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- EMDB-43023: 60S ribosome biogenesis intermediate (Dbp10 catalytic structure -... -

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Basic information

Entry
Database: EMDB / ID: EMD-43023
Title60S ribosome biogenesis intermediate (Dbp10 catalytic structure - Low-pass filtered locally refined map)
Map dataLow pass filtered map of locally refined map (Dbp10 catalytic state)
Sample
  • Complex: 60S ribosome biogenesis intermediate
    • Protein or peptide: ATP-dependent RNA helicase DBP10
KeywordsRNA Binding / RNA BINDING PROTEIN
Function / homology
Function and homology information


maturation of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / preribosome, large subunit precursor / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / assembly of large subunit precursor of preribosome / rRNA processing / RNA helicase activity / RNA helicase / nucleolus / ATP hydrolysis activity / RNA binding ...maturation of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / preribosome, large subunit precursor / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / assembly of large subunit precursor of preribosome / rRNA processing / RNA helicase activity / RNA helicase / nucleolus / ATP hydrolysis activity / RNA binding / ATP binding / identical protein binding
Similarity search - Function
DBP10, C-terminal / DDX54/DBP10 family / DBP10CT (NUC160) domain / DBP10CT (NUC160) domain / : / DEAD-box subfamily ATP-dependent helicases signature. / RNA helicase, DEAD-box type, Q motif / ATP-dependent RNA helicase DEAD-box, conserved site / DEAD-box RNA helicase Q motif profile. / DEAD/DEAH box helicase domain ...DBP10, C-terminal / DDX54/DBP10 family / DBP10CT (NUC160) domain / DBP10CT (NUC160) domain / : / DEAD-box subfamily ATP-dependent helicases signature. / RNA helicase, DEAD-box type, Q motif / ATP-dependent RNA helicase DEAD-box, conserved site / DEAD-box RNA helicase Q motif profile. / DEAD/DEAH box helicase domain / DEAD/DEAH box helicase / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ATP-dependent RNA helicase DBP10
Similarity search - Component
Biological speciesSaccharomyces cerevisiae BY4741 (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsCruz VE / Weirich CS / Peddada N / Erzberger JP
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R01GM135617-01 United States
Robert A. Welch FoundationI-1897 United States
Cancer Prevention and Research Institute of Texas (CPRIT)RR150074 United States
CitationJournal: Nat Commun / Year: 2024
Title: The DEAD-box ATPase Dbp10/DDX54 initiates peptidyl transferase center formation during 60S ribosome biogenesis.
Authors: Victor E Cruz / Christine S Weirich / Nagesh Peddada / Jan P Erzberger /
Abstract: DEAD-box ATPases play crucial roles in guiding rRNA restructuring events during the biogenesis of large (60S) ribosomal subunits, but their precise molecular functions are currently unknown. In this ...DEAD-box ATPases play crucial roles in guiding rRNA restructuring events during the biogenesis of large (60S) ribosomal subunits, but their precise molecular functions are currently unknown. In this study, we present cryo-EM reconstructions of nucleolar pre-60S intermediates that reveal an unexpected, alternate secondary structure within the nascent peptidyl-transferase-center (PTC). Our analysis of three sequential nucleolar pre-60S intermediates reveals that the DEAD-box ATPase Dbp10/DDX54 remodels this alternate base pairing and enables the formation of the rRNA junction that anchors the mature form of the universally conserved PTC A-loop. Post-catalysis, Dbp10 captures rRNA helix H61, initiating the concerted exchange of biogenesis factors during late nucleolar 60S maturation. Our findings show that Dbp10 activity is essential for the formation of the ribosome active site and reveal how this function is integrated with subsequent assembly steps to drive the biogenesis of the large ribosomal subunit.
History
DepositionDec 4, 2023-
Header (metadata) releaseMay 1, 2024-
Map releaseMay 1, 2024-
UpdateMay 1, 2024-
Current statusMay 1, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43023.png

Downloads & links

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Map

FileDownload / File: emd_43023.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLow pass filtered map of locally refined map (Dbp10 catalytic state)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 400 pix.
= 432. Å		1.08 Å/pix.
x 400 pix.
= 432. Å		1.08 Å/pix.
x 400 pix.
= 432. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.002
Minimum - Maximum-0.015224732 - 0.022578854
Average (Standard dev.)0.000019663521 (±0.0005902748)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 432.00003 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half Map A

Fileemd_43023_half_map_1.map
AnnotationHalf Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map B

Fileemd_43023_half_map_2.map
AnnotationHalf 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 : 60S ribosome biogenesis intermediate

EntireName: 60S ribosome biogenesis intermediate
Components
  • Complex: 60S ribosome biogenesis intermediate
    • Protein or peptide: ATP-dependent RNA helicase DBP10

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Supramolecule #1: 60S ribosome biogenesis intermediate

SupramoleculeName: 60S ribosome biogenesis intermediate / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast)

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Macromolecule #1: ATP-dependent RNA helicase DBP10

MacromoleculeName: ATP-dependent RNA helicase DBP10 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast)
Molecular weightTheoretical: 113.126469 KDa
SequenceString: MAGVQKRKRD LEDQDDNGSE EDDIAFDIAN EIALNDSESD ANDSDSEVEA DYGPNDVQDV IEYSSDEEEG VNNKKKAENK DIKKKKNSK KEIAAFPMLE MSDDENNASG KTQTGDDEDD VNEYFSTNNL EKTKHKKGSF PSFGLSKIVL NNIKRKGFRQ P TPIQRKTI ...String:
MAGVQKRKRD LEDQDDNGSE EDDIAFDIAN EIALNDSESD ANDSDSEVEA DYGPNDVQDV IEYSSDEEEG VNNKKKAENK DIKKKKNSK KEIAAFPMLE MSDDENNASG KTQTGDDEDD VNEYFSTNNL EKTKHKKGSF PSFGLSKIVL NNIKRKGFRQ P TPIQRKTI PLILQSRDIV GMARTGSGKT AAFILPMVEK LKSHSGKIGA RAVILSPSRE LAMQTFNVFK DFARGTELRS VL LTGGDSL EEQFGMMMTN PDVIIATPGR FLHLKVEMNL DLKSVEYVVF DEADRLFEMG FQEQLNELLA SLPTTRQTLL FSA TLPNSL VDFVKAGLVN PVLVRLDAET KVSENLEMLF LSSKNADREA NLLYILQEII KIPLATSEQL QKLQNSNNEA DSDS DDEND RQKKRRNFKK EKFRKQKMPA ANELPSEKAT ILFVPTRHHV EYISQLLRDC GYLISYIYGT LDQHARKRQL YNFRA GLTS ILVVTDVAAR GVDIPMLANV INYTLPGSSK IFVHRVGRTA RAGNKGWAYS IVAENELPYL LDLELFLGKK ILLTPM YDS LVDVMKKRWI DEGKPEYQFQ PPKLSYTKRL VLGSCPRLDV EGLGDLYKNL MSSNFDLQLA KKTAMKAEKL YYRTRTS AS PESLKRSKEI ISSGWDAQNA FFGKNEEKEK LDFLAKLQNR RNKETVFEFT RNPDDEMAVF MKRRRKQLAP IQRKATER R ELLEKERMAG LSHSIEDEIL KGDDGETGYT VSEDALKEFE DADQLLEAQE NENKKKKKPK SFKDPTFFLS HYAPAGDIQ DKQLQITNGF ANDAAQAAYD LNSDDKVQVH KQTATVKWDK KRKKYVNTQG IDNKKYIIGE SGQKIAASFR SGRFDDWSKA RNLKPLKVG SRETSIPSNL LEDPSQGPAA NGRTVRGKFK HKQMKAPKMP DKHRDNYYSQ KKKVEKALQS GISVKGYNNA P GLRSELKS TEQIRKDRII AEKKRAKNAR PSKKRKF

UniProtKB: ATP-dependent RNA helicase DBP10

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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

Concentration0.5 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
50.0 mMBis-Tris-Propane
125.0 mMSodium ChlorideNaCl
25.0 mMPotassium ChlorideKCl
10.0 mMMagnesium ChlorideMgCl2
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 39.3 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.9 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6em1

Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4)
Details: High-resolution map was low pass filtered to 6A to allow domain docking
Number images used: 195000
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)

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