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- PDB-8uk3: The rotavirus VP5*/VP8* conformational transition permeabilizes m... -

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Basic information

Entry
Database: PDB / ID: 8uk3
TitleThe rotavirus VP5*/VP8* conformational transition permeabilizes membranes to Ca2+ (class 6 reconstruction)
Components
  • Outer capsid glycoprotein VP7
  • Outer capsid protein VP4
KeywordsVIRAL PROTEIN / Non-enveloped virus / viral particle / entry / membrane-penetration / rotavirus / VP4 / VP5* / VP8* / calcium / Ca2+ / liposome
Function / homology
Function and homology information


host cell endoplasmic reticulum lumen / T=13 icosahedral viral capsid / host cell rough endoplasmic reticulum / host cytoskeleton / viral outer capsid / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / host cell endoplasmic reticulum-Golgi intermediate compartment / receptor-mediated virion attachment to host cell / virion attachment to host cell ...host cell endoplasmic reticulum lumen / T=13 icosahedral viral capsid / host cell rough endoplasmic reticulum / host cytoskeleton / viral outer capsid / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / host cell endoplasmic reticulum-Golgi intermediate compartment / receptor-mediated virion attachment to host cell / virion attachment to host cell / host cell plasma membrane / membrane / metal ion binding
Similarity search - Function
Glycoprotein VP7 / Glycoprotein VP7, domain 1 / Glycoprotein VP7, domain 2 / Glycoprotein VP7 / Haemagglutinin outer capsid protein VP4, concanavalin-like domain / Outer Capsid protein VP4 (Hemagglutinin) Concanavalin-like domain / Rotavirus VP4 helical domain / Rotavirus VP4 helical domain / Outer capsid protein VP4 / Rotavirus VP4, membrane interaction domain superfamily ...Glycoprotein VP7 / Glycoprotein VP7, domain 1 / Glycoprotein VP7, domain 2 / Glycoprotein VP7 / Haemagglutinin outer capsid protein VP4, concanavalin-like domain / Outer Capsid protein VP4 (Hemagglutinin) Concanavalin-like domain / Rotavirus VP4 helical domain / Rotavirus VP4 helical domain / Outer capsid protein VP4 / Rotavirus VP4, membrane interaction domain superfamily / Rotavirus VP4, membrane interaction domain / Rotavirus VP4 membrane interaction domain / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Outer capsid protein VP4 / Outer capsid glycoprotein VP7
Similarity search - Component
Biological speciesSimian rotavirus A strain RRV
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 8 Å
AuthorsDe Sautu, M. / Herrmann, T. / Jenni, S. / Harrison, S.C.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)CA13202 United States
CitationJournal: PLoS Pathog / Year: 2024
Title: The rotavirus VP5*/VP8* conformational transition permeabilizes membranes to Ca2.
Authors: Marilina de Sautu / Tobias Herrmann / Gustavo Scanavachi / Simon Jenni / Stephen C Harrison /
Abstract: Rotaviruses infect cells by delivering into the cytosol a transcriptionally active inner capsid particle (a "double-layer particle": DLP). Delivery is the function of a third, outer layer, which ...Rotaviruses infect cells by delivering into the cytosol a transcriptionally active inner capsid particle (a "double-layer particle": DLP). Delivery is the function of a third, outer layer, which drives uptake from the cell surface into small vesicles from which the DLPs escape. In published work, we followed stages of rhesus rotavirus (RRV) entry by live-cell imaging and correlated them with structures from cryogenic electron microscopy and tomography (cryo-EM and cryo-ET). The virus appears to wrap itself in membrane, leading to complete engulfment and loss of Ca2+ from the vesicle produced by the wrapping. One of the outer-layer proteins, VP7, is a Ca2+-stabilized trimer; loss of Ca2+ releases both VP7 and the other outer-layer protein, VP4, from the particle. VP4, activated by cleavage into VP8* and VP5*, is a trimer that undergoes a large-scale conformational rearrangement, reminiscent of the transition that viral fusion proteins undergo to penetrate a membrane. The rearrangement of VP5* thrusts a 250-residue, C-terminal segment of each of the three subunits outward, while allowing the protein to remain attached to the virus particle and to the cell being infected. We proposed that this segment inserts into the membrane of the target cell, enabling Ca2+ to cross. In the work reported here, we show the validity of key aspects of this proposed sequence. By cryo-EM studies of liposome-attached virions ("triple-layer particles": TLPs) and single-particle fluorescence imaging of liposome-attached TLPs, we confirm insertion of the VP4 C-terminal segment into the membrane and ensuing generation of a Ca2+ "leak". The results allow us to formulate a molecular description of early events in entry. We also discuss our observations in the context of other work on double-strand RNA virus entry.
History
DepositionOct 12, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 27, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 17, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
1: Outer capsid protein VP4
f: Outer capsid glycoprotein VP7
2: Outer capsid protein VP4
g: Outer capsid glycoprotein VP7
3: Outer capsid protein VP4
h: Outer capsid glycoprotein VP7
i: Outer capsid glycoprotein VP7
j: Outer capsid glycoprotein VP7
k: Outer capsid glycoprotein VP7
l: Outer capsid glycoprotein VP7
m: Outer capsid glycoprotein VP7
n: Outer capsid glycoprotein VP7
o: Outer capsid glycoprotein VP7
p: Outer capsid glycoprotein VP7
q: Outer capsid glycoprotein VP7
r: Outer capsid glycoprotein VP7
a: Outer capsid glycoprotein VP7
b: Outer capsid glycoprotein VP7
c: Outer capsid glycoprotein VP7
d: Outer capsid glycoprotein VP7
e: Outer capsid glycoprotein VP7
hetero molecules


Theoretical massNumber of molelcules
Total (without water)934,57093
Polymers928,42421
Non-polymers6,14672
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Outer capsid protein VP4 / Hemagglutinin


Mass: 86655.586 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Simian rotavirus A strain RRV / Cell line (production host): MA104 / Production host: Chlorocebus aethiops (grivet) / References: UniProt: G0YZG6
#2: Protein
Outer capsid glycoprotein VP7


Mass: 37136.531 Da / Num. of mol.: 18
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Simian rotavirus A strain RRV / Cell line (production host): MA104 / Production host: Chlorocebus aethiops (grivet) / References: UniProt: P12476
#3: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 18 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#4: Chemical...
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 54 / Source method: obtained synthetically / Formula: Ca
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Simian rotavirus A strain RRV / Type: VIRUS / Entity ID: #1-#2 / Source: NATURAL
Source (natural)Organism: Simian rotavirus A strain RRV
Details of virusEmpty: NO / Enveloped: NO / Isolate: STRAIN / Type: VIRION
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 8 Å / Resolution method: OTHER / Num. of particles: 70018 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingPDB-ID: 6WXG

6wxg


Accession code: 6WXG / Source name: PDB / Type: experimental model

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