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- EMDB-25562: Kinetically trapped Pseudomonas-phage PaP3 portal protein - delta... -

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Basic information

Entry
Database: EMDB / ID: EMD-25562
TitleKinetically trapped Pseudomonas-phage PaP3 portal protein - delta barrel mutant class-3
Map data
Sample
  • Complex: Undecameric complex of phage PaP3 portal, C-terminal truncated mutant
    • Protein or peptide: Portal protein
KeywordsPortal protein / Dodecamer / VIRAL PROTEIN
Function / homologyORF.04
Function and homology information
Biological speciesPhage PaP3 (virus) / Pseudomonas virus PaP3
Methodsingle particle reconstruction / cryo EM / Resolution: 6.24 Å
AuthorsHou C-FD / Swanson NA
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 GM100888, R35 GM140733-0, P30 CA56036, HSSN261200800001E United States
CitationJournal: J Mol Biol / Year: 2022
Title: Cryo-EM Structure of a Kinetically Trapped Dodecameric Portal Protein from the Pseudomonas-phage PaP3.
Authors: Chun-Feng David Hou / Nicholas A Swanson / Fenglin Li / Ruoyu Yang / Ravi K Lokareddy / Gino Cingolani /
Abstract: Portal proteins are dodecameric assemblies that occupy a unique 5-fold vertex of the icosahedral capsid of tailed bacteriophages and herpesviruses. The portal vertex interrupts the icosahedral ...Portal proteins are dodecameric assemblies that occupy a unique 5-fold vertex of the icosahedral capsid of tailed bacteriophages and herpesviruses. The portal vertex interrupts the icosahedral symmetry, and in vivo, its assembly and incorporation in procapsid are controlled by the scaffolding protein. Ectopically expressed portal oligomers are polymorphic in solution, and portal rings built by a different number of subunits have been documented in the literature. In this paper, we describe the cryo-EM structure of the portal protein from the Pseudomonas-phage PaP3, which we determined at 3.4 Å resolution. Structural analysis revealed a dodecamer with helical rather than rotational symmetry, which we hypothesize is kinetically trapped. The helical assembly was stabilized by local mispairing of portal subunits caused by the slippage of crown and barrel helices that move like a lever with respect to the portal body. Removing the C-terminal barrel promoted assembly of undecameric and dodecameric rings with quasi-rotational symmetry, suggesting that the barrel contributes to subunits mispairing. However, ΔC-portal rings were intrinsically asymmetric, with most particles having one open portal subunit interface. Together, these data expand the structural repertoire of viral portal proteins to Pseudomonas-phages and shed light on the unexpected plasticity of the portal protein quaternary structure.
History
DepositionNov 25, 2021-
Header (metadata) releaseMar 30, 2022-
Map releaseMar 30, 2022-
UpdateFeb 28, 2024-
Current statusFeb 28, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_25562.png

Downloads & links

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Map

FileDownload / File: emd_25562.map.gz / Format: CCP4 / Size: 11.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.82 Å
Density
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.025165206 - 0.076010115
Average (Standard dev.)0.00065622246 (±0.006525926)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions144144144
Spacing144144144
CellA=B=C: 262.08002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_25562_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_25562_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_25562_half_map_2.map
Projections & Slices
AxesZYX

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Sample components

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Entire : Undecameric complex of phage PaP3 portal, C-terminal truncated mutant

EntireName: Undecameric complex of phage PaP3 portal, C-terminal truncated mutant
Components
  • Complex: Undecameric complex of phage PaP3 portal, C-terminal truncated mutant
    • Protein or peptide: Portal protein

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Supramolecule #1: Undecameric complex of phage PaP3 portal, C-terminal truncated mutant

SupramoleculeName: Undecameric complex of phage PaP3 portal, C-terminal truncated mutant
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Phage PaP3 (virus)

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Macromolecule #1: Portal protein

MacromoleculeName: Portal protein / type: protein_or_peptide / ID: 1 / Number of copies: 11 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas virus PaP3
Molecular weightTheoretical: 80.981344 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAKRRKIKPM DDEQVLRHLD QLVNDALDFN SSELSKQRSE ALKYYFGEPF GNERPGKSGI VSRDVQETVD WIMPSLMKVF TSGGQVVKY EPDTAEDVEQ AEQETEYVNY LFMRKNEGFK VMFDWFQDTL MMKTGVVKVY VEEVLKPTFE RFSGLSEDMV A DILSDPDT ...String:
MAKRRKIKPM DDEQVLRHLD QLVNDALDFN SSELSKQRSE ALKYYFGEPF GNERPGKSGI VSRDVQETVD WIMPSLMKVF TSGGQVVKY EPDTAEDVEQ AEQETEYVNY LFMRKNEGFK VMFDWFQDTL MMKTGVVKVY VEEVLKPTFE RFSGLSEDMV A DILSDPDT SILAQSVDDD GTYTIKIRKD KKKREIKVLC VKPENFLVDR LATCIDDARF LCHREKYTVS DLRLLGVPED VI EELPYDE YEFSDSQPER LVRDNFDMTG QLQYNSGDDA EANREVWASE CYTLLDVDGD GISELRRILY VGDYIISNEP WDC RPFADL NAYRIAHKFH GMSVYDKIRD IQEIRSVLMR NIMDNIYRTN QGRSVVLDGQ VNLEDLLTNE AAGIVRVKSM NSIT PLETP QLSGEVYGML DRLEADRGKR TGITDRTRGL DQNTLHSNQA AMSVNQLMTA AEQQIDLIAR MFAETGVKRL FQLLH DHAI KYQNQEEVFQ LRGKWVAVNP ANWRERSDLT VTVGIGNMNK DQQMLHLMRI WEMAQAVVGG GGLGVLVSEQ NLYNIL KEV TENAGYKDPD RFWTNPNSPE ALQAKAIREQ KEAQPKPEDI KAQADAQRAQ SDALAKQAEA QMKQVEAQIR LAEIELK KQ EAVLQQREMA LKEAELQLER DRFTWERARN EAEYHLEATQ ARAAYIGDGK VPETKKPTKA VRR

UniProtKB: ORF.04

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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

Concentration3.5 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
20.0 mMTris-HClTris
200.0 mMNaClSodium chloride
5.0 mMEDTAEthylenediaminetetraacetic acid
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS GLACIOS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 150000
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Average electron dose: 50.0 e/Å2

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Image processing

Particle selectionNumber selected: 1200000
Startup modelType of model: NONE
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.24 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 85000
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL / Overall B value: 129 / Target criteria: Correlation coefficient
Output model

PDB-7sz6:
Kinetically trapped Pseudomonas-phage PaP3 portal protein - delta barrel mutant class-3

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