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- EMDB-10745: Leishmania RNA virus capsid with exogenous mRNA -

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Basic information

Entry
Database: EMDB / ID: EMD-10745
TitleLeishmania RNA virus capsid with exogenous mRNA
Map data
Sample
  • Virus: Leishmania RNA virus 1 - LgM5313
Biological speciesLeishmania RNA virus 1 - LgM5313
Methodsingle particle reconstruction / cryo EM / Resolution: 3.76 Å
AuthorsProchazkova M
Funding support Czech Republic, 1 items
OrganizationGrant numberCountry
European Research Council (ERC) Czech Republic
CitationJournal: J Virol / Year: 2021
Title: Capsid Structure of RNA Virus 1.
Authors: Michaela Procházková / Tibor Füzik / Danyil Grybchuk / Francesco Luca Falginella / Lucie Podešvová / Vyacheslav Yurchenko / Robert Vácha / Pavel Plevka /
Abstract: parasites cause a variety of symptoms, including mucocutaneous leishmaniasis, which results in the destruction of the mucous membranes of the nose, mouth, and throat. The species of carrying RNA ... parasites cause a variety of symptoms, including mucocutaneous leishmaniasis, which results in the destruction of the mucous membranes of the nose, mouth, and throat. The species of carrying RNA virus 1 (LRV1), from the family , are more likely to cause severe disease and are less sensitive to treatment than those that do not contain the virus. Although the importance of LRV1 for the severity of leishmaniasis was discovered a long time ago, the structure of the virus remained unknown. Here, we present a cryo-electron microscopy reconstruction of the virus-like particle of LRV1 determined to a resolution of 3.65 Å. The capsid has icosahedral symmetry and is formed by 120 copies of a capsid protein assembled in asymmetric dimers. RNA genomes of viruses from the family are synthetized, but not capped at the 5' end, by virus RNA polymerases. To protect viral RNAs from degradation, capsid proteins of the L-A totivirus cleave the 5' caps of host mRNAs, creating decoys to overload the cellular RNA quality control system. Capsid proteins of LRV1 form positively charged clefts, which may be the cleavage sites for the 5' cap of mRNAs. The putative RNA binding site of LRV1 is distinct from that of the related L-A virus. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative decapping site. Such inhibitors may be developed into a treatment for mucocutaneous leishmaniasis caused by LRV1-positive species of Twelve million people worldwide suffer from leishmaniasis, resulting in more than 30 thousand deaths annually. The disease has several variants that differ in their symptoms. The mucocutaneous form, which leads to disintegration of the nasal septum, lips, and palate, is caused predominantly by parasites carrying RNA virus 1 (LRV1). Here, we present the structure of the LRV1 capsid determined using cryo-electron microscopy. Capsid proteins of a related totivirus, L-A virus, protect viral RNAs from degradation by cleaving the 5' caps of host mRNAs. Capsid proteins of LRV1 may have the same function. We show that the LRV1 capsid contains positively charged clefts that may be sites for the cleavage of mRNAs of cells. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative mRNA cleavage site. Such inhibitors may be used as treatments for mucocutaneous leishmaniasis.
History
DepositionMar 11, 2020-
Header (metadata) releaseNov 11, 2020-
Map releaseNov 11, 2020-
UpdateApr 14, 2021-
Current statusApr 14, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10745.png

Downloads & links

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Map

FileDownload / File: emd_10745.map.gz / Format: CCP4 / Size: 620.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.063 Å
Density
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.12939958 - 0.22170044
Average (Standard dev.)0.0015629168 (±0.012472558)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-273-273-273
Dimensions546546546
Spacing546546546
CellA=B=C: 580.398 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0631.0631.063
M x/y/z546546546
origin x/y/z0.0000.0000.000
length x/y/z580.398580.398580.398
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-273-273-273
NC/NR/NS546546546
D min/max/mean-0.1290.2220.002

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Supplemental data

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Sample components

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Entire : Leishmania RNA virus 1 - LgM5313

EntireName: Leishmania RNA virus 1 - LgM5313
Components
  • Virus: Leishmania RNA virus 1 - LgM5313

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Supramolecule #1: Leishmania RNA virus 1 - LgM5313

SupramoleculeName: Leishmania RNA virus 1 - LgM5313 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Assembled particles mixed with isolated Leishmania mRNA
NCBI-ID: 1285422 / Sci species name: Leishmania RNA virus 1 - LgM5313 / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: OTHER / Virus enveloped: No / Virus empty: Yes
Host (natural)Organism: Leishmania guyanensis (eukaryote) / Strain: MHOM/BR/75/M4147
Host systemOrganism: Escherichia coli BL21(DE3) (bacteria)
Virus shellShell ID: 1 / Name: capsid / Diameter: 422.0 Å / T number (triangulation number): 2

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration15 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
300.0 mMNaClSodium chloridesodium chloride
50.0 mMTris
1.0 mMEDTAEthylenediaminetetraacetic acid
5.0 mMB-mebetamercaptoethanol

Details: Supplemented with isolated mRNA from Leishmania sp.
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 70 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK II / Details: blot force 3 blot time 5 wait time 10.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 0.0005 µm / Nominal defocus min: -0.0005 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 8000 / Average exposure time: 0.5 sec. / Average electron dose: 21.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 16715
CTF correctionSoftware - Name: CTFFIND (ver. 4)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1m1c


Details: Initial model constructed.
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 2.1)
Final 3D classificationNumber classes: 3 / Avg.num./class: 7000 / Software - Name: RELION
Final angle assignmentType: PROJECTION MATCHING
Projection matching processing - Angular sampling: 1.8 degrees
Software - Name: RELION
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 3.76 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 9932
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

1m1c


Chain - Chain ID: A
DetailsPDB ID: 6Y83 fits the map
RefinementSpace: RECIPROCAL / Protocol: RIGID BODY FIT / Overall B value: 174 / Target criteria: R-factor

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