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- EMDB-17064: Spherical assembly from truncated PVY coat protein with K177E mutation -

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Basic information

Entry
Database: EMDB / ID: EMD-17064
TitleSpherical assembly from truncated PVY coat protein with K177E mutation
Map datafinal trCPK177E spherical particle sharp cryoEM map
Sample
  • Complex: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
    • Protein or peptide: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
Keywordsspherical particle / trCP / K177E / Potyvirus / PVY / VIRUS LIKE PARTICLE
Biological speciesPotato virus Y strain NTN
Methodsingle particle reconstruction / cryo EM / Resolution: 12.75 Å
AuthorsKavcic L / Kezar A / Podobnik M
Funding support Slovenia, 2 items
OrganizationGrant numberCountry
Slovenian Research AgencyP1-0391 Slovenia
Slovenian Research AgencyJ7-7248 Slovenia
CitationJournal: Commun Chem / Year: 2024
Title: From structural polymorphism to structural metamorphosis of the coat protein of flexuous filamentous potato virus Y.
Authors: Luka Kavčič / Andreja Kežar / Neža Koritnik / Magda Tušek Žnidarič / Tajda Klobučar / Žiga Vičič / Franci Merzel / Ellie Holden / Justin L P Benesch / Marjetka Podobnik /
Abstract: The structural diversity and tunability of the capsid proteins (CPs) of various icosahedral and rod-shaped viruses have been well studied and exploited in the development of smart hybrid ...The structural diversity and tunability of the capsid proteins (CPs) of various icosahedral and rod-shaped viruses have been well studied and exploited in the development of smart hybrid nanoparticles. However, the potential of CPs of the wide-spread flexuous filamentous plant viruses remains to be explored. Here, we show that we can control the shape, size, RNA encapsidation ability, symmetry, stability and surface functionalization of nanoparticles through structure-based design of CP from potato virus Y (PVY). We provide high-resolution insight into CP-based self-assemblies, ranging from large polymorphic or monomorphic filaments to smaller annular, cubic or spherical particles. Furthermore, we show that we can prevent CP self-assembly in bacteria by fusion with a cleavable protein, enabling controlled nanoparticle formation in vitro. Understanding the remarkable structural diversity of PVY CP not only provides possibilities for the production of biodegradable nanoparticles, but may also advance future studies of CP's polymorphism in a biological context.
History
DepositionApr 7, 2023-
Header (metadata) releaseJan 24, 2024-
Map releaseJan 24, 2024-
UpdateJan 31, 2024-
Current statusJan 31, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_17064.png

Downloads & links

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Map

FileDownload / File: emd_17064.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationfinal trCPK177E spherical particle sharp cryoEM map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.63 Å/pix.
x 300 pix.
= 489. Å		1.63 Å/pix.
x 300 pix.
= 489. Å		1.63 Å/pix.
x 300 pix.
= 489. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.63 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.26
Minimum - Maximum-0.30947614 - 0.75922054
Average (Standard dev.)-0.0015992993 (±0.06700913)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 489.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_17064_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: trCPK177E spherical particle raw cryoEM map

Fileemd_17064_additional_1.map
AnnotationtrCPK177E spherical particle raw cryoEM map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: trCPK177E spherical particle ab-initio reconstruction result

Fileemd_17064_additional_2.map
AnnotationtrCPK177E spherical particle ab-initio reconstruction result
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: trCPK177E spherical particle half A cryoEM map

Fileemd_17064_half_map_1.map
AnnotationtrCPK177E spherical particle half A cryoEM map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: trCPK177E spherical particle half B cryoEM map

Fileemd_17064_half_map_2.map
AnnotationtrCPK177E spherical particle half B cryoEM map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : truncated coat protein (dN49C40) with C-terminal His tag and K177...

EntireName: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
Components
  • Complex: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
    • Protein or peptide: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation

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Supramolecule #1: truncated coat protein (dN49C40) with C-terminal His tag and K177...

SupramoleculeName: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Sample contains a mixture of cubic particles and spherical particles.
Source (natural)Organism: Potato virus Y strain NTN

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Macromolecule #1: truncated coat protein (dN49C40) with C-terminal His tag and K177...

MacromoleculeName: truncated coat protein (dN49C40) with C-terminal His tag and K177E mutation
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Potato virus Y strain NTN / Strain: NTN
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: GITSKMRMPK SKGATVLNLE HLLEYAPQQI DISNTRATQS QFDTWYEAVQ LAYDIGETEM PTVMNGLMVW CIENGTSPNI NGVWVMMDGD EQVEYPLKPI VENAKPTLRQ IMAHFSDVAE AYIEMRNKEE PYMPRYGLVR NLRDGSLARY AFDFYEVTSR TPVRAREAHI ...String:
GITSKMRMPK SKGATVLNLE HLLEYAPQQI DISNTRATQS QFDTWYEAVQ LAYDIGETEM PTVMNGLMVW CIENGTSPNI NGVWVMMDGD EQVEYPLKPI VENAKPTLRQ IMAHFSDVAE AYIEMRNKEE PYMPRYGLVR NLRDGSLARY AFDFYEVTSR TPVRAREAHI QMKAAALKSE NLYFQGLEHH HHHH

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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

Concentration3 mg/mL
BufferpH: 7.4
Details: 1.8 mM KH2PO4, 10.1 mM Na2HPO4, 140 mM NaCl, 2.7 mM KCl, pH 7.4
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number real images: 501 / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.1 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 150000

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Image processing

Particle selectionNumber selected: 111383
Startup modelType of model: NONE
Details: Initial model was obtained by ab-initio reconstruction
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 12.75 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 3452
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationSoftware - Name: cryoSPARC
FSC plot (resolution estimation)

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