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- PDB-8tvr: In situ cryo-EM structure of bacteriophage P22 tail hub protein: ... -

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Basic information

Entry
Database: PDB / ID: 8tvr
TitleIn situ cryo-EM structure of bacteriophage P22 tail hub protein: tailspike protein complex at 2.8A resolution
Components
  • Packaged DNA stabilization protein gp10
  • Tail spike protein
KeywordsVIRAL PROTEIN / phage / bacteriophage / tail spike protein / TSP / gene product 9 (gp9) / Packaged DNA stabilization protein / gene product 10 (gp10) / STRUCTURAL PROTEIN
Function / homology
Function and homology information


endo-1,3-alpha-L-rhamnosidase activity / symbiont entry into host cell via disruption of host cell envelope lipopolysaccharide / virus tail, fiber / symbiont entry into host cell via disruption of host cell envelope / symbiont entry into host / Hydrolases; Glycosylases; Glycosidases, i.e. enzymes that hydrolyse O- and S-glycosyl compounds / adhesion receptor-mediated virion attachment to host cell / metabolic process / virion attachment to host cell
Similarity search - Function
Bacteriophage P22, Gp10, DNA-stabilising / Phage stabilisation protein / P22 tailspike C-terminal domain / Salmonella phage P22 tail-spike / Bacteriophage P22 tailspike, N-terminal / Phage P22 tailspike-like, N-terminal domain superfamily / Head binding / Autotransporter, pectate lyase C-like domain superfamily / Pectin lyase fold/virulence factor
Similarity search - Domain/homology
Tail spike protein / Packaged DNA stabilization protein gp10
Similarity search - Component
Biological speciesSalmonella phage P22 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsIglesias, S. / 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: Tail spike protein
B: Tail spike protein
C: Tail spike protein
S: Packaged DNA stabilization protein gp10
D: Tail spike protein
E: Tail spike protein
F: Tail spike protein
G: Packaged DNA stabilization protein gp10
H: Tail spike protein
I: Tail spike protein
J: Tail spike protein
K: Packaged DNA stabilization protein gp10
L: Tail spike protein
M: Tail spike protein
N: Tail spike protein
O: Packaged DNA stabilization protein gp10
P: Tail spike protein
Q: Tail spike protein
R: Tail spike protein
T: Packaged DNA stabilization protein gp10
V: Tail spike protein
W: Tail spike protein
X: Tail spike protein
Y: Packaged DNA stabilization protein gp10


Theoretical massNumber of molelcules
Total (without water)1,609,69624
Polymers1,609,69624
Non-polymers00
Water00
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
Tail spike protein


Mass: 71923.523 Da / Num. of mol.: 18 / Source method: isolated from a natural source / Source: (natural) Salmonella phage P22 (virus) / References: UniProt: P12528
#2: Protein
Packaged DNA stabilization protein gp10


Mass: 52512.020 Da / Num. of mol.: 6 / Source method: isolated from a natural source / Source: (natural) Salmonella phage P22 (virus) / References: UniProt: P26749

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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
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 softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 41597
SymmetryPoint symmetry: C6 (6 fold cyclic)
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 38155 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00339342
ELECTRON MICROSCOPYf_angle_d0.51653442
ELECTRON MICROSCOPYf_dihedral_angle_d4.8175418
ELECTRON MICROSCOPYf_chiral_restr0.0495934
ELECTRON MICROSCOPYf_plane_restr0.0046996

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