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- EMDB-21670: Large ribosomal subunit of Halococcus morrhuae, an archaeon with ... -

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Basic information

Entry
Database: EMDB / ID: EMD-21670
TitleLarge ribosomal subunit of Halococcus morrhuae, an archaeon with an unusual 5S rRNA insertion
Map dataFinal map of Halococcus morrhuae large subunit
Sample
  • Complex: Large ribosomal subunit of Halococcus morrhuae
Biological speciesHalococcus morrhuae (archaea)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.2 Å
AuthorsTirumalai MR / Kaelber JT / Fox GE
Funding support United States, 4 items
OrganizationGrant numberCountry
National Aeronautic Space Administration (NASA, United States)NNX14AK36G United States
National Aeronautic Space Administration (NASA, United States)80NSSC18K1139 United States
National Aeronautic Space Administration (NASA, United States)NNX14AK16G United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM103832 United States
CitationJournal: FEBS Open Bio / Year: 2020
Title: Cryo-electron microscopy visualization of a large insertion in the 5S ribosomal RNA of the extremely halophilic archaeon Halococcus morrhuae.
Authors: Madhan R Tirumalai / Jason T Kaelber / Donghyun R Park / Quyen Tran / George E Fox /
Abstract: The extreme halophile Halococcus morrhuae (ATCC 17082) contains a 108-nucleotide insertion in its 5S rRNA. Large rRNA expansions in Archaea are rare. This one almost doubles the length of the 5S rRNA. ...The extreme halophile Halococcus morrhuae (ATCC 17082) contains a 108-nucleotide insertion in its 5S rRNA. Large rRNA expansions in Archaea are rare. This one almost doubles the length of the 5S rRNA. In order to understand how such an insertion is accommodated in the ribosome, we obtained a cryo-electron microscopy reconstruction of the native large subunit at subnanometer resolution. The insertion site forms a four-way junction that fully preserves the canonical 5S rRNA structure. Moving away from the junction site, the inserted region is conformationally flexible and does not pack tightly against the large subunit. The high-salt requirement of the H. morrhuae ribosomes for their stability conflicted with the low-salt threshold for cryo-electron microscopy procedures. Despite this obstacle, this is the first cryo-electron microscopy map of Halococcus ribosomes.
History
DepositionApr 7, 2020-
Header (metadata) releaseSep 16, 2020-
Map releaseSep 16, 2020-
UpdateOct 14, 2020-
Current statusOct 14, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.62
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.62
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21670.png

Downloads & links

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Map

FileDownload / File: emd_21670.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFinal map of Halococcus morrhuae large subunit
Voxel sizeX=Y=Z: 1.74 Å
Density
Contour LevelBy AUTHOR: 0.62 / Movie #1: 0.62
Minimum - Maximum-0.29695523 - 1.6128983
Average (Standard dev.)0.00093241315 (±0.10554897)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 417.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.741.741.74
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z417.600417.600417.600
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ161186271
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.2971.6130.001

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Supplemental data

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Mask #1

Fileemd_21670_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unmasked unfiltered half-map A

Fileemd_21670_half_map_1.map
AnnotationUnmasked unfiltered half-map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unmasked unfiltered half-map B

Fileemd_21670_half_map_2.map
AnnotationUnmasked unfiltered half-map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Large ribosomal subunit of Halococcus morrhuae

EntireName: Large ribosomal subunit of Halococcus morrhuae
Components
  • Complex: Large ribosomal subunit of Halococcus morrhuae

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Supramolecule #1: Large ribosomal subunit of Halococcus morrhuae

SupramoleculeName: Large ribosomal subunit of Halococcus morrhuae / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Halococcus morrhuae (archaea) / Strain: ATCC 17082
Molecular weightTheoretical: 1.5 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
Component:
ConcentrationFormulaName
3.4 molarKClpotassium chloride
60.0 millimolarMgC2H3O2magnesium acetate
30.0 millimolarNH2C(CH2OH)3HClTris HCl
7.0 millimolarHOCH2CH2SHmercaptoethanol

Details: pure water was added to the sample on the grid
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV
Details: 2 microliters of sample were applied to the grid, then 6 microliters of pure water was applied to the reverse face of the grid..

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Electron microscopy

MicroscopeJEOL 3200FSC
Specialist opticsEnergy filter - Name: In-column Omega Filter
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Frames/image: 1-40 / Number grids imaged: 2 / Number real images: 902 / Average exposure time: 8.0 sec. / Average electron dose: 26.0 e/Å2 / Details: some images collected manually (no serialEM)
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 5.716 µm / Calibrated defocus min: 0.915 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 4.1 mm / Nominal magnification: 20000
Sample stageSpecimen holder model: JEOL 3200FSC CRYOHOLDER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 140367
CTF correctionSoftware - Name: cryoSPARC (ver. 2.14.2)
Startup modelType of model: NONE / Details: cryoSPARC ab initio reconstruction
Final reconstructionNumber classes used: 22 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 6.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.14.2) / Details: masked FSC calculated automatically in cryoSPARC / Number images used: 99337
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 2.14.2)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 2.14.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

4v9f

RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: correlation coefficient

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