+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-25474 | |||||||||
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Title | CryoEM structure of the N-Terminal deleted Rix7 AAA-ATPase | |||||||||
Map data | ||||||||||
Sample |
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Keywords | AAA-ATPase / ribosome biogenesis / substrate translocation / RIBOSOMAL PROTEIN | |||||||||
Function / homology | Function and homology information preribosome binding / ribosome biogenesis / ATP hydrolysis activity / RNA binding / ATP binding / nucleus Similarity search - Function | |||||||||
Biological species | Chaetomium thermophilum (fungus) / synthetic construct (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.88 Å | |||||||||
Authors | Kocaman S / Stanley RE | |||||||||
Funding support | United States, 1 items
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Citation | Journal: PNAS Nexus / Year: 2022 Title: Communication network within the essential AAA-ATPase Rix7 drives ribosome assembly. Authors: Seda Kocaman / Yu-Hua Lo / Juno M Krahn / Mack Sobhany / Venkata P Dandey / Matthew L Petrovich / Suhas K Etigunta / Jason G Williams / Leesa J Deterding / Mario J Borgnia / Robin E Stanley / Abstract: Rix7 is an essential AAA+ ATPase that functions during the early stages of ribosome biogenesis. Rix7 is composed of three domains including an N-terminal domain (NTD) and two AAA+ domains (D1 and ...Rix7 is an essential AAA+ ATPase that functions during the early stages of ribosome biogenesis. Rix7 is composed of three domains including an N-terminal domain (NTD) and two AAA+ domains (D1 and D2) that assemble into an asymmetric stacked hexamer. It was recently established that Rix7 is a presumed protein translocase that removes substrates from preribosomes by translocating them through its central pore. However, how the different domains of Rix7 coordinate their activities within the overall hexameric structure was unknown. We captured cryo-electron microscopy (EM) structures of single and double Walker B variants of full length Rix7. The disordered NTD was not visible in the cryo-EM reconstructions, but cross-linking mass spectrometry revealed that the NTD can associate with the central channel in vitro. Deletion of the disordered NTD enabled us to obtain a structure of the Rix7 hexamer to 2.9 Å resolution, providing high resolution details of critical motifs involved in substrate translocation and interdomain communication. This structure coupled with cell-based assays established that the linker connecting the D1 and D2 domains as well as the pore loops lining the central channel are essential for formation of the large ribosomal subunit. Together, our work shows that Rix7 utilizes a complex communication network to drive ribosome biogenesis. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_25474.map.gz | 8.6 MB | EMDB map data format | |
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Header (meta data) | emd-25474-v30.xml emd-25474.xml | 11.6 KB 11.6 KB | Display Display | EMDB header |
Images | emd_25474.png | 68.9 KB | ||
Filedesc metadata | emd-25474.cif.gz | 5.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-25474 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-25474 | HTTPS FTP |
-Validation report
Summary document | emd_25474_validation.pdf.gz | 539 KB | Display | EMDB validaton report |
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Full document | emd_25474_full_validation.pdf.gz | 538.6 KB | Display | |
Data in XML | emd_25474_validation.xml.gz | 4.4 KB | Display | |
Data in CIF | emd_25474_validation.cif.gz | 4.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-25474 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-25474 | HTTPS FTP |
-Related structure data
Related structure data | 7swlMC 7t0vC 7t3iC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_25474.map.gz / Format: CCP4 / Size: 10.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.0545 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : CryoEM structure of the N-terminal-deleted Rix7 AAA-ATPase
Entire | Name: CryoEM structure of the N-terminal-deleted Rix7 AAA-ATPase |
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Components |
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-Supramolecule #1: CryoEM structure of the N-terminal-deleted Rix7 AAA-ATPase
Supramolecule | Name: CryoEM structure of the N-terminal-deleted Rix7 AAA-ATPase type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 |
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Source (natural) | Organism: Chaetomium thermophilum (fungus) |
Molecular weight | Theoretical: 100 KDa |
-Macromolecule #1: Rix7
Macromolecule | Name: Rix7 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: Chaetomium thermophilum (fungus) |
Molecular weight | Theoretical: 69.008492 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MASMTGGQQM GRGSMSILDI AGVDDTLQRL LKEVWFPLRG GEACEKMGYR YDNGVLLHGP SGCGKTTLAH AIAGSIGVAF IPVSAPSVI GGTSGESEKN IRDVFDEAIR LAPCLIFLDQ IDAIAGRRES ANKGMESRIV AEIMNGMDRI RQNTPLGKNV V VLAATNRP ...String: MASMTGGQQM GRGSMSILDI AGVDDTLQRL LKEVWFPLRG GEACEKMGYR YDNGVLLHGP SGCGKTTLAH AIAGSIGVAF IPVSAPSVI GGTSGESEKN IRDVFDEAIR LAPCLIFLDQ IDAIAGRRES ANKGMESRIV AEIMNGMDRI RQNTPLGKNV V VLAATNRP EFLDPAIRRR FSVEIDMGMP SERAREQILR SLTRDLSLAD DINFKELAKM TPGYVGSDLQ YVVKAAVSES FQ ANIDSLL AQARAKHPAD HLANVSQPQR DWLLLEAHRD EEVSWPSTKI TMEQFRKAVS LVQPASKREG FSTIPDTTWS HVG ALEDVR KKLEMSIIGP IKNPELFTRV GIKPAAGILL WGPPGCGKTL VAKAVANESK ANFISIKGPE LLNKYVGESE RAVR QLFSR AKSSAPCILF FDQMDALVPR RDDSLSDASA RVVNTLLTEL DGVGDRSGIY VIGATNRPDM IDEAIRRPGR LGTSI YVGL PSAEDRVKIL KTLYRNTVKA PKKREGTNGE DVDMTDAAAE QQHQGTTDAD LEKVALDLRC TGFSGADLGN LMQAAA QAC LERVYTQRQQ KRKEGGSVAE EEEIEPVITM EDWEKALNEV KPSVKDPEKY MHSGFAAALE HHHHHH UniProtKB: AAA+ ATPase domain-containing protein |
-Macromolecule #2: polyleucine
Macromolecule | Name: polyleucine / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 2.733803 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: LLLLLLLLLL LLLLLLLLLL LLLL |
-Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 9 / Formula: ATP |
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Molecular weight | Theoretical: 507.181 Da |
Chemical component information | ChemComp-ATP: |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 9 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 5 / Number of copies: 1 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: PDB ENTRY |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 2.88 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 1200000 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |