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- EMDB-1250: Structure of the E. coli signal recognition particle bound to a t... -

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Basic information

Entry
Database: EMDB / ID: EMD-1250
TitleStructure of the E. coli signal recognition particle bound to a translating ribosome.
Map dataThis is the map file of the E. coli signal recognition particle bound to a ribosome nascent chain complex.
Sample
  • Sample: E. coli ribosome nascent chain complexes and signal recognition particle
  • Complex: E.coli 70S ribosome
  • RNA: A-site tRNA
  • RNA: P-site tRNA
  • RNA: E-site tRNA
  • Ligand: E. coli SRP
Function / homology
Function and homology information


signal recognition particle / signal-recognition-particle GTPase / 7S RNA binding / SRP-dependent cotranslational protein targeting to membrane / GTPase activity / GTP binding / ATP hydrolysis activity
Similarity search - Function
Signal recognition particle protein / SRP/SRP receptor, N-terminal / Signal recognition particle, SRP54 subunit / Signal recognition particle, SRP54 subunit, M-domain / Signal recognition particle, SRP54 subunit, M-domain superfamily / Signal peptide binding domain / SRP54-type proteins GTP-binding domain signature. / Signal recognition particle SRP54, helical bundle / Signal recognition particle SRP54, N-terminal domain superfamily / SRP54-type protein, helical bundle domain ...Signal recognition particle protein / SRP/SRP receptor, N-terminal / Signal recognition particle, SRP54 subunit / Signal recognition particle, SRP54 subunit, M-domain / Signal recognition particle, SRP54 subunit, M-domain superfamily / Signal peptide binding domain / SRP54-type proteins GTP-binding domain signature. / Signal recognition particle SRP54, helical bundle / Signal recognition particle SRP54, N-terminal domain superfamily / SRP54-type protein, helical bundle domain / SRP54-type protein, helical bundle domain / Signal recognition particle, SRP54 subunit, GTPase domain / SRP54-type protein, GTPase domain / SRP54-type protein, GTPase domain / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Signal recognition particle 54 kDa protein / Signal recognition particle protein
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 15.5 Å
AuthorsSchaffitzel C / Oswald M / Berger I / Ishikawa T / Abrahams JP / Koerten HK / Koning RI / Ban N
CitationJournal: Nature / Year: 2006
Title: Structure of the E. coli signal recognition particle bound to a translating ribosome.
Authors: Christiane Schaffitzel / Miro Oswald / Imre Berger / Takashi Ishikawa / Jan Pieter Abrahams / Henk K Koerten / Roman I Koning / Nenad Ban /
Abstract: The prokaryotic signal recognition particle (SRP) targets membrane proteins into the inner membrane. It binds translating ribosomes and screens the emerging nascent chain for a hydrophobic signal ...The prokaryotic signal recognition particle (SRP) targets membrane proteins into the inner membrane. It binds translating ribosomes and screens the emerging nascent chain for a hydrophobic signal sequence, such as the transmembrane helix of inner membrane proteins. If such a sequence emerges, the SRP binds tightly, allowing the SRP receptor to lock on. This assembly delivers the ribosome-nascent chain complex to the protein translocation machinery in the membrane. Using cryo-electron microscopy and single-particle reconstruction, we obtained a 16 A structure of the Escherichia coli SRP in complex with a translating E. coli ribosome containing a nascent chain with a transmembrane helix anchor. We also obtained structural information on the SRP bound to an empty E. coli ribosome. The latter might share characteristics with a scanning SRP complex, whereas the former represents the next step: the targeting complex ready for receptor binding. High-resolution structures of the bacterial ribosome and of the bacterial SRP components are available, and their fitting explains our electron microscopic density. The structures reveal the regions that are involved in complex formation, provide insight into the conformation of the SRP on the ribosome and indicate the conformational changes that accompany high-affinity SRP binding to ribosome nascent chain complexes upon recognition of the signal sequence.
History
DepositionJul 28, 2006-
Header (metadata) releaseJul 29, 2006-
Map releaseJul 29, 2008-
UpdateMar 13, 2013-
Current statusMar 13, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.35
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.35
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-2iy3
  • Surface level: 0.35
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-2iy3
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1250.gif

Downloads & links

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Map

FileDownload / File: emd_1250.map.gz / Format: CCP4 / Size: 5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThis is the map file of the E. coli signal recognition particle bound to a ribosome nascent chain complex.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
3.81 Å/pix.
x 110 pix.
= 419.1 Å		3.81 Å/pix.
x 110 pix.
= 419.1 Å		3.81 Å/pix.
x 110 pix.
= 419.1 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 3.81 Å
Density

Histogram

Histogram (log scale)
Contour Level1: 0.425 / Movie #1: 0.35
Minimum - Maximum-2.11013 - 3.62066
Average (Standard dev.)0.00000000003178 (±0.322362)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions110110110
Spacing110110110
CellA=B=C: 419.1 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.813.813.81
M x/y/z110110110
origin x/y/z0.0000.0000.000
length x/y/z419.100419.100419.100
α/β/γ90.00090.00090.000
start NX/NY/NZ-64-64-64
NX/NY/NZ128128128
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS110110110
D min/max/mean-2.1103.6210.000

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Supplemental data

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Sample components

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Entire : E. coli ribosome nascent chain complexes and signal recognition p...

EntireName: E. coli ribosome nascent chain complexes and signal recognition particle
Components
  • Sample: E. coli ribosome nascent chain complexes and signal recognition particle
  • Complex: E.coli 70S ribosome
  • RNA: A-site tRNA
  • RNA: P-site tRNA
  • RNA: E-site tRNA
  • Ligand: E. coli SRP

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Supramolecule #1000: E. coli ribosome nascent chain complexes and signal recognition p...

SupramoleculeName: E. coli ribosome nascent chain complexes and signal recognition particle
type: sample / ID: 1000 / Oligomeric state: One SRP binds to one ribosome. / Number unique components: 5

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Supramolecule #1: E.coli 70S ribosome

SupramoleculeName: E.coli 70S ribosome / type: complex / ID: 1 / Recombinant expression: No / Ribosome-details: ribosome-prokaryote: ALL
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 2.5 MDa

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Macromolecule #1: A-site tRNA

MacromoleculeName: A-site tRNA / type: rna / ID: 1 / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 25 KDa

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Macromolecule #2: P-site tRNA

MacromoleculeName: P-site tRNA / type: rna / ID: 2 / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 25 KDa

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Macromolecule #3: E-site tRNA

MacromoleculeName: E-site tRNA / type: rna / ID: 3 / Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 25 KDa

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Macromolecule #4: E. coli SRP

MacromoleculeName: E. coli SRP / type: ligand / ID: 4 / Name.synonym: Signal recognition particle / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Escherichia coli (E. coli) / Location in cell: cytosol
Molecular weightTheoretical: 90 KDa
Recombinant expressionOrganism: BL21StarDE3 / Recombinant plasmid: pET24a_Ffh and pUC19_Ffs

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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

Concentration0.25 mg/mL
BufferpH: 7.5
Details: 50 mM Hepes-KOH pH 7.5, 100 mM KCl, 25 mM MgCl2, 1 mM DTT, 1 mM GTP
GridDetails: carbon-coated lacey formvar grids
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: HOMEMADE PLUNGER
Details: Vitrification instrument: home-built environmental chamber and vitrification device
Method: blot for 1.5 sec before plunging

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Electron microscopy

MicroscopeFEI TECNAI F20
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: OTHER / Number real images: 251 / Average electron dose: 10 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 51000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 50000
Sample stageSpecimen holder: cryo stage / Specimen holder model: GATAN LIQUID NITROGEN
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 15.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Spider / Number images used: 24382

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Atomic model buiding 1

Initial modelPDB ID:

1jpn


Chain - Chain ID: A
SoftwareName: NOMAD
DetailsProtocol: normal mode based flexible fitting. Fitting of SRP atomic model into isolated EM density
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: correlation coefficient
Output model

PDB-2iy3:
Structure of the E. Coli Signal Regognition Particle

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Atomic model buiding 2

Initial modelPDB ID:

1qzw


Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B
SoftwareName: NOMAD
DetailsProtocol: normal mode based flexible fitting. Fitting of SRP atomic model into isolated EM density
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: correlation coefficient
Output model

PDB-2iy3:
Structure of the E. Coli Signal Regognition Particle

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