[English] 日本語
Yorodumi
- EMDB-5389: An RNA Degradation Machine Sculpted by Ro Autoantigen and Noncodi... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-5389
TitleAn RNA Degradation Machine Sculpted by Ro Autoantigen and Noncoding RNA
Map datanegative stain single particle reconstruction of GraFix-prepared Rsr/Y RNA/PNPase complex
Sample
  • Sample: Rsr/Y RNA/PNPase complex
  • Protein or peptide: Ro sixty-related
  • Protein or peptide: polynucleotide phosphorylase
  • RNA: Y RNA
Keywordsexonuclease / Y RNA / degradation machine
Biological speciesDeinococcus radiodurans (radioresistant)
Methodsingle particle reconstruction / negative staining / Resolution: 25.0 Å
AuthorsChen X / Taylor DW / Wang HW / Wolin SL
CitationJournal: Cell / Year: 2013
Title: An RNA degradation machine sculpted by Ro autoantigen and noncoding RNA.
Authors: Xinguo Chen / David W Taylor / Casey C Fowler / Jorge E Galan / Hong-Wei Wang / Sandra L Wolin /
Abstract: Many bacteria contain an ortholog of the Ro autoantigen, a ring-shaped protein that binds noncoding RNAs (ncRNAs) called Y RNAs. In the only studied bacterium, Deinococcus radiodurans, the Ro ...Many bacteria contain an ortholog of the Ro autoantigen, a ring-shaped protein that binds noncoding RNAs (ncRNAs) called Y RNAs. In the only studied bacterium, Deinococcus radiodurans, the Ro ortholog Rsr functions in heat-stress-induced ribosomal RNA (rRNA) maturation and starvation-induced rRNA decay. However, the mechanism by which this conserved protein and its associated ncRNAs act has been obscure. We report that Rsr and the exoribonuclease polynucleotide phosphorylase (PNPase) form an RNA degradation machine that is scaffolded by Y RNA. Single-particle electron microscopy, followed by docking of atomic models into the reconstruction, suggests that Rsr channels single-stranded RNA into the PNPase cavity. Biochemical assays reveal that Rsr and Y RNA adapt PNPase for effective degradation of structured RNAs. A Ro ortholog and ncRNA also associate with PNPase in Salmonella Typhimurium. Our studies identify another ribonucleoprotein machine and demonstrate that ncRNA, by tethering a protein cofactor, can alter the substrate specificity of an enzyme.
History
DepositionFeb 6, 2012-
Header (metadata) releaseFeb 10, 2012-
Map releaseApr 3, 2013-
UpdateApr 10, 2013-
Current statusApr 10, 2013Processing site: RCSB / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.67
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 3.67
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5389_1.png

Downloads & links

-
Map

FileDownload / File: emd_5389.map.gz / Format: CCP4 / Size: 1.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationnegative stain single particle reconstruction of GraFix-prepared Rsr/Y RNA/PNPase complex
Voxel sizeX=Y=Z: 4.36 Å
Density
Contour LevelBy AUTHOR: 3.67 / Movie #1: 3.67
Minimum - Maximum-3.97276688 - 14.19131088
Average (Standard dev.)0.0 (±0.99999869)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions727272
Spacing727272
CellA=B=C: 313.92 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.364.364.36
M x/y/z727272
origin x/y/z0.0000.0000.000
length x/y/z313.920313.920313.920
α/β/γ90.00090.00090.000
start NX/NY/NZ-62-62-62
NX/NY/NZ125125125
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS727272
D min/max/mean-3.97314.191-0.000

-
Supplemental data

-
Sample components

-
Entire : Rsr/Y RNA/PNPase complex

EntireName: Rsr/Y RNA/PNPase complex
Components
  • Sample: Rsr/Y RNA/PNPase complex
  • Protein or peptide: Ro sixty-related
  • Protein or peptide: polynucleotide phosphorylase
  • RNA: Y RNA

-
Supramolecule #1000: Rsr/Y RNA/PNPase complex

SupramoleculeName: Rsr/Y RNA/PNPase complex / type: sample / ID: 1000 / Details: The sample was monodisperse and homogeneous.
Oligomeric state: one Rsr-Y RNA complex binds to one PNPase trimer
Number unique components: 3
Molecular weightExperimental: 375 KDa / Theoretical: 365 KDa / Method: glycerol gradient

-
Macromolecule #1: Ro sixty-related

MacromoleculeName: Ro sixty-related / type: protein_or_peptide / ID: 1 / Name.synonym: Rsr, Ro 60 kDa autoantigen / Details: His-tag Rsr / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Deinococcus radiodurans (radioresistant)
Molecular weightTheoretical: 62 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: RSFDuet-1

-
Macromolecule #3: polynucleotide phosphorylase

MacromoleculeName: polynucleotide phosphorylase / type: protein_or_peptide / ID: 3 / Name.synonym: PNPase / Details: Strep-tag PNPase / Number of copies: 3 / Oligomeric state: Trimer / Recombinant expression: Yes
Source (natural)Organism: Deinococcus radiodurans (radioresistant)
Molecular weightTheoretical: 262 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant plasmid: RSFDuet-1

-
Macromolecule #2: Y RNA

MacromoleculeName: Y RNA / type: rna / ID: 2
Details: Y RNA generated with correct 3' end using hammerhead ribozyme
Classification: OTHER / Structure: OTHER / Synthetic?: No
Source (natural)Organism: Deinococcus radiodurans (radioresistant)
Molecular weightTheoretical: 42 KDa

-
Experimental details

-
Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.1 mg/mL
BufferpH: 7.5
Details: 20 mM HEPES, 50 mM NaCl, 2 mM beta-mercaptoethanol, 1 mM MgCl2, 1 mM MnCl2, 1 mM Petabloc
StainingType: NEGATIVE
Details: Grids with adsorbed protein were stained consecutively with 3 droplets of 2% w/v uranyl acetate for 10 seconds each.
GridDetails: Homemade holey carbon grids with a thin layer of carbon over the holes
VitrificationCryogen name: NONE / Instrument: OTHER

-
Electron microscopy

MicroscopeFEI TECNAI 12
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.3 mm / Nominal defocus max: 1.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 49000
Sample stageSpecimen holder model: OTHER
Alignment procedureLegacy - Astigmatism: object lens astigmatism was corrected at 42,000 magnification
DateJul 12, 2009
Image recordingCategory: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number real images: 40 / Average electron dose: 20 e/Å2

-
Image processing

CTF correctionDetails: Each particle in IMAGIC
Final two d classificationNumber classes: 50
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 25.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: LEGINON, EMAN2, SPARX / Number images used: 9000
DetailsThe particles were selected manually using boxer in EMAN.

-
Atomic model buiding 1

Initial modelPDB ID:

1e3p


Chain - Chain ID: A
SoftwareName: Chimera
DetailsProtocol: Rigid body using Fit-in-Map. A trimer of the crystal structure of PNPase was created using COOT and used for fitting.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

-
Atomic model buiding 2

Initial modelPDB ID:

1yvp


Chain - #0 - Chain ID: B / Chain - #1 - Chain ID: E / Chain - #2 - Chain ID: F / Chain - #3 - Chain ID: H
SoftwareName: Chimera
DetailsProtocol: Rigid body using Fit-in-Map. Combined with the double-stranded portion of the misfolded substrate RNA from PDB 2I91.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

-
Atomic model buiding 3

Initial modelPDB ID:

2i91


Chain - #0 - Chain ID: C / Chain - #1 - Chain ID: D
SoftwareName: Chimera
DetailsProtocol: Rigid body using Fit-in-Map. The double-stranded portion of the misfolded substrate RNA from this structure was combined with PDB 1YVP to create a model of Rsr bound by Y RNA and a full substrate RNA with single-stranded tail.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more