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- PDB-7jqq: The bacteriophage Phi-29 viral genome packaging motor assembly -

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Basic information

Entry
Database: PDB / ID: 7jqq
TitleThe bacteriophage Phi-29 viral genome packaging motor assembly
Components
  • (DNA (60-MER)) x 2
  • DNA packaging protein
  • pRNA (117-MER)
KeywordsMOTOR PROTEIN / packaging motor / ATPase
Function / homology
Function and homology information


viral DNA genome packaging / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / ATP hydrolysis activity / DNA binding / RNA binding / ATP binding
Similarity search - Function
Podovirus DNA packaging protein / Podovirus DNA encapsidation protein (Gp16) / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / DNA / DNA (> 10) / RNA / RNA (> 10) / RNA (> 100) / DNA packaging protein
Similarity search - Component
Biological speciesBacillus phage phi29 (virus)
Bacillus virus phi29
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsWhite, M.A. / Woodson, M. / Morais, M.C.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM122979 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM127365 United States
CitationJournal: Sci Adv / Year: 2021
Title: A viral genome packaging motor transitions between cyclic and helical symmetry to translocate dsDNA.
Authors: Michael Woodson / Joshua Pajak / Bryon P Mahler / Wei Zhao / Wei Zhang / Gaurav Arya / Mark A White / Paul J Jardine / Marc C Morais /
Abstract: Molecular segregation and biopolymer manipulation require the action of molecular motors to do work by applying directional forces to macromolecules. The additional strand conserved E (ASCE) ring ...Molecular segregation and biopolymer manipulation require the action of molecular motors to do work by applying directional forces to macromolecules. The additional strand conserved E (ASCE) ring motors are an ancient family of molecular motors responsible for diverse biological polymer manipulation tasks. Viruses use ASCE segregation motors to package their genomes into their protein capsids and provide accessible experimental systems due to their relative simplicity. We show by cryo-EM-focused image reconstruction that ASCE ATPases in viral double-stranded DNA (dsDNA) packaging motors adopt helical symmetry complementary to their dsDNA substrates. Together with previous data, our results suggest that these motors cycle between helical and planar configurations, providing a possible mechanism for directional translocation of DNA. Similar changes in quaternary structure have been observed for proteasome and helicase motors, suggesting an ancient and common mechanism of force generation that has been adapted for specific tasks over the course of evolution.
History
DepositionAug 11, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 19, 2021Provider: repository / Type: Initial release
Revision 1.1Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.2May 1, 2024Group: Data processing / Category: em_experiment / Item: _em_experiment.entity_assembly_id

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
A: DNA packaging protein
B: DNA packaging protein
C: DNA packaging protein
D: DNA packaging protein
E: DNA packaging protein
K: pRNA (117-MER)
L: pRNA (117-MER)
M: pRNA (117-MER)
N: pRNA (117-MER)
O: pRNA (117-MER)
F: DNA (60-MER)
G: DNA (60-MER)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)421,02318
Polymers419,38112
Non-polymers1,6436
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy, This is the focused CryoEM reconstruction of the motor assembly attached to the phage prohead at the portal.
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area40340 Å2
ΔGint-248 kcal/mol
Surface area187500 Å2

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Components

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DNA chain , 2 types, 2 molecules FG

#3: DNA chain DNA (60-MER)


Mass: 18491.848 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bacillus virus phi29
#4: DNA chain DNA (60-MER)


Mass: 18491.850 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus virus phi29 / Production host: unidentified (others)

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Protein / RNA chain , 2 types, 10 molecules ABCDEKLMNO

#1: Protein
DNA packaging protein / ATPase gp16 / Gene product 16 / gp16 / Protein p16


Mass: 39010.406 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus phage phi29 (virus) / Gene: 16 / Plasmid: pSUMO-SacI-XhoI / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3)
References: UniProt: P11014, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
#2: RNA chain
pRNA (117-MER)


Mass: 37469.012 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus virus phi29 / Production host: unidentified (others)

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Non-polymers , 2 types, 6 molecules

#5: Chemical ChemComp-AGS / PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / ATP-GAMMA-S / ADENOSINE 5'-(3-THIOTRIPHOSPHATE) / ADENOSINE 5'-(GAMMA-THIOTRIPHOSPHATE) / ADENOSINE-5'-DIPHOSPHATE MONOTHIOPHOSPHATE


Mass: 523.247 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H16N5O12P3S / Comment: ATP-gamma-S, energy-carrying molecule analogue*YM
#6: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Mg

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Details

Has ligand of interestN
Sequence detailsThe authors state that the genomic DNA sequences are unknown and the complementary GTCA repeat ...The authors state that the genomic DNA sequences are unknown and the complementary GTCA repeat sequences are just placeholders.

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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: Bacillus virus phi29 viral genome packaging motor assembly
Type: VIRUS / Entity ID: #1-#4 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Bacillus virus phi29
Details of virusEmpty: NO / Enveloped: NO / Isolate: SPECIES / Type: VIRION
Natural hostOrganism: Bacillus subtilis / Strain: A12
Buffer solutionpH: 7.8
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMTris(HOCH2)3CNH21
25 mMmagnesium chlorideMgCl21
350 mMsodium chlorideNaCl1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: Quantifoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 4200 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 4200 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 41.4 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter name: GIF 2002
Image scansWidth: 7676 / Height: 7420 / Movie frames/image: 45 / Used frames/image: 0-44

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
EM software
IDNameVersionCategory
4CTFFIND2.03CTF correction
10RELION2.03initial Euler assignment
11RELION2.03final Euler assignment
12RELION2.03classification
13RELION2.13D reconstruction
CTF correctionType: PHASE FLIPPING ONLY
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 12526 / Algorithm: BACK PROJECTION
Details: The final step before reconstruction was classification without alignment, which was done with even/odd sets mixed together, but even/odd were completely independent for all angular assignment steps
Num. of class averages: 1 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 55.8 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.003930472
ELECTRON MICROSCOPYf_angle_d0.861244568
ELECTRON MICROSCOPYf_chiral_restr0.0445408
ELECTRON MICROSCOPYf_plane_restr0.00533128
ELECTRON MICROSCOPYf_dihedral_angle_d25.2510061

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