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- EMDB-35432: C2 reconstruction of the concave tetramer in the cube-like assemb... -

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Basic information

Entry
Database: EMDB / ID: EMD-35432
TitleC2 reconstruction of the concave tetramer in the cube-like assembly of 201Phi2-1 gp105
Map dataThe cryo-EM map of the phage-encoded nucleus-like shell protein gp105 concave tetramer reconstructed with C2 symmetry imposed.
Sample
  • Complex: 201Phi2-1 gp105
    • Protein or peptide: Chimallin
Function / homologyhost cell cytoplasm / Chimallin
Function and homology information
Biological speciesPseudomonas phage 201phi2-1 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.09 Å
AuthorsLiu Z / Xiang Y
Funding support China, 1 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)2021YFA1300204 China
CitationJournal: Front Microbiol / Year: 2023
Title: Structural studies of the nucleus-like assembly of jumbo bacteriophage 201φ2-1.
Authors: Zhe Liu / Ye Xiang /
Abstract: The jumbo phages encode proteins that assemble to form a nucleus-like compartment in infected cells. Here we report the cryo-EM structure and biochemistry characterization of gp105, a protein that is ...The jumbo phages encode proteins that assemble to form a nucleus-like compartment in infected cells. Here we report the cryo-EM structure and biochemistry characterization of gp105, a protein that is encoded by the jumbo phage 201φ2-1 and is involved in the formation of the nucleus-like compartment in phage 201φ2-1 infected . We found that, although most gp105 molecules are in the monomeric state in solution, a small portion of gp105 assemble to form large sheet-like assemblies and small cube-like particles. Reconstruction of the cube-like particles showed that the particle consists of six flat head-to-tail tetramers arranged into an octahedral cube. The four molecules at the contact interface of two head-to-tail tetramers are 2-fold symmetry-related and constitute a concave tetramer. Further reconstructions without applying symmetry showed that molecules in the particles around the distal ends of a 3-fold axis are highly dynamic and have the tendency to open up the assembly. Local classifications and refinements of the concave tetramers in the cube-like particle resulted in a map of the concave tetramer at a resolution of 4.09 Å. Structural analysis of the concave tetramer indicates that the N and C terminal fragments of gp105 are important for mediating the intermolecular interactions, which was further confirmed by mutagenesis studies. Biochemistry assays showed that, in solution, the cube-like particles of gp105 are liable to either disassemble to form the monomers or recruit more molecules to form the high molecular weight lattice-like assembly. We also found that monomeric gp105s can self-assemble to form large sheet-like assemblies , and the assembly of gp105 is a reversible dynamic process and temperature-dependent. Taken together, our results revealed the dynamic assembly of gp105, which helps to understand the development and function of the nucleus-like compartment assembled by phage-encoded proteins.
History
DepositionFeb 20, 2023-
Header (metadata) releaseMay 3, 2023-
Map releaseMay 3, 2023-
UpdateMay 17, 2023-
Current statusMay 17, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_35432.png

Downloads & links

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Map

FileDownload / File: emd_35432.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe cryo-EM map of the phage-encoded nucleus-like shell protein gp105 concave tetramer reconstructed with C2 symmetry imposed.
Voxel sizeX=Y=Z: 1.25 Å
Density
Contour LevelBy AUTHOR: 0.018
Minimum - Maximum-0.07349291 - 0.1013904
Average (Standard dev.)0.00030418325 (±0.004104856)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 225.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: The cryo-EM half map of the phage-encoded nucleus-like...

Fileemd_35432_half_map_1.map
AnnotationThe cryo-EM half map of the phage-encoded nucleus-like shell protein gp105 concave tetramer reconstructed with C2 symmetry imposed.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: The cryo-EM half map of the phage-encoded nucleus-like...

Fileemd_35432_half_map_2.map
AnnotationThe cryo-EM half map of the phage-encoded nucleus-like shell protein gp105 concave tetramer reconstructed with C2 symmetry imposed.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 201Phi2-1 gp105

EntireName: 201Phi2-1 gp105
Components
  • Complex: 201Phi2-1 gp105
    • Protein or peptide: Chimallin

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Supramolecule #1: 201Phi2-1 gp105

SupramoleculeName: 201Phi2-1 gp105 / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas phage 201phi2-1 (virus)

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Macromolecule #1: Chimallin

MacromoleculeName: Chimallin / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas phage 201phi2-1 (virus)
Molecular weightTheoretical: 70.654516 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MIRDTATNTT QTQAAPQQAP AQQFTQAPQE KPMQSTQSQP TPSYAGTGGI NSQFTRSGNV QGGDARASEA LTVFTRLKEQ AVAQQDLAD DFSILRFDRD QHQVGWSSLV IAKQISLNGQ PVIAVRPLIL PNNSIELPKR KTNIVNGMQT DVIESDIDVG T VFSAQYFN ...String:
MIRDTATNTT QTQAAPQQAP AQQFTQAPQE KPMQSTQSQP TPSYAGTGGI NSQFTRSGNV QGGDARASEA LTVFTRLKEQ AVAQQDLAD DFSILRFDRD QHQVGWSSLV IAKQISLNGQ PVIAVRPLIL PNNSIELPKR KTNIVNGMQT DVIESDIDVG T VFSAQYFN RLSTYVQNTL GKPGAKVVLA GPFPIPADLV LKDSELQLRN LLIKSVNACD DILALHSGER PFTIAGLKGQ QG ETLAAKV DIRTQPLHDT VGNPIRADIV VTTQRVRRNG QQENEFYETD VKLNQVAMFT NLERTPQAQA QTLFPNQQQV ATP APWVAS VVITDVRNAD GIQANTPEMY WFALSNAFRS THGHAWARPF LPMTGVAKDM KDIGALGWMS ALRNRIDTKA ANFD DAQFG QLMLSQVQPN PVFQIDLNRM GETAQMDSLQ LDAAGGPNAQ KAAATIIRQI NNLGGGGFER FFDHTTQPIL ERTGQ VIDL GNWFDGDEKR DRRDLDNLAA LNAAEGNENE FWGFYGAQLN PNLHPDLRNR QSRNYDRQYL GSTVTYTGKA ERCTYN AKF IEALDRYLAE AGLQITMDNT SVLNSGQRFM GNSVIGNNMV SGQAQVHSAY AGTQGFNTQY QTGPSSFYAL EHHHHHH

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 1.2 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Details: We used AlphaFold for the structure prediction of the nucleus-like shell protein (NLSP) gp105. The predicted model was used as a reference model for model building.
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 4
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionNumber classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 4.09 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 290996
FSC plot (resolution estimation)

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