboratory [Grenoble] (EMBL), EMBL Eukaryotic Expression facility, IBS Electron Miroscopy Platform, ANR-19-CE11-0024,HiPathBunya,Caractérisation des machineries de réplication de Bunyavirus hautement pathogènes par une approche de biologie structurale intégrée(2019), Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Architecture et fonction des macromolécules biologiques (AFMB), and Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Aix Marseille Université (AMU)

Subjects
0301 basic medicine, La Crosse virus, Transcription, Genetic, Protein Conformation, Science, [SDV]Life Sciences [q-bio], Arenaviridae, viruses, 030106 microbiology, General Physics and Astronomy, Crystallography, X-Ray, Genome, Article, General Biochemistry, Genetics and Molecular Biology, Viral Proteins, 03 medical and health sciences, chemistry.chemical_compound, Protein Domains, Cryoelectron microscopy, Transcription (biology), RNA polymerase, lcsh:Science, ComputingMilieux_MISCELLANEOUS, Polymerase, X-ray crystallography, 030304 developmental biology, 0303 health sciences, Multidisciplinary, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, 030306 microbiology, Chemistry, RNA, General Chemistry, RNA-Dependent RNA Polymerase, 3. Good health, Cell biology, 030104 developmental biology, Duplex (building), biology.protein, lcsh:Q, Linker
Abstract

Bunyavirales is an order of segmented negative-strand RNA viruses comprising several life-threatening pathogens against which no effective treatment is currently available. Replication and transcription of the RNA genome constitute essential processes performed by the virally encoded multi-domain RNA-dependent RNA polymerase. Here, we describe the complete high-resolution cryo-EM structure of La Crosse virus polymerase. It reveals the presence of key protruding C-terminal domains, notably the cap-binding domain, which undergoes large movements related to its role in transcription initiation, and a zinc-binding domain that displays a fold not previously observed. We capture the polymerase structure at pre-initiation and elongation states, uncovering the coordinated movement of the priming loop, mid-thumb ring linker and lid domain required for the establishment of a ten-base-pair template-product RNA duplex before strand separation into respective exit tunnels. These structural details and the observed dynamics of key functional elements will be instrumental for structure-based development of polymerase inhibitors., RNA-dependent RNA polymerases from segmented negative stranded RNA viruses catalyze genome replication and viral transcription. Here, the authors present the cryo-EM structure of full-length La Crosse virus polymerase and structurally characterize the pre-initiation and elongation states, which is of interest for the development of polymerase inhibitors.

Published
2020
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2. Structural snapshots of La Crosse virus polymerase reveal the mechanisms underlying $Peribunyaviridae$ replication and transcription

Author

Benoît Arragain, Quentin Durieux Trouilleton, Florence Baudin, Jan Provaznik, Nayara Azevedo, Stephen Cusack, Guy Schoehn, Hélène Malet, Groupe Imagerie microscopique d'assemblages complexes / Microscopic Imaging of complex Assemblies group (IBS MICA), Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), EMBL Heidelberg, European Molecular Biology Laboratory [Grenoble] (EMBL), Institut Universitaire de France (IUF), Ministère de l'Education nationale, de l’Enseignement supérieur et de la Recherche (M.E.N.E.S.R.), EMBL Eukaryotic Expression facility, IBS Electron Miroscopy Platform, EMBL GeneCore, Institut Universitaire de France, ANR-19-CE11-0024,HiPathBunya,Caractérisation des machineries de réplication de Bunyavirus hautement pathogènes par une approche de biologie structurale intégrée(2019), and European Project: FDT202012010396

Subjects
Transcription, Genetic, Protein Conformation, General Physics and Astronomy, Genome, Viral, Virus Replication, General Biochemistry, Genetics and Molecular Biology, Cell Line, MESH: Protein Conformation, La Crosse virus, MESH: Sequence Analysis, RNA, Humans, Multidisciplinary, MESH: Humans, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Sequence Analysis, RNA, MESH: Transcription, Genetic, Cryoelectron Microscopy, MESH: Virus Replication, General Chemistry, RNA-Dependent RNA Polymerase, MESH: Cell Line, HEK293 Cells, MESH: La Crosse virus, MESH: RNA, Viral, MESH: RNA-Dependent RNA Polymerase, MESH: HEK293 Cells, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, RNA, Viral, MESH: Cryoelectron Microscopy, MESH: Genome, Viral
Abstract

Segmented negative-strand RNA bunyaviruses encode a multi-functional polymerase that performs genome replication and transcription. Here, we establish conditions for in vitro activity of La Crosse virus polymerase and visualize its conformational dynamics by cryo-electron microscopy, unveiling the precise molecular mechanics underlying its essential activities. We find that replication initiation is coupled to distal duplex promoter formation, endonuclease movement, prime-and-realign loop extension and closure of the polymerase core that direct the template towards the active site. Transcription initiation depends on C-terminal region closure and endonuclease movements that prompt primer cleavage prior to primer entry in the active site. Product realignment after priming, observed in replication and transcription, is triggered by the prime-and-realign loop. Switch to elongation results in polymerase reorganization and core region opening to facilitate template-product duplex formation in the active site cavity. The uncovered detailed mechanics should be helpful for the future design of antivirals counteracting bunyaviral life threatening pathogens.

Published
2022
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3. High resolution cryo-EM structure of the helical RNA-bound Hantaan virus nucleocapsid reveals its assembly mechanisms

Author

Benoît Arragain, Juan Reguera, Ambroise Desfosses, Irina Gutsche, Guy Schoehn, Hélène Malet, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Architecture et fonction des macromolécules biologiques (AFMB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), IDEX IRS grant University of Grenoble (G7H-IRS17H50), Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
replication, Insecta, QH301-705.5, Protein Conformation, Science, viruses, Structural Biology and Molecular Biophysics, Short Report, Genome, Viral, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, 0302 clinical medicine, molecular biophysics, structural biology, Animals, Humans, Biology (General), Nucleocapsid, 030304 developmental biology, nucleoprotein, Bunyavirales, 0303 health sciences, General Immunology and Microbiology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], General Neuroscience, Virus Assembly, Cryoelectron Microscopy, General Medicine, 3. Good health, Hantaan virus, Virus, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], HEK293 Cells, Medicine, RNA, Viral, Hantaan, 030217 neurology & neurosurgery, Hantavirus
Abstract

[u'Rats and mice sometimes transmit hantaviruses, a family of microbes that can cause deadly human diseases. For example, the Hantaan virus leads to haemorrhagic fevers that are potentially fatal. There are no vaccine or even drugs against these infections.', u'To multiply, viruses must insert their genetic material inside a cell. While the body often detects and destroys foreign genetic information, hantaviruses can still evade our defences. Molecules called nucleoproteins bind to the viral genome, hiding it away in long helices called nucleocapsids. When the virus needs to replicate, an enzyme opens up the nucleocapsid, reads and copies the genetic code, and then closes the helix. Yet, researchers know little about the details of this process, or even the structure of the nucleocapsid.', u'Here, Arragain et al. use a method called cryo-electron microscopy to examine and piece together the exact 3D structure of the Hantaan virus nucleocapsid. This was possible because the new technique allows scientists to observe biological molecules at an unprecedented, near atomic resolution. The resulting model reveals that the viral genome nests into a groove inside the nucleocapsid. It also shows that specific interactions between nucleoproteins stabilise the helix. Finally, the model helps to provide hypotheses on how the enzyme could read the genome without breaking the capsid.', u'Mapping out the structure and the interactions of the nucleocapsid is the first step towards finding molecules that could destabilise the helix and neutralise the virus: this could help fight both the Hantaan virus and other members of its deadly family.'], Negative-strand RNA viruses condense their genome into helical nucleocapsids that constitute essential templates for viral replication and transcription. The intrinsic flexibility of nucleocapsids usually prevents their full-length structural characterisation at high resolution. Here, we describe purification of full-length recombinant metastable helical nucleocapsid of Hantaan virus (Hantaviridae family, Bunyavirales order) and determine its structure at 3.3 Å resolution by cryo-electron microscopy. The structure reveals the mechanisms of helical multimerisation via sub-domain exchanges between protomers and highlights nucleotide positions in a continuous positively charged groove compatible with viral genome binding. It uncovers key sites for future structure-based design of antivirals that are currently lacking to counteract life-threatening hantavirus infections. The structure also suggests a model of nucleoprotein-polymerase interaction that would enable replication and transcription solely upon local disruption of the nucleocapsid.

Published
2018
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