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- PDB-8tvu: In situ cryo-EM structure of bacteriophage P22 portal protein: he... -

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Basic information

Entry
Database: PDB / ID: 8tvu
TitleIn situ cryo-EM structure of bacteriophage P22 portal protein: head-to-tail protein complex at 3.0A resolution
Components
  • Peptidoglycan hydrolase gp4
  • Portal protein
KeywordsVIRAL PROTEIN / phage / bacteriophage / portal protein / head-to-tail protein / gene product 1 (gp1) / gene product 4 (gp4 / STRUCTURAL PROTEIN
Function / homology
Function and homology information


viral DNA genome packaging, headful / symbiont entry into host cell via disruption of host cell wall peptidoglycan / viral portal complex / symbiont genome ejection through host cell envelope, short tail mechanism / symbiont entry into host cell via disruption of host cell envelope / viral DNA genome packaging / virus tail / virion assembly / hydrolase activity
Similarity search - Function
Tail accessory factor GP4 / Phage P22-like portal protein / Peptidoglycan hydrolase Gp4 superfamily / P22 tail accessory factor / Phage P22-like portal protein
Similarity search - Domain/homology
Portal protein / Peptidoglycan hydrolase gp4
Similarity search - Component
Biological speciesSalmonella phage P22 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å
AuthorsIglesias, S.M. / Cingolani, G. / Feng-Hou, C.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)GM140733 United States
CitationJournal: J Mol Biol / Year: 2023
Title: Molecular Architecture of Salmonella Typhimurium Virus P22 Genome Ejection Machinery.
Authors: Stephano M Iglesias / Ravi K Lokareddy / Ruoyu Yang / Fenglin Li / Daniel P Yeggoni / Chun-Feng David Hou / Makayla N Leroux / Juliana R Cortines / Justin C Leavitt / Mary Bird / Sherwood R ...Authors: Stephano M Iglesias / Ravi K Lokareddy / Ruoyu Yang / Fenglin Li / Daniel P Yeggoni / Chun-Feng David Hou / Makayla N Leroux / Juliana R Cortines / Justin C Leavitt / Mary Bird / Sherwood R Casjens / Simon White / Carolyn M Teschke / Gino Cingolani /
Abstract: Bacteriophage P22 is a prototypical member of the Podoviridae superfamily. Since its discovery in 1952, P22 has become a paradigm for phage transduction and a model for icosahedral viral capsid ...Bacteriophage P22 is a prototypical member of the Podoviridae superfamily. Since its discovery in 1952, P22 has become a paradigm for phage transduction and a model for icosahedral viral capsid assembly. Here, we describe the complete architecture of the P22 tail apparatus (gp1, gp4, gp10, gp9, and gp26) and the potential location and organization of P22 ejection proteins (gp7, gp20, and gp16), determined using cryo-EM localized reconstruction, genetic knockouts, and biochemical analysis. We found that the tail apparatus exists in two equivalent conformations, rotated by ∼6° relative to the capsid. Portal protomers make unique contacts with coat subunits in both conformations, explaining the 12:5 symmetry mismatch. The tail assembles around the hexameric tail hub (gp10), which folds into an interrupted β-propeller characterized by an apical insertion domain. The tail hub connects proximally to the dodecameric portal protein and head-to-tail adapter (gp4), distally to the trimeric tail needle (gp26), and laterally to six trimeric tailspikes (gp9) that attach asymmetrically to gp10 insertion domain. Cryo-EM analysis of P22 mutants lacking the ejection proteins gp7 or gp20 and biochemical analysis of purified recombinant proteins suggest that gp7 and gp20 form a molecular complex associated with the tail apparatus via the portal protein barrel. We identified a putative signal transduction pathway from the tailspike to the tail needle, mediated by three flexible loops in the tail hub, that explains how lipopolysaccharide (LPS) is sufficient to trigger the ejection of the P22 DNA in vitro.
History
DepositionAug 18, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 29, 2023Provider: repository / Type: Initial release
Revision 1.1Dec 6, 2023Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.title

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Portal protein
a: Peptidoglycan hydrolase gp4
B: Portal protein
C: Peptidoglycan hydrolase gp4
D: Portal protein
E: Peptidoglycan hydrolase gp4
F: Portal protein
G: Peptidoglycan hydrolase gp4
H: Portal protein
I: Peptidoglycan hydrolase gp4
J: Portal protein
K: Peptidoglycan hydrolase gp4
L: Portal protein
M: Peptidoglycan hydrolase gp4
N: Portal protein
O: Peptidoglycan hydrolase gp4
P: Portal protein
Q: Peptidoglycan hydrolase gp4
R: Portal protein
S: Peptidoglycan hydrolase gp4
T: Portal protein
V: Peptidoglycan hydrolase gp4
W: Portal protein
X: Peptidoglycan hydrolase gp4


Theoretical massNumber of molelcules
Total (without water)1,210,49224
Polymers1,210,49224
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Portal protein


Mass: 82829.375 Da / Num. of mol.: 12 / Source method: isolated from a natural source / Source: (natural) Salmonella phage P22 (virus) / References: UniProt: P26744
#2: Protein
Peptidoglycan hydrolase gp4


Mass: 18044.959 Da / Num. of mol.: 12 / Source method: isolated from a natural source / Source: (natural) Salmonella phage P22 (virus) / References: UniProt: P26746

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Salmonella phage P22 / Type: VIRUS / Entity ID: all / Source: NATURAL
Molecular weightUnits: MEGADALTONS / Experimental value: NO
Source (natural)Organism: Salmonella phage P22 (virus)
Details of virusEmpty: NO / Enveloped: NO / Isolate: OTHER / Type: VIRION
Natural hostOrganism: Salmonella enterica
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER / Nominal magnification: 29000 X / Calibrated magnification: 29000 X / Nominal defocus max: 2100 nm / Nominal defocus min: 800 nm / Calibrated defocus min: 800 nm / Calibrated defocus max: 2100 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 1.08 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
Image scansWidth: 11520 / Height: 8184

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Processing

EM software
IDNameVersionCategory
2PHENIX1.20.1_4487:model refinement
10RELION3.1.2initial Euler assignment
11RELION3.1.2final Euler assignment
12RELION3.1.2classification
13RELION3.1.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C12 (12 fold cyclic)
3D reconstructionResolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 38151 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00474340
ELECTRON MICROSCOPYf_angle_d0.669100800
ELECTRON MICROSCOPYf_dihedral_angle_d3.8889972
ELECTRON MICROSCOPYf_chiral_restr0.04210908
ELECTRON MICROSCOPYf_plane_restr0.00513344

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