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- EMDB-5948: Three-dimensional structure of a protozoal dsRNA virus that infec... -

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Entry
Database: EMDB / ID: EMD-5948
TitleThree-dimensional structure of a protozoal dsRNA virus that infects enteric pathogen Giardia lamblia
Map dataIcosahedral reconstruction of giardiavirus
Sample
  • Sample: GLV virion
  • Virus: Giardia lamblia virus
KeywordsGiardia lamblia virus / protozoal virus / dsRNA virus / icosahedral image reconstruction / cryoTEM / infects Giardia lamblia
Biological speciesGiardia lamblia virus
Methodsingle particle reconstruction / cryo EM / Resolution: 6.0 Å
AuthorsJanssen ME / Takagi Y / Parent KN / Cardone G / Fichorova RN / Nibert ML / Baker TS
CitationJournal: J Virol / Year: 2015
Title: Three-dimensional structure of a protozoal double-stranded RNA virus that infects the enteric pathogen Giardia lamblia.
Authors: Mandy E W Janssen / Yuko Takagi / Kristin N Parent / Giovanni Cardone / Max L Nibert / Timothy S Baker /
Abstract: Giardia lamblia virus (GLV) is a small, nonenveloped, nonsegmented double-stranded RNA (dsRNA) virus infecting Giardia lamblia, the most common protozoan pathogen of the human intestine and a major ...Giardia lamblia virus (GLV) is a small, nonenveloped, nonsegmented double-stranded RNA (dsRNA) virus infecting Giardia lamblia, the most common protozoan pathogen of the human intestine and a major agent of waterborne diarrheal disease worldwide. GLV (genus Giardiavirus) is a member of family Totiviridae, along with several other groups of protozoal or fungal viruses, including Leishmania RNA viruses and Trichomonas vaginalis viruses. Interestingly, GLV is more closely related than other Totiviridae members to a group of recently discovered metazoan viruses that includes penaeid shrimp infectious myonecrosis virus (IMNV). Moreover, GLV is the only known protozoal dsRNA virus that can transmit efficiently by extracellular means, also like IMNV. In this study, we used transmission electron cryomicroscopy and icosahedral image reconstruction to examine the GLV virion at an estimated resolution of 6.0 Å. Its outermost diameter is 485 Å, making it the largest totivirus capsid analyzed to date. Structural comparisons of GLV and other totiviruses highlighted a related "T=2" capsid organization and a conserved helix-rich fold in the capsid subunits. In agreement with its unique capacity as a protozoal dsRNA virus to survive and transmit through extracellular environments, GLV was found to be more thermoresistant than Trichomonas vaginalis virus 1, but no specific protein machinery to mediate cell entry, such as the fiber complexes in IMNV, could be localized. These and other structural and biochemical findings provide a basis for future work to dissect the cell entry mechanism of GLV into a "primitive" (early-branching) eukaryotic host and an important enteric pathogen of humans.
IMPORTANCE: Numerous pathogenic bacteria, including Corynebacterium diphtheriae, Salmonella enterica, and Vibrio cholerae, are infected with lysogenic bacteriophages that contribute significantly to ...IMPORTANCE: Numerous pathogenic bacteria, including Corynebacterium diphtheriae, Salmonella enterica, and Vibrio cholerae, are infected with lysogenic bacteriophages that contribute significantly to bacterial virulence. In line with this phenomenon, several pathogenic protozoa, including Giardia lamblia, Leishmania species, and Trichomonas vaginalis are persistently infected with dsRNA viruses, and growing evidence indicates that at least some of these protozoal viruses can likewise enhance the pathogenicity of their hosts. Understanding of these protozoal viruses, however, lags far behind that of many bacteriophages. Here, we investigated the dsRNA virus that infects the widespread enteric parasite Giardia lamblia. Using electron cryomicroscopy and icosahedral image reconstruction, we determined the virion structure of Giardia lamblia virus, obtaining new information relating to its assembly, stability, functions in cell entry and transcription, and similarities and differences with other dsRNA viruses. The results of our study set the stage for further mechanistic work on the roles of these viruses in protozoal virulence.
History
DepositionApr 16, 2014-
Header (metadata) releaseApr 30, 2014-
Map releaseNov 12, 2014-
UpdateMay 27, 2015-
Current statusMay 27, 2015Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.1
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 1.1
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 1.1
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_5948.map.gz / Format: CCP4 / Size: 808.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationIcosahedral reconstruction of giardiavirus
Voxel sizeX=Y=Z: 1.09 Å
Density
Contour LevelBy AUTHOR: 1.1 / Movie #1: 1.1
Minimum - Maximum-3.38433194 - 6.48850393
Average (Standard dev.)0.05024409 (±0.66671234)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions601601601
Spacing601601601
CellA=B=C: 655.09 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.091.091.09
M x/y/z601601601
origin x/y/z0.0000.0000.000
length x/y/z655.090655.090655.090
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS601601601
D min/max/mean-3.3846.4890.050

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Supplemental data

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Sample components

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Entire : GLV virion

EntireName: GLV virion
Components
  • Sample: GLV virion
  • Virus: Giardia lamblia virus

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Supramolecule #1000: GLV virion

SupramoleculeName: GLV virion / type: sample / ID: 1000 / Oligomeric state: icosahedral / Number unique components: 1

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Supramolecule #1: Giardia lamblia virus

SupramoleculeName: Giardia lamblia virus / type: virus / ID: 1 / Name.synonym: GLV / NCBI-ID: 29255 / Sci species name: Giardia lamblia virus / Sci species strain: WBI / Database: NCBI / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No / Syn species name: GLV
Host (natural)Organism: Giardia intestinalis (eukaryote) / Strain: WBI / synonym: PROTOZOA
Virus shellShell ID: 1 / Diameter: 485 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

Concentration0.02 mg/mL
BufferpH: 7.2 / Details: 50 mM HEPES, 500 mM NaCl, 20 mM MgCl2
GridDetails: Quantifoil R2/2
VitrificationCryogen name: ETHANE / Chamber humidity: 99 % / Chamber temperature: 90 K / Instrument: HOMEMADE PLUNGER / Method: Blot 5 sec before plunging

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 58050 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.26 mm / Nominal defocus max: 3.41 µm / Nominal defocus min: 0.78 µm / Nominal magnification: 59000
Sample stageSpecimen holder model: OTHER
TemperatureMin: 90 K / Max: 96 K / Average: 93 K
Alignment procedureLegacy - Astigmatism: Objective lens astigmatism was corrected at 135,000 times magnification.
DateApr 18, 2012
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Digitization - Sampling interval: 6.35 µm / Number real images: 101 / Average electron dose: 25 e/Å2 / Bits/pixel: 8
Tilt angle min0
Tilt angle max0
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: robem
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 6.0 Å / Resolution method: OTHER / Software - Name: auto3dem
Details: Particles were selected and preprocessed using RobEM.
Number images used: 14125
DetailsParticles were selected and preprocessed using RobEM. Image reconstruction was performed using Auto3DEM.

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