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- EMDB-20526: CryoEM Structure of Pyocin R2 - precontracted - trunk -

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Basic information

Entry
Database: EMDB / ID: EMD-20526
TitleCryoEM Structure of Pyocin R2 - precontracted - trunk
Map dataCryoEM Structure of Pyocin R2
Sample
  • Organelle or cellular component: Pyocin R2
    • Protein or peptide: Pyocin tube PA0623
    • Protein or peptide: Pyocin sheath PA0622
Keywordsbacteriocin / pyocin / helix / ANTIMICROBIAL PROTEIN
Function / homologyTail tube protein / Phage tail tube protein FII / Tail sheath protein, subtilisin-like domain / Phage tail sheath protein subtilisin-like domain / Tail sheath protein, C-terminal domain / Phage tail sheath C-terminal domain / Probable bacteriophage protein / Probable bacteriophage protein
Function and homology information
Biological speciesPseudomonas aeruginosa PAO1 (bacteria) / Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Methodhelical reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsGe P / Avaylon J
Funding support United States, Switzerland, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM071940 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R21AI085318 United States
Swiss National Science Foundation31003A_146284 Switzerland
National Science Foundation (NSF, United States)XSEDE MCB140140 United States
CitationJournal: Nature / Year: 2020
Title: Action of a minimal contractile bactericidal nanomachine.
Authors: Peng Ge / Dean Scholl / Nikolai S Prokhorov / Jaycob Avaylon / Mikhail M Shneider / Christopher Browning / Sergey A Buth / Michel Plattner / Urmi Chakraborty / Ke Ding / Petr G Leiman / Jeff ...Authors: Peng Ge / Dean Scholl / Nikolai S Prokhorov / Jaycob Avaylon / Mikhail M Shneider / Christopher Browning / Sergey A Buth / Michel Plattner / Urmi Chakraborty / Ke Ding / Petr G Leiman / Jeff F Miller / Z Hong Zhou /
Abstract: R-type bacteriocins are minimal contractile nanomachines that hold promise as precision antibiotics. Each bactericidal complex uses a collar to bridge a hollow tube with a contractile sheath loaded ...R-type bacteriocins are minimal contractile nanomachines that hold promise as precision antibiotics. Each bactericidal complex uses a collar to bridge a hollow tube with a contractile sheath loaded in a metastable state by a baseplate scaffold. Fine-tuning of such nucleic acid-free protein machines for precision medicine calls for an atomic description of the entire complex and contraction mechanism, which is not available from baseplate structures of the (DNA-containing) T4 bacteriophage. Here we report the atomic model of the complete R2 pyocin in its pre-contraction and post-contraction states, each containing 384 subunits of 11 unique atomic models of 10 gene products. Comparison of these structures suggests the following sequence of events during pyocin contraction: tail fibres trigger lateral dissociation of baseplate triplexes; the dissociation then initiates a cascade of events leading to sheath contraction; and this contraction converts chemical energy into mechanical force to drive the iron-tipped tube across the bacterial cell surface, killing the bacterium.
History
DepositionJul 30, 2019-
Header (metadata) releaseAug 28, 2019-
Map releaseApr 15, 2020-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6pyt
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6pyt
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20526.png

Downloads & links

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Map

FileDownload / File: emd_20526.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM Structure of Pyocin R2
Voxel sizeX=Y=Z: 1.041 Å
Density
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.09599879 - 0.14136648
Average (Standard dev.)0.00020344139 (±0.007855937)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-192-192-192
Dimensions384384384
Spacing384384384
CellA=B=C: 399.74402 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0411.0411.041
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z399.744399.744399.744
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ460460460
MAP C/R/S123
start NC/NR/NS-192-192-192
NC/NR/NS384384384
D min/max/mean-0.0960.1410.000

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Supplemental data

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Sample components

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Entire : Pyocin R2

EntireName: Pyocin R2
Components
  • Organelle or cellular component: Pyocin R2
    • Protein or peptide: Pyocin tube PA0623
    • Protein or peptide: Pyocin sheath PA0622

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Supramolecule #1: Pyocin R2

SupramoleculeName: Pyocin R2 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas aeruginosa PAO1 (bacteria)

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Macromolecule #1: Pyocin tube PA0623

MacromoleculeName: Pyocin tube PA0623 / type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 17.957352 KDa
SequenceString:
MIPQTLTNTN LFIDGVSFAG DVPSLTLPKL AVKTEQYRAG GMDAPVSIDM GLEAMEAKFS TNGARREALN FFGLADQSAF NGVFRGSFK GQKGASVPVV ATLRGLLKEV DPGDWKAGEK AEFKYAVAVS YYKLEVDGRE VYEIDPVNGV RAINGVDQLA G MRNDLGL

UniProtKB: Probable bacteriophage protein

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Macromolecule #2: Pyocin sheath PA0622

MacromoleculeName: Pyocin sheath PA0622 / type: protein_or_peptide / ID: 2 / Number of copies: 12 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 41.247332 KDa
SequenceString: MSFFHGVTVT NVDIGARTIA LPASSVIGLC DVFTPGAQAS AKPNVPVLLT SKKDAAAAFG IGSSIYLACE AIYNRAQAVI VAVGVETAE TPEAQASAVI GGISAAGERT GLQALLDGKS RFNAQPRLLV APGHSAQQAV ATAMDGLAEK LRAIAILDGP N STDEAAVA ...String:
MSFFHGVTVT NVDIGARTIA LPASSVIGLC DVFTPGAQAS AKPNVPVLLT SKKDAAAAFG IGSSIYLACE AIYNRAQAVI VAVGVETAE TPEAQASAVI GGISAAGERT GLQALLDGKS RFNAQPRLLV APGHSAQQAV ATAMDGLAEK LRAIAILDGP N STDEAAVA YAKNFGSKRL FMVDPGVQVW DSATNAARNA PASAYAAGLF AWTDAEYGFW SSPSNKEIKG VTGTSRPVEF LD GDETCRA NLLNNANIAT IIRDDGYRLW GNRTLSSDSK WAFVTRVRTM DLVMDAILAG HKWAVDRGIT KTYVKDVTEG LRA FMRDLK NQGAVINFEV YADPDLNSAS QLAQGKVYWN IRFTDVPPAE NPNFRVEVTD QWLTEVLDVA

UniProtKB: Probable bacteriophage protein

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationNameFormula
10.0 mMTris
130.0 mMSaltNaClSodium chloride
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Details: Baked, glow discharged for 4 min before use
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated defocus max: 3.4 µm / Calibrated defocus min: 1.1 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.16 µm / Nominal defocus min: 2.16 µm / Nominal magnification: 130000
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Lower energy threshold: -10 eV / Energy filter - Upper energy threshold: 10 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 80.0 K / Max: 81.0 K
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 3-13 / Number grids imaged: 1 / Number real images: 7331 / Average exposure time: 10.0 sec. / Average electron dose: 80.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Segment selectionNumber selected: 15684
Startup modelType of model: NONE
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 1.4)
Final reconstructionNumber classes used: 1
Applied symmetry - Helical parameters - Δz: 39.46 Å
Applied symmetry - Helical parameters - Δ&Phi: 18.30 °
Applied symmetry - Helical parameters - Axial symmetry: C6 (6 fold cyclic)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Software - details: +IHRSR / Number images used: 15487

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-6pyt:
CryoEM Structure of Pyocin R2 - precontracted - trunk

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