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- EMDB-24271: State E2 nucleolar 60S ribosomal biogenesis intermediate - Spb1-M... -

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Basic information

Entry
Database: EMDB / ID: EMD-24271
TitleState E2 nucleolar 60S ribosomal biogenesis intermediate - Spb1-MTD locally refined map
Map dataState E2 Spb1-MTD locally refined map
Sample
  • Complex: Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.
    • RNA: x 1 types
    • Protein or peptide: x 15 types
  • Ligand: x 2 types
Keywordsribosome biogenesis / DEAD-box ATPases / methyltransferase / nucleolus / RIBOSOME
Function / homology
Function and homology information


25S rRNA (cytosine2870-C5)-methyltransferase / 27S pre-rRNA (guanosine2922-2'-O)-methyltransferase / Noc1p-Noc2p complex / Noc2p-Noc3p complex / rRNA (guanosine-2'-O-)-methyltransferase activity / rRNA (uridine-2'-O-)-methyltransferase activity / rRNA (guanine) methyltransferase activity / rRNA (cytosine-C5-)-methyltransferase activity / rRNA base methylation / positive regulation of ATP-dependent activity ...25S rRNA (cytosine2870-C5)-methyltransferase / 27S pre-rRNA (guanosine2922-2'-O)-methyltransferase / Noc1p-Noc2p complex / Noc2p-Noc3p complex / rRNA (guanosine-2'-O-)-methyltransferase activity / rRNA (uridine-2'-O-)-methyltransferase activity / rRNA (guanine) methyltransferase activity / rRNA (cytosine-C5-)-methyltransferase activity / rRNA base methylation / positive regulation of ATP-dependent activity / maturation of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / rRNA methylation / maturation of 5.8S rRNA / Major pathway of rRNA processing in the nucleolus and cytosol / SRP-dependent cotranslational protein targeting to membrane / GTP hydrolysis and joining of the 60S ribosomal subunit / ribosomal large subunit binding / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / Formation of a pool of free 40S subunits / preribosome, large subunit precursor / ATPase activator activity / L13a-mediated translational silencing of Ceruloplasmin expression / translational elongation / ribosomal large subunit export from nucleus / regulation of translational fidelity / ribosomal subunit export from nucleus / maturation of LSU-rRNA / translation initiation factor activity / maturation of LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / assembly of large subunit precursor of preribosome / ribosomal large subunit biogenesis / cytosolic ribosome assembly / maintenance of translational fidelity / rRNA processing / protein transport / ATPase binding / large ribosomal subunit rRNA binding / ribosomal large subunit assembly / cytoplasmic translation / cytosolic large ribosomal subunit / structural constituent of ribosome / translation / GTPase activity / mRNA binding / GTP binding / nucleolus / mitochondrion / RNA binding / nucleoplasm / nucleus / cytosol / cytoplasm
Similarity search - Function
Nucleolar complex protein 2 / Noc2p family / RNA (C5-cytosine) methyltransferase, NOP2 / Nop2p / Bacterial Fmu (Sun)/eukaryotic nucleolar NOL1/Nop2p, conserved site / NOL1/NOP2/sun family signature. / Ribosomal RNA methyltransferase, SPB1-like, C-terminal / Ribosomal RNA methyltransferase Spb1, domain of unknown function DUF3381 / AdoMet-dependent rRNA methyltransferase SPB1-like / Spb1 C-terminal domain ...Nucleolar complex protein 2 / Noc2p family / RNA (C5-cytosine) methyltransferase, NOP2 / Nop2p / Bacterial Fmu (Sun)/eukaryotic nucleolar NOL1/Nop2p, conserved site / NOL1/NOP2/sun family signature. / Ribosomal RNA methyltransferase, SPB1-like, C-terminal / Ribosomal RNA methyltransferase Spb1, domain of unknown function DUF3381 / AdoMet-dependent rRNA methyltransferase SPB1-like / Spb1 C-terminal domain / Ribosomal RNA methyltransferase Spb1, DUF3381 / : / Ribosomal RNA large subunit methyltransferase E / Domain of unknown function DUF2423 / YBL028C ribosome biogenesis factor, N-terminal domain / SAM-dependent methyltransferase RsmB/NOP2-type / RNA (C5-cytosine) methyltransferase / : / 16S rRNA methyltransferase RsmB/F / SAM-dependent MTase RsmB/NOP-type domain profile. / Guanine nucleotide-binding protein-like 3, N-terminal domain / GNL3L/Grn1 putative GTPase / : / GTP-binding protein, orthogonal bundle domain superfamily / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain / Circularly permuted (CP)-type guanine nucleotide-binding (G) domain profile. / Translation initiation factor IF6 / eIF-6 family / translation initiation factor 6 / 50S ribosome-binding GTPase / GTP binding domain / metallochaperone-like domain / TRASH domain / Ribosomal RNA methyltransferase, FtsJ domain / FtsJ-like methyltransferase / Ribosomal protein L31e, conserved site / Ribosomal protein L31e signature. / Ribosomal protein L31e / Ribosomal protein L31e domain superfamily / Ribosomal protein L31e / Ribosomal_L31e / Ribosomal protein L24e-related / Ribosomal protein L24e/L24 superfamily / Ribosomal protein L24e / Ribosomal protein L3, domain 3, archaeal type superfamily / Ribosomal protein L3, archaeal/eukaryotic type / Ribosomal protein L14P, conserved site / Ribosomal protein L14 signature. / Ribosomal protein L14p/L23e / Ribosomal protein L14P / Ribosomal protein L14 superfamily / Ribosomal protein L14p/L23e / Ribosomal protein L3, conserved site / Ribosomal protein L3 signature. / Ribosomal protein L3 / Ribosomal protein L3 / Translation protein, beta-barrel domain superfamily / S-adenosyl-L-methionine-dependent methyltransferase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Large ribosomal subunit protein eL31A / Large ribosomal subunit protein uL14A / Large ribosomal subunit protein uL3 / 27S pre-rRNA (guanosine(2922)-2'-O)-methyltransferase / UPF0642 protein YBL028C / Nucleolar complex protein 2 / Nuclear GTP-binding protein NUG1 / 25S rRNA (cytosine(2870)-C(5))-methyltransferase / Ribosome biogenesis protein RLP24 / Eukaryotic translation initiation factor 6
Similarity search - Component
Biological speciesSaccharomyces cerevisiae BY4741 (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.13 Å
AuthorsCruz VE / Sekulski K / Peddada N / Erzberger JP
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM135617-01 United States
Cancer Prevention and Research Institute of Texas (CPRIT)RR150074 United States
Robert A. Welch FoundationI-1897 United States
CitationJournal: Nat Struct Mol Biol / Year: 2022
Title: Sequence-specific remodeling of a topologically complex RNP substrate by Spb4.
Authors: Victor Emmanuel Cruz / Kamil Sekulski / Nagesh Peddada / Carolin Sailer / Sahana Balasubramanian / Christine S Weirich / Florian Stengel / Jan P Erzberger /
Abstract: DEAD-box ATPases are ubiquitous enzymes essential in all aspects of RNA biology. However, the limited in vitro catalytic activities described for these enzymes are at odds with their complex cellular ...DEAD-box ATPases are ubiquitous enzymes essential in all aspects of RNA biology. However, the limited in vitro catalytic activities described for these enzymes are at odds with their complex cellular roles, most notably in driving large-scale RNA remodeling steps during the assembly of ribonucleoproteins (RNPs). We describe cryo-EM structures of 60S ribosomal biogenesis intermediates that reveal how context-specific RNA unwinding by the DEAD-box ATPase Spb4 results in extensive, sequence-specific remodeling of rRNA secondary structure. Multiple cis and trans interactions stabilize Spb4 in a post-catalytic, high-energy intermediate that drives the organization of the three-way junction at the base of rRNA domain IV. This mechanism explains how limited strand separation by DEAD-box ATPases is leveraged to provide non-equilibrium directionality and ensure efficient and accurate RNP assembly.
History
DepositionJun 21, 2021-
Header (metadata) releaseNov 9, 2022-
Map releaseNov 9, 2022-
UpdateJun 5, 2024-
Current statusJun 5, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24271.png

Downloads & links

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Map

FileDownload / File: emd_24271.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationState E2 Spb1-MTD locally refined map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 420 pix.
= 453.6 Å		1.08 Å/pix.
x 420 pix.
= 453.6 Å		1.08 Å/pix.
x 420 pix.
= 453.6 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: 0.018
Minimum - Maximum-0.048888553 - 0.10175384
Average (Standard dev.)0.000036645804 (±0.0018181521)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 453.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: State E2 Spb1-MTD locally refined half map 2

Fileemd_24271_half_map_1.map
AnnotationState E2 Spb1-MTD locally refined half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: State E2 Spb1-MTD locally refined half map 1

Fileemd_24271_half_map_2.map
AnnotationState E2 Spb1-MTD locally refined half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.

EntireName: Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.
Components
  • Complex: Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.
    • RNA: 25S rRNA
    • Protein or peptide: Ribosomal RNA-processing protein 17
    • Protein or peptide: Nucleolar complex protein 2
    • Protein or peptide: 60S ribosomal protein L23-A
    • Protein or peptide: Ribosome biogenesis protein RLP24
    • Protein or peptide: 27S pre-rRNA (guanosine(2922)-2'-O)-methyltransferase
    • Protein or peptide: 60S ribosome biogenesis factor Nog1
    • Protein or peptide: 60S ribosomal protein L31-A
    • Protein or peptide: 60S ribosomal protein L22-A
    • Protein or peptide: 60S ribosome ribosomal protein L19A
    • Protein or peptide: 60S ribosomal protein L30
    • Protein or peptide: 60S ribosomal protein L3
    • Protein or peptide: UPF0642 protein YBL028C
    • Protein or peptide: Nuclear GTP-binding protein NUG1
    • Protein or peptide: Eukaryotic translation initiation factor 6
    • Protein or peptide: 25S rRNA (cytosine(2870)-C(5))-methyltransferase
  • Ligand: S-ADENOSYL-L-HOMOCYSTEINE
  • Ligand: water

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Supramolecule #1: Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.

SupramoleculeName: Nucleolar 60S intermediate purified with tags on Ytm1 and Spb4.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#16
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast)
Molecular weightTheoretical: 3.3 MDa

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Macromolecule #1: 25S rRNA

MacromoleculeName: 25S rRNA / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 121.655477 KDa
SequenceString: AUAGGGUGAA GCCAGAGGAA ACUCUGGUGG AGGCUCGAAA AUGGAAUUAG ACUGAAGGUC GGCGCCCUGG AGUUAUCUUU UCUUGUUUA UCCCGCUUCG GCCCCAAGGU GAACAGCCUC UAAUGUAGAU AAGGGAAGUC GGGGGAAUCU GACUGUUAAC G AGAUUCCC ...String:
AUAGGGUGAA GCCAGAGGAA ACUCUGGUGG AGGCUCGAAA AUGGAAUUAG ACUGAAGGUC GGCGCCCUGG AGUUAUCUUU UCUUGUUUA UCCCGCUUCG GCCCCAAGGU GAACAGCCUC UAAUGUAGAU AAGGGAAGUC GGGGGAAUCU GACUGUUAAC G AGAUUCCC ACUGUCCCUA UCUACUCGGG GAACUGGCUU GUCGGCUCUU CCUAUCAUAC CAUUCGGUAA GCGUUGGAU (OMU)(OMG)(PSU)UCACCCA CUAAUAGGGA ACGUG(A2M)GCUG AAUUGAACUU AGGAACAGUU CAUUCGGAUA AUUG GUUUU UGCGGCUGUC UGAUCAGGCA UUGCCGCGAA GCUACCAUCC GAAUCUAUAU AUUGUAAGCU GAGAUUAAGC CU

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Macromolecule #2: Ribosomal RNA-processing protein 17

MacromoleculeName: Ribosomal RNA-processing protein 17 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 13.077044 KDa
SequenceString:
NRQILTRGKQ SKKFGTDEVT FDKDSRLDYL TGFHKRKLQR QKKAQEFIKE QERLRKIEER QKIRQERKEV MEEQLKTFKE SLNLKFRYL TKNERRINQR KANDNK

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Macromolecule #3: Nucleolar complex protein 2

MacromoleculeName: Nucleolar complex protein 2 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 4.74966 KDa
SequenceString:
KVSKSTKKFQ SKHLKHTLDQ RRKEKIQKKR IQGRRGNKT

UniProtKB: Nucleolar complex protein 2

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Macromolecule #4: 60S ribosomal protein L23-A

MacromoleculeName: 60S ribosomal protein L23-A / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 14.218625 KDa
SequenceString:
NGAQGTKFRI SLGLPVGAIM NCADNSGARN LYIIAVKGSG SRLNRLPAAS LGDMVMATVK KGKPELRKKV MPAIVVRQAK SWRRRDGVF LYFEDNAGVI ANPKGEMKGS AITGPVGKEC ADLWPRVASN SGVVV

UniProtKB: Large ribosomal subunit protein uL14A

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Macromolecule #5: Ribosome biogenesis protein RLP24

MacromoleculeName: Ribosome biogenesis protein RLP24 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 7.786305 KDa
SequenceString:
MRIYQCHFCS SPCYPGHGIM FVRNDAKEFR FCRSKCHKAF KQRRNPRKLK WTKAFRKAAG KELAV

UniProtKB: Ribosome biogenesis protein RLP24

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Macromolecule #6: 27S pre-rRNA (guanosine(2922)-2'-O)-methyltransferase

MacromoleculeName: 27S pre-rRNA (guanosine(2922)-2'-O)-methyltransferase / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
EC number: 27S pre-rRNA (guanosine2922-2'-O)-methyltransferase
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 45.405633 KDa
SequenceString: MGKTQKKNSK GRLDRYYYLA KEKGYRARSS FKIIQINEKY GHFLEKSKVV IDLCAAPGSW CQVASKLCPV NSLIIGVDIV PMKPMPNVI TFQSDITTED CRSKLRGYMK TWKADTVLHD GAPNVGLGWV QDAFTQSQLT LQALKLAVEN LVVNGTFVTK I FRSKDYNK ...String:
MGKTQKKNSK GRLDRYYYLA KEKGYRARSS FKIIQINEKY GHFLEKSKVV IDLCAAPGSW CQVASKLCPV NSLIIGVDIV PMKPMPNVI TFQSDITTED CRSKLRGYMK TWKADTVLHD GAPNVGLGWV QDAFTQSQLT LQALKLAVEN LVVNGTFVTK I FRSKDYNK LIWVFQQLFE KVEATKPPAS RNVSAEIFVV CKGFKAPKRL DPRLLDPKEV FEELPDGQQN MESKIYNPEK KV RKRQGYE EGDNLLYHET SILDFVRTED PISMLGEMNK FTIDENDHEW KILKKLKQTT DEFRSCIEDL KVLGKKDFKM ILR WRKIAR EILGIEVKDD AKTEIEVVPL TEEEQIEKDL QGLQEKQRLN VKRERRRKNE MKQKELQRMQ MNM

UniProtKB: 27S pre-rRNA (guanosine(2922)-2'-O)-methyltransferase

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Macromolecule #7: 60S ribosome biogenesis factor Nog1

MacromoleculeName: 60S ribosome biogenesis factor Nog1 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
EC number: 27S pre-rRNA (guanosine2922-2'-O)-methyltransferase
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 8.783554 KDa
SequenceString:
IVLNRTQRKT PTVIRPGFKI TRIRAFYMRK VKYTGEGMPE ILDGKNVYIR NRQKTMIAEA RNRKSLKNKA IMPR

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Macromolecule #8: 60S ribosomal protein L31-A

MacromoleculeName: 60S ribosomal protein L31-A / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 7.025393 KDa
SequenceString:
TINLHKRLHG VSFKKRAPRA VKEIKKFAKL HMGTDDVRLA PELNQAIWKR GVKGVEYRLR

UniProtKB: Large ribosomal subunit protein eL31A

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Macromolecule #9: 60S ribosomal protein L22-A

MacromoleculeName: 60S ribosomal protein L22-A / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 2.350864 KDa
SequenceString:
KYLKYLTKKY LKKNQLRD

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Macromolecule #10: 60S ribosome ribosomal protein L19A

MacromoleculeName: 60S ribosome ribosomal protein L19A / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 7.490019 KDa
SequenceString:
KAVTVHSKRL PSQVVWIRRL RVLRRLLAKY RIDKHLYHVL YKESKGNAFK HKRALVEHII QA

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Macromolecule #11: 60S ribosomal protein L30

MacromoleculeName: 60S ribosomal protein L30 / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 4.390093 KDa
SequenceString:
LGYIIIAANT PVLRKSVYYF QGGNNELGTA VGKLFRVGVV S

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Macromolecule #12: 60S ribosomal protein L3

MacromoleculeName: 60S ribosomal protein L3 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 42.452184 KDa
SequenceString: GHLGFLPRKR AASIRARVKA FPKDDRSKPV ALTSFLGYKA GMTTIVRDLD RPGSKFHKRE VVEAVTVVDT PPVVVVGVVG YVETPRGLR SLTTVWAEHL SDEVKRRFYK NWYKSKKKAF TKYSAKYAQD GAGIERELAR IKKYASVVRV LVHTQIRKTP L AQKKAHLA ...String:
GHLGFLPRKR AASIRARVKA FPKDDRSKPV ALTSFLGYKA GMTTIVRDLD RPGSKFHKRE VVEAVTVVDT PPVVVVGVVG YVETPRGLR SLTTVWAEHL SDEVKRRFYK NWYKSKKKAF TKYSAKYAQD GAGIERELAR IKKYASVVRV LVHTQIRKTP L AQKKAHLA EIQLNGGSIS EKVDWAREHF EKTVAVDSVF EQNEMIDAIA VTKGHGFEGV THRWGTKKLP RKTHRGLRKV AC IGAWHPA HVMWSVARAG QRGYHSRTSI NHKIYRVGKG DDEANGATSF DRTKKTITPM GGFVHYGEIK NDFIMVKGCI PGN RKRIVT LRKSLYTNTS RKALEEVSLK WIDTASKFGK GRFQTPAEKH AFMGTLKKDL

UniProtKB: Large ribosomal subunit protein uL3

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Macromolecule #13: UPF0642 protein YBL028C

MacromoleculeName: UPF0642 protein YBL028C / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 646.78 Da
SequenceString:
AKSLRA

UniProtKB: UPF0642 protein YBL028C

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Macromolecule #14: Nuclear GTP-binding protein NUG1

MacromoleculeName: Nuclear GTP-binding protein NUG1 / type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 977.232 Da
SequenceString:
MRVRKRQ

UniProtKB: Nuclear GTP-binding protein NUG1

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Macromolecule #15: Eukaryotic translation initiation factor 6

MacromoleculeName: Eukaryotic translation initiation factor 6 / type: protein_or_peptide / ID: 15 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 26.476605 KDa
SequenceString: MATRTQFENS NEIGVFSKLT NTYCLVAVGG SENFYSAFEA ELGDAIPIVH TTIAGTRIIG RMTAGNRRGL LVPTQTTDQE LQHLRNSLP DSVKIQRVEE RLSALGNVIC CNDYVALVHP DIDRETEELI SDVLGVEVFR QTISGNILVG SYCSLSNQGG L VHPQTSVQ ...String:
MATRTQFENS NEIGVFSKLT NTYCLVAVGG SENFYSAFEA ELGDAIPIVH TTIAGTRIIG RMTAGNRRGL LVPTQTTDQE LQHLRNSLP DSVKIQRVEE RLSALGNVIC CNDYVALVHP DIDRETEELI SDVLGVEVFR QTISGNILVG SYCSLSNQGG L VHPQTSVQ DQEELSSLLQ VPLVAGTVNR GSSVVGAGMV VNDYLAVTGL DTTAPELSVI ESIFRLQDAQ PESISGNLRD TL IETYS

UniProtKB: Eukaryotic translation initiation factor 6

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Macromolecule #16: 25S rRNA (cytosine(2870)-C(5))-methyltransferase

MacromoleculeName: 25S rRNA (cytosine(2870)-C(5))-methyltransferase / type: protein_or_peptide / ID: 16 / Number of copies: 1 / Enantiomer: LEVO / EC number: 25S rRNA (cytosine2870-C5)-methyltransferase
Source (natural)Organism: Saccharomyces cerevisiae BY4741 (yeast) / Strain: BY4741
Molecular weightTheoretical: 3.821383 KDa
SequenceString:
LAQTLVNRGV NLQPIGSWTK VGLQIFDSQV PIGATP

UniProtKB: 25S rRNA (cytosine(2870)-C(5))-methyltransferase

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Macromolecule #17: S-ADENOSYL-L-HOMOCYSTEINE

MacromoleculeName: S-ADENOSYL-L-HOMOCYSTEINE / type: ligand / ID: 17 / Number of copies: 1 / Formula: SAH
Molecular weightTheoretical: 384.411 Da
Chemical component information

ChemComp-SAH:
S-ADENOSYL-L-HOMOCYSTEINE

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Macromolecule #18: water

MacromoleculeName: water / type: ligand / ID: 18 / Number of copies: 2 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration0.45 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
50.0 mMCH2[CH2NHC(CH2OH)3]2Bis-Tris-Propane
150.0 mMNaClSodium Chloride
10.0 mMMgCl2Magnesium Chloride
1.0 mMC9H15O6PHClTCEP
0.01 % (w/v)NP-40
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.3
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) / Number grids imaged: 1 / Number real images: 4523 / Average exposure time: 0.05 sec. / Average electron dose: 1.2 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 81000
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

Particle selectionNumber selected: 825096
Startup modelType of model: EMDB MAP
EMDB ID:

3891

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.13 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 198000
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final 3D classificationNumber classes: 4 / Avg.num./class: 118000 / Software - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT
Output model

PDB-7naf:
State E2 nucleolar 60S ribosomal biogenesis intermediate - Spb1-MTD local model

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