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- EMDB-23560: 5-fold sub-particle reconstruction from Trichomonas vaginalis vir... -

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Basic information

Entry
Database: EMDB / ID: EMD-23560
Title5-fold sub-particle reconstruction from Trichomonas vaginalis virus 2 capsid
Map dataTVV viral Capsid sub-particle (C5)
Sample
  • Virus: Trichomonas vaginalis virus 2
    • Protein or peptide: Capsid protein
KeywordsCapsid / Viral Protein
Function / homologyCapsid protein
Function and homology information
Biological speciesTrichomonas vaginalis virus 2
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsZhou ZH / Stevens AW
Funding support United States, 10 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Dental and Craniofacial Research (NIH/NIDCR)DE028583 United States
National Institutes of Health/National Institute of Dental and Craniofacial Research (NIH/NIDCR)DE025567 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI094386 United States
National Institutes of Health/Office of the Director1S10OD018111 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1U24GM116792 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI103182 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R33AI119721 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI007323 United States
National Science Foundation (NSF, United States)DBI-1338135 United States
National Science Foundation (NSF, United States)DMR-1548924 United States
CitationJournal: mBio / Year: 2021
Title: Atomic Structure of the Trichomonas vaginalis Double-Stranded RNA Virus 2.
Authors: Alexander Stevens / Katherine Muratore / Yanxiang Cui / Patricia J Johnson / Z Hong Zhou /
Abstract: , the causative pathogen for the most common nonviral sexually transmitted infection worldwide, is itself frequently infected with one or more of the four types of small double-stranded RNA (dsRNA) ..., the causative pathogen for the most common nonviral sexually transmitted infection worldwide, is itself frequently infected with one or more of the four types of small double-stranded RNA (dsRNA) viruses (TVV1 to 4, genus , family ). Each TVV encloses a nonsegmented genome within a single-layered capsid and replicates entirely intracellularly, like many dsRNA viruses, and unlike those in the family. Here, we have determined the structure of TVV2 by cryo-electron microscopy (cryoEM) at 3.6 Å resolution and derived an atomic model of its capsid. TVV2 has an icosahedral, T = 2*, capsid comprised of 60 copies of the icosahedral asymmetric unit (a dimer of the two capsid shell protein [CSP] conformers, CSP-A and CSP-B), typical of icosahedral dsRNA virus capsids. However, unlike the robust CSP-interlocking interactions such as the use of auxiliary "clamping" proteins among , only lateral CSP interactions are observed in TVV2, consistent with an assembly strategy optimized for TVVs' intracellular-only replication cycles within their protozoan host. The atomic model reveals both a mostly negatively charged capsid interior, which is conducive to movement of the loosely packed genome, and channels at the 5-fold vertices, which we suggest as routes of mRNA release during transcription. Structural comparison of TVV2 to the L-A virus reveals a conserved helix-rich fold within the CSP and putative guanylyltransferase domain along the capsid exterior, suggesting conserved mRNA maintenance strategies among This first atomic structure of a TVV provides a framework to guide future biochemical investigations into the interplay between and its viruses. viruses (TVVs) are double-stranded RNA (dsRNA) viruses that cohabitate in , the causative pathogen of trichomoniasis, the most common nonviral sexually transmitted disease worldwide. Featuring an unsegmented dsRNA genome encoding a single capsid shell protein (CSP), TVVs contrast with multisegmented dsRNA viruses, such as the diarrhea-causing rotavirus, whose larger genome is split into 10 dsRNA segments encoding 5 unique capsid proteins. To determine how TVVs incorporate the requisite functionalities for viral replication into their limited proteome, we derived the atomic model of TVV2, a first for TVVs. Our results reveal the intersubunit interactions driving CSP association for capsid assembly and the properties that govern organization and maintenance of the viral genome. Structural comparison between TVV2 capsids and those of distantly related dsRNA viruses indicates conserved strategies of nascent RNA release and a putative viral guanylyltransferase domain implicated in the cytoplasmic maintenance of viral messenger and genomic RNA.
History
DepositionMar 2, 2021-
Header (metadata) releaseApr 7, 2021-
Map releaseApr 7, 2021-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.014
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.014
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lwy
  • Surface level: 0.014
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7lwy
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23560.png

Downloads & links

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Map

FileDownload / File: emd_23560.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTVV viral Capsid sub-particle (C5)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.36 Å/pix.
x 256 pix.
= 348.16 Å		1.36 Å/pix.
x 256 pix.
= 348.16 Å		1.36 Å/pix.
x 256 pix.
= 348.16 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.36 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.014 / Movie #1: 0.014
Minimum - Maximum-0.021746783 - 0.050101865
Average (Standard dev.)0.00026565584 (±0.002994053)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 348.16 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.361.361.36
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z348.160348.160348.160
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ160160160
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0220.0500.000

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Supplemental data

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Sample components

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Entire : Trichomonas vaginalis virus 2

EntireName: Trichomonas vaginalis virus 2
Components
  • Virus: Trichomonas vaginalis virus 2
    • Protein or peptide: Capsid protein

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Supramolecule #1: Trichomonas vaginalis virus 2

SupramoleculeName: Trichomonas vaginalis virus 2 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Virus particles isolated from lysed trichomonas vaginalis
NCBI-ID: 674954 / Sci species name: Trichomonas vaginalis virus 2 / Sci species strain: 2 / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Trichomonas vaginalis G3 (eukaryote) / Strain: G3
Molecular weightTheoretical: 9.40 MDa
Virus shellShell ID: 1 / Name: Capsid / Diameter: 430.0 Å / T number (triangulation number): 1

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Macromolecule #1: Capsid protein

MacromoleculeName: Capsid protein / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Trichomonas vaginalis virus 2
Molecular weightTheoretical: 74.037117 KDa
SequenceString: SNPRLTKVLD EMSKKPCVNI NEIRKMIRNF QPQFIQPRNG NRPNAQPRTV DSFEWVVRIQ STVETQLLGA TNTVPQQTLN LDISFTDDS TTITPASIPG SISMLDNSRH IPAIQSMIQN FKARYLGSLQ DTAQLQSPQY PQLLAYLFGQ LIAIKDRLDL F RPSNPLSL ...String:
SNPRLTKVLD EMSKKPCVNI NEIRKMIRNF QPQFIQPRNG NRPNAQPRTV DSFEWVVRIQ STVETQLLGA TNTVPQQTLN LDISFTDDS TTITPASIPG SISMLDNSRH IPAIQSMIQN FKARYLGSLQ DTAQLQSPQY PQLLAYLFGQ LIAIKDRLDL F RPSNPLSL ADALFGFTLA QNARPRYDDH RHAKACQGPL VIPAATNSDC GPCGFVQINA NQGLTLPLGA CLFVNPETVN DQ SFQDFLW LIFATHHRMP NQMQNNWPFS LNIVSTCAAP GRQAPHAGEL TDERVRLALD TGHRILLSMF NDDEETLRYY QRK GIETMF RPCCFYTEGG LLRKATRYVS MVPLNGLYYY NGATSYVVSP IHTDAHPGIT AAIESFVDIM VLQAVFSFSG PKVV AAKVN ASQIDAAMVF GPAVAEGDGF VYDPLRPAPP LSAFYTEFIH RPAEQRIFQM AMSQIYGSHA PLIIANVINS IHNCK TKIV NNKLRATFVR RPPGAPHLKA DTAIINRFHD PELAYALGIL ADGIAPLDGS HEYNVLDELD YLFNGGDIRN CFGLNA LNT RGLGQIVHIR PKREPGKRPR RGFYTTLDGQ VHPVTQDAPL DEIYHWRDHG NLTRPYSCHI LDSQGLEFAD VSNGRSR GK ILVVVNSPLK TCAAYQGPSF APK

UniProtKB: Capsid protein

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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

BufferpH: 7.4
Component:
ConcentrationFormulaName
150.0 mMNaClSodium Chloride
50.0 mMTristris(hydroxymethyl)aminomethane
2.0 mMMgCl2Magnesium Chloride
1.0 mMDTTDithiothreitol
1.0 XHALTHalt Protease Inhibitor Cocktail (100X)

Details: Sterile Solution was prepared fresh to prevent contamination.
GridModel: PELCO Ultrathin Carbon with Lacey Carbon / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE
Details: The grids were manually plunged into liquid ethane..

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Number real images: 2177 / Average exposure time: 0.2 sec. / Average electron dose: 17.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 5076
Startup modelType of model: OTHER / Details: a gaussian ball
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 29916
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)

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Atomic model buiding 1

DetailsModel was manually built into EM map using Coot, then PHENIX was used for real-space refinement
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-7lwy:
TVV viral capsid protein

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