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- EMDB-42565: Focus refined map of HflXr bound to the Listeria innocua ribosome... -

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Basic information

Entry
Database: EMDB / ID: EMD-42565
TitleFocus refined map of HflXr bound to the Listeria innocua ribosome in complex with HPF and E-site tRNA (structure II-B)
Map dataFocus refined map of HflXr from Structure II-B (Main map)
Sample
  • Complex: Listeria innocua 70S ribosome in complex with HPF, HflXr and E-site tRNA
Keywordscryo-EM / recycling / HflXr / HPF / time-resolved / RIBOSOME
Biological speciesListeria innocua (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsSeely SM / Basu RS / Gagnon MG
Funding support United States, 4 items
OrganizationGrant numberCountry
Other governmentUniversity of Texas Medical Branch Startup Funds United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM136936 United States
Welch FoundationH-2032-20230405 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)T32-GM008280 United States
CitationJournal: Nucleic Acids Res / Year: 2024
Title: Mechanistic insights into the alternative ribosome recycling by HflXr.
Authors: Savannah M Seely / Ritwika S Basu / Matthieu G Gagnon /
Abstract: During stress conditions such as heat shock and antibiotic exposure, ribosomes stall on messenger RNAs, leading to inhibition of protein synthesis. To remobilize ribosomes, bacteria use rescue ...During stress conditions such as heat shock and antibiotic exposure, ribosomes stall on messenger RNAs, leading to inhibition of protein synthesis. To remobilize ribosomes, bacteria use rescue factors such as HflXr, a homolog of the conserved housekeeping GTPase HflX that catalyzes the dissociation of translationally inactive ribosomes into individual subunits. Here we use time-resolved cryo-electron microscopy to elucidate the mechanism of ribosome recycling by Listeria monocytogenes HflXr. Within the 70S ribosome, HflXr displaces helix H69 of the 50S subunit and induces long-range movements of the platform domain of the 30S subunit, disrupting inter-subunit bridges B2b, B2c, B4, B7a and B7b. Our findings unveil a unique ribosome recycling strategy by HflXr which is distinct from that mediated by RRF and EF-G. The resemblance between HflXr and housekeeping HflX suggests that the alternative ribosome recycling mechanism reported here is universal in the prokaryotic kingdom.
History
DepositionOct 31, 2023-
Header (metadata) releaseFeb 28, 2024-
Map releaseFeb 28, 2024-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42565.png

Downloads & links

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Map

FileDownload / File: emd_42565.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFocus refined map of HflXr from Structure II-B (Main map)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 512 pix.
= 435.2 Å		0.85 Å/pix.
x 512 pix.
= 435.2 Å		0.85 Å/pix.
x 512 pix.
= 435.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.21
Minimum - Maximum-0.70163083 - 1.404609
Average (Standard dev.)-0.012902381 (±0.03487016)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 435.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_42565_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Focus refined map of HflXr from Structure II-B (Sharpened map)

Fileemd_42565_additional_1.map
AnnotationFocus refined map of HflXr from Structure II-B (Sharpened map)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Focus refined map of HflXr from Structure II-B (Half map)

Fileemd_42565_half_map_1.map
AnnotationFocus refined map of HflXr from Structure II-B (Half map)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Focus refined map of HflXr from Structure II-B (Half map)

Fileemd_42565_half_map_2.map
AnnotationFocus refined map of HflXr from Structure II-B (Half map)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Listeria innocua 70S ribosome in complex with HPF, HflXr and E-si...

EntireName: Listeria innocua 70S ribosome in complex with HPF, HflXr and E-site tRNA
Components
  • Complex: Listeria innocua 70S ribosome in complex with HPF, HflXr and E-site tRNA

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Supramolecule #1: Listeria innocua 70S ribosome in complex with HPF, HflXr and E-si...

SupramoleculeName: Listeria innocua 70S ribosome in complex with HPF, HflXr and E-site tRNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#52
Source (natural)Organism: Listeria innocua (bacteria)
Molecular weightTheoretical: 2.5 MDa

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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

BufferpH: 7.6
GridModel: Quantifoil R2/1 / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 295 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: INTEGRATING / Number grids imaged: 1 / Number real images: 10151 / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 96000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.1.2) / Number images used: 27543
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1.2)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.1.2)

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Atomic model buiding 1

Initial modelPDB ID:

7nhn


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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