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- EMDB-3285: Cryo-electron microscopy structure of ribosome-bound initiation f... -
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Basic information
Entry | Database: EMDB / ID: EMD-3285 | |||||||||
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Title | Cryo-electron microscopy structure of ribosome-bound initiation factor 2 | |||||||||
![]() | Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. | |||||||||
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![]() | Initiation Factor 2 / ribosome / translation / translation initiation / IF2 / translational GTPase / GTPase | |||||||||
Function / homology | ![]() guanosine tetraphosphate binding / stringent response / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / ribosomal small subunit binding / transcriptional attenuation / endoribonuclease inhibitor activity ...guanosine tetraphosphate binding / stringent response / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / ribosomal small subunit binding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / four-way junction DNA binding / translational termination / DnaA-L2 complex / chaperone-mediated protein folding / translation repressor activity / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / translation initiation factor activity / response to cold / mRNA regulatory element binding translation repressor activity / ribosome assembly / positive regulation of RNA splicing / assembly of large subunit precursor of preribosome / transcription elongation factor complex / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / DNA endonuclease activity / ribosomal large subunit assembly / transcription antitermination / response to reactive oxygen species / translational initiation / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / response to radiation / mRNA 5'-UTR binding / large ribosomal subunit / ribosome biogenesis / ribosome binding / regulation of translation / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / transferase activity / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / cytoplasmic translation / tRNA binding / molecular adaptor activity / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / translation / response to antibiotic / GTPase activity / negative regulation of DNA-templated transcription / mRNA binding / GTP binding / DNA binding / RNA binding / zinc ion binding / membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.6 Å | |||||||||
![]() | Sprink T / Ramrath DJF / Yamamoto H / Yamamoto K / Loerke J / Ismer J / Hildebrand PW / Scheerer P / Buerger J / Mielke T / Spahn CMT | |||||||||
![]() | ![]() Title: Structures of ribosome-bound initiation factor 2 reveal the mechanism of subunit association. Authors: Thiemo Sprink / David J F Ramrath / Hiroshi Yamamoto / Kaori Yamamoto / Justus Loerke / Jochen Ismer / Peter W Hildebrand / Patrick Scheerer / Jörg Bürger / Thorsten Mielke / Christian M T Spahn / ![]() Abstract: Throughout the four phases of protein biosynthesis-initiation, elongation, termination, and recycling-the ribosome is controlled and regulated by at least one specified translational guanosine ...Throughout the four phases of protein biosynthesis-initiation, elongation, termination, and recycling-the ribosome is controlled and regulated by at least one specified translational guanosine triphosphatase (trGTPase). Although the structural basis for trGTPase interaction with the ribosome has been solved for the last three steps of translation, the high-resolution structure for the key initiation trGTPase, initiation factor 2 (IF2), complexed with the ribosome, remains elusive. We determine the structure of IF2 complexed with a nonhydrolyzable guanosine triphosphate analog and initiator fMet-tRNAi (Met) in the context of the Escherichia coli ribosome to 3.7-Å resolution using cryo-electron microscopy. The structural analysis reveals previously unseen intrinsic conformational modes of the 70S initiation complex, establishing the mutual interplay of IF2 and initator transfer RNA (tRNA) with the ribsosome and providing the structural foundation for a mechanistic understanding of the final steps of translation initiation. | |||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 97.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 12.8 KB 12.8 KB | Display Display | ![]() |
Images | ![]() | 407.1 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3jcnMC ![]() 6559C ![]() 3jcjC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.23 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes...
Entire | Name: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. |
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Components |
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-Supramolecule #1000: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes...
Supramolecule | Name: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. type: sample / ID: 1000 / Number unique components: 3 |
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Molecular weight | Experimental: 2.5 MDa / Theoretical: 2.6 MDa |
-Supramolecule #1: 70S ribosome
Supramolecule | Name: 70S ribosome / type: complex / ID: 1 / Name.synonym: 70S Details: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. Recombinant expression: No / Ribosome-details: ribosome-prokaryote: ALL |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 2.5 MDa / Theoretical: 2.6 MDa |
-Macromolecule #1: fMet-tRNAiMet
Macromolecule | Name: fMet-tRNAiMet / type: rna / ID: 1 / Name.synonym: initiator tRNA Details: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. Classification: TRANSFER / Structure: DOUBLE HELIX / Synthetic?: No |
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Source (natural) | Organism: ![]() ![]() |
-Macromolecule #2: Initiation Factor 2
Macromolecule | Name: Initiation Factor 2 / type: protein_or_peptide / ID: 2 / Name.synonym: IF2 Details: Initiation Factor 2 was stalled on Escherichia coli 70S ribosomes by the non-hydrolysable GTP analogue GDPNP in the presence of fMet-tRNAiMet and mRNA. Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Experimental: 97 KDa / Theoretical: 97 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Translation initiation factor IF-2 |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 Details: 20 mM Hepes-KOH pH 7.5, 15 mM magnesium acetate, 150 mM potassium acetate, 4 mM -mercapthoethanol, 2 mM spermidine and 0.05 mM spermine |
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Grid | Details: Quantifoil R3-3 Cu 300 mesh with 2 nm carbon support film |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK I / Method: blot for 2-4 seconds before plunging |
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Electron microscopy #1
Microscopy ID | 1 |
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Microscope | FEI POLARA 300 |
Date | Aug 26, 2013 |
Image recording | Category: CCD / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number real images: 918 / Average electron dose: 20 e/Å2 / Details: Automated data collection using Leginon |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 39000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 7.18 µm / Nominal defocus min: 0.64 µm / Nominal magnification: 31000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Electron microscopy #2
Microscopy ID | 2 |
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Microscope | FEI POLARA 300 |
Date | Jun 10, 2015 |
Image recording | Category: CCD / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number real images: 2797 / Average electron dose: 20 e/Å2 / Details: Automated data collection using Leginon |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 39000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 7.57 µm / Nominal defocus min: 0.19 µm / Nominal magnification: 31000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
Details | To avoid overfitting, the data was refined in a resolution-limited scheme using SPIDER |
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CTF correction | Details: CTFFIND4 |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.6 Å / Resolution method: OTHER / Software - Name: EMAN2, CTFFIND4, SPIDER, SPARX Details: Final maps were calculated from two datasets. To avoid overfitting, the data was refined in a resolution-limited scheme using SPIDER Number images used: 14872 |