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- EMDB-22379: Herpes Simplex Virus Type 1 A-capsid with Portal Vertex -

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Basic information

Entry
Database: EMDB / ID: EMD-22379
TitleHerpes Simplex Virus Type 1 A-capsid with Portal Vertex
Map dataHerpes sipmlex virus Type 1 A-capsid with the portal vertex
Sample
  • Virus: Human alphaherpesvirus 1 (Herpes simplex virus type 1)
Biological speciesHuman alphaherpesvirus 1 (Herpes simplex virus type 1)
Methodsubtomogram averaging / cryo EM / Resolution: 42.57 Å
AuthorsBuch MHC / Heymann JB / Newcomb WW / Winkler DC / Steven AC
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)Intramural Research Program United States
CitationJournal: mBio / Year: 2021
Title: Cryo-Electron Tomography of the Herpesvirus Procapsid Reveals Interactions of the Portal with the Scaffold and a Shift on Maturation.
Authors: Michael H C Buch / William W Newcomb / Dennis C Winkler / Alasdair C Steven / J Bernard Heymann /
Abstract: Herpes simplex virus 1 (HSV-1) requires seven proteins to package its genome through a vertex in its capsid, one of which is the portal protein, pUL6. The portal protein is also thought to facilitate ...Herpes simplex virus 1 (HSV-1) requires seven proteins to package its genome through a vertex in its capsid, one of which is the portal protein, pUL6. The portal protein is also thought to facilitate assembly of the procapsid. While the portal has been visualized in mature capsids, we aimed to elucidate its role in the assembly and maturation of procapsids using cryo-electron tomography (cryoET). We identified the portal vertex in individual procapsids, calculated a subtomogram average, and compared that with the portal vertex in empty mature capsids (A-capsids). The resulting maps show the portal on the interior surface with its narrower end facing outwards, while maintaining close contact with the capsid shell. In the procapsid, the portal is embedded in the underlying scaffold, suggesting that assembly involves a portal-scaffold complex. During maturation, the capsid shell angularizes with a corresponding outward movement of the vertices. We found that in A-capsids, the portal translocates outward further than the adjacent capsomers and strengthens its contacts with the capsid shell. Our methodology also allowed us to determine the number of portal vertices in each capsid, with most having one per capsid, but some none or two, and rarely three. The predominance of a single portal per capsid supports facilitation of the assembly of the procapsid. Herpes simplex virus 1 (HSV-1) infects a majority of humans, causing mostly mild disease but in some cases progressing toward life-threatening encephalitis. Understanding the life cycle of the virus is important to devise countermeasures. Production of the virion starts with the assembly of an icosahedral procapsid, which includes DNA packaging proteins at a vertex, one of which is the dodecameric portal protein. The procapsid then undergoes maturation and DNA packaging through the portal, driven by a terminase complex. We used cryo-electron tomography to visualize the portal in procapsids and compare them to mature empty capsids. We found the portal located inside one vertex interacting with the scaffold protein in the procapsid. On maturation, the scaffold is cleaved and dissociates, the capsid angularizes, and the portal moves outward, interacting closely with the capsid shell. These transformations may provide a basis for the development of drugs to prevent HSV-1 infections.
History
DepositionJul 30, 2020-
Header (metadata) releaseMar 10, 2021-
Map releaseMar 10, 2021-
UpdateSep 22, 2021-
Current statusSep 22, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: -0.06
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: -0.06
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22379.png

Downloads & links

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Map

FileDownload / File: emd_22379.map.gz / Format: CCP4 / Size: 347.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHerpes sipmlex virus Type 1 A-capsid with the portal vertex
Voxel sizeX=Y=Z: 3.66 Å
Density
Contour LevelBy AUTHOR: 0.0624 / Movie #1: -0.06
Minimum - Maximum-0.48863298 - 0.27769145
Average (Standard dev.)6.443275e-06 (±0.09214036)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-225-225-225
Dimensions450450450
Spacing450450450
CellA=B=C: 1647.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.663.663.66
M x/y/z450450450
origin x/y/z0.0000.0000.000
length x/y/z1647.0001647.0001647.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-225-225-225
NC/NR/NS450450450
D min/max/mean-0.4890.2780.000

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Supplemental data

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Mask #1

Fileemd_22379_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Herpes Simplex Virus Type 1 A-capsid with Portal Vertex

Fileemd_22379_half_map_1.map
AnnotationHerpes Simplex Virus Type 1 A-capsid with Portal Vertex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Herpes Simplex Virus Type 1 A-capsid with Portal Vertex

Fileemd_22379_half_map_2.map
AnnotationHerpes Simplex Virus Type 1 A-capsid with Portal Vertex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human alphaherpesvirus 1

EntireName: Human alphaherpesvirus 1 (Herpes simplex virus type 1)
Components
  • Virus: Human alphaherpesvirus 1 (Herpes simplex virus type 1)

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Supramolecule #1: Human alphaherpesvirus 1

SupramoleculeName: Human alphaherpesvirus 1 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Capsids were obtained from infected Vero cells. Purification steps were carried out at 4 deg. Harvested cells were resuspended in TNE buffer and lysed by sonication. Nuclei were lysed with 1. ...Details: Capsids were obtained from infected Vero cells. Purification steps were carried out at 4 deg. Harvested cells were resuspended in TNE buffer and lysed by sonication. Nuclei were lysed with 1.5% TX-100, and a mix of A-, B-, and C-capsids was pelleted through a 30% sucrose cushion. The resuspended capsid mix was applied to a 20-50% sucrose gradient, and the band containing A-capsids was isolated.
NCBI-ID: 10298 / Sci species name: Human alphaherpesvirus 1 / Sci species strain: KOS / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: SPECIES / Virus enveloped: Yes / Virus empty: No
Host (natural)Organism: Homo sapiens (human)
Host systemOrganism: Chlorocebus aethiops (grivet)
Virus shellShell ID: 1 / Name: A-capsid / Diameter: 1250.0 Å / T number (triangulation number): 16

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMTrisTris Buffer
500.0 mMNaClSodium Chloride
1.0 mMEDTAEthylene Diamine Tetraacetate
GridModel: Quantifoil / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER
Details: Grids were plasma-cleaned in a Model 1020 plasma cleaner (Fischione, Export, PA).
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: LEICA EM GP
Details: Sample was mixed with Aurion Anionic 10 nm BSA gold tracer (EMS) at a 1:1 ratio, and 3 uL were applied to Quantifoil copper grids covered with 300-mesh holey carbon (EMS). Excess liquid was ...Details: Sample was mixed with Aurion Anionic 10 nm BSA gold tracer (EMS) at a 1:1 ratio, and 3 uL were applied to Quantifoil copper grids covered with 300-mesh holey carbon (EMS). Excess liquid was blotted off for 2.5 s, and grids were flash-frozen in liquid ethane..

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Electron microscopy

MicroscopeJEOL 2200FS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Average exposure time: 1.0 sec. / Average electron dose: 3.1 e/Å2
Details: Tilts were imaged in movie mode with 3 movies at on average 0.5 seconds each.
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 20000
Sample stageSpecimen holder model: GATAN 914 HIGH TILT LIQUID NITROGEN CRYO TRANSFER TOMOGRAPHY HOLDER
Cooling holder cryogen: NITROGEN

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Image processing

Final reconstructionApplied symmetry - Point group: C5 (5 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 42.57 Å / Resolution method: FSC 0.33 CUT-OFF / Software - Name: Bsoft (ver. 2.07) / Number subtomograms used: 164
ExtractionNumber tomograms: 21 / Number images used: 164 / Method: Particles were picked by hand / Software - Name: Bsoft (ver. 2.0.7)
CTF correctionSoftware - Name: Bsoft (ver. 2.07)
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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