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- PDB-8cas: Cryo-EM structure of native Otu2-bound ubiquitinated 48S initiati... -

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Basic information

Entry
Database: PDB / ID: 8cas
TitleCryo-EM structure of native Otu2-bound ubiquitinated 48S initiation complex (partial)
Components
  • (40S ribosomal protein ...) x 30
  • (60S ribosomal protein ...) x 2
  • (Eukaryotic translation initiation factor 3 subunit ...) x 5
  • 18S ribosomal RNA
  • Eukaryotic translation initiation factor 4C
  • Guanine nucleotide-binding protein subunit beta-like protein
  • OTU domain-containing protein 2
  • RLI1 isoform 1
  • RPS15 isoform 1
  • RPS29A isoform 1
  • SUI2 isoform 1
  • SUI3 isoform 1
  • TIF5 isoform 1
  • mRNA
  • protein-synthesizing GTPase
  • tRNA
KeywordsRIBOSOME / Translation / Ubiquitin / Deubiquitinating enzyme / Initiation / Complex
Function / homology
Function and homology information


eukaryotic translation initiation factor 3 complex, eIF3e / eukaryotic translation initiation factor 3 complex, eIF3m / translation reinitiation / incipient cellular bud site / multi-eIF complex / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex / protein-synthesizing GTPase / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, LSU-rRNA,5S) / formation of cytoplasmic translation initiation complex ...eukaryotic translation initiation factor 3 complex, eIF3e / eukaryotic translation initiation factor 3 complex, eIF3m / translation reinitiation / incipient cellular bud site / multi-eIF complex / eukaryotic translation initiation factor 3 complex / eukaryotic 43S preinitiation complex / protein-synthesizing GTPase / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, LSU-rRNA,5S) / formation of cytoplasmic translation initiation complex / eukaryotic 48S preinitiation complex / negative regulation of glucose mediated signaling pathway / negative regulation of translational frameshifting / Negative regulators of DDX58/IFIH1 signaling / positive regulation of translational fidelity / Protein methylation / RMTs methylate histone arginines / mTORC1-mediated signalling / Protein hydroxylation / ribosome-associated ubiquitin-dependent protein catabolic process / GDP-dissociation inhibitor activity / positive regulation of nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / Formation of the ternary complex, and subsequently, the 43S complex / Translation initiation complex formation / Ribosomal scanning and start codon recognition / preribosome, small subunit precursor / mRNA destabilization / 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 / Formation of a pool of free 40S subunits / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / L13a-mediated translational silencing of Ceruloplasmin expression / ribosomal large subunit export from nucleus / endonucleolytic cleavage to generate mature 3'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / G-protein alpha-subunit binding / positive regulation of protein kinase activity / regulation of translational fidelity / Ub-specific processing proteases / ribosomal small subunit export from nucleus / translation regulator activity / ribosomal subunit export from nucleus / endonucleolytic cleavage in ITS1 to separate SSU-rRNA from 5.8S rRNA and LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / DNA-(apurinic or apyrimidinic site) endonuclease activity / translation initiation factor binding / translational initiation / translation initiation factor activity / cellular response to amino acid starvation / ribosome assembly / rescue of stalled ribosome / 90S preribosome / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of SSU-rRNA / small-subunit processome / positive regulation of apoptotic signaling pathway / protein kinase C binding / maintenance of translational fidelity / ribosomal large subunit assembly / cytoplasmic stress granule / modification-dependent protein catabolic process / rRNA processing / protein tag activity / ribosome biogenesis / ribosome binding / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cysteine-type deubiquitinase activity / cytosolic large ribosomal subunit / tRNA binding / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / protein ubiquitination / structural constituent of ribosome / translation / positive regulation of protein phosphorylation / G protein-coupled receptor signaling pathway / negative regulation of gene expression / GTPase activity / mRNA binding / ubiquitin protein ligase binding / nucleolus / GTP binding / ATP hydrolysis activity / mitochondrion / RNA binding / zinc ion binding / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm / cytosol
Similarity search - Function
: / Eukaryotic translation initiation factor 3 subunit G / Eukaryotic translation initiation factor 3 subunit G, N-terminal / eIF3G, RNA recognition motif / Eukaryotic translation initiation factor 3 subunit G / Eukaryotic translation initiation factor 3 subunit I / RLI, domain 1 / RLI1 / RNase L inhibitor RLI-like, possible metal-binding domain / Eukaryotic translation initiation factor 3 subunit B ...: / Eukaryotic translation initiation factor 3 subunit G / Eukaryotic translation initiation factor 3 subunit G, N-terminal / eIF3G, RNA recognition motif / Eukaryotic translation initiation factor 3 subunit G / Eukaryotic translation initiation factor 3 subunit I / RLI, domain 1 / RLI1 / RNase L inhibitor RLI-like, possible metal-binding domain / Eukaryotic translation initiation factor 3 subunit B / eIF3B, RNA recognition motif / Possible Fer4-like domain in RNase L inhibitor, RLI / Eukaryotic translation initiation factor 3 subunit C, N-terminal domain / Eukaryotic translation initiation factor 3 subunit C / Eukaryotic translation initiation factor 3 subunit 8 N-terminus / Translation initiation factor, beta propellor-like domain / Eukaryotic translation initiation factor 3 subunit A / OTU-like cysteine protease / Eukaryotic translation initiation factor eIF2A / Translation initiation factor IF2/IF5 / Translation initiation factor IF2/IF5 domain / Translation initiation factor IF2/IF5, N-terminal / Translation initiation factor IF2/IF5, zinc-binding / Domain found in IF2B/IF5 / domain present in translation initiation factor eIF2B and eIF5 / Translation initiation factor 1A (eIF-1A), conserved site / Eukaryotic initiation factor 1A signature. / OTU domain / OTU domain profile. / eukaryotic translation initiation factor 1A / Translation initiation factor 1A (eIF-1A) / eIF4-gamma/eIF5/eIF2-epsilon / Domain at the C-termini of GCD6, eIF-2B epsilon, eIF-4 gamma and eIF-5 / W2 domain / W2 domain profile. / IF2a, S1-like domain / Translation initiation factor 2, alpha subunit / Translation initiation factor 2, alpha subunit, middle domain superfamily / Translation initiation factor 2, alpha subunit, C-terminal / Eukaryotic translation initiation factor 2 alpha subunit / Initiation factor eIF2 gamma, domain 2 / Initiation factor eIF2 gamma, GTP-binding domain / Initiation factor eIF2 gamma, C-terminal / Initiation factor eIF2 gamma, C terminal / RNA-binding domain, S1, IF1 type / Translation initiation factor 1A / IF-1 / S1 domain IF1 type profile. / 4Fe-4S binding domain / RNA-binding domain, S1 / PCI/PINT associated module / motif in proteasome subunits, Int-6, Nip-1 and TRIP-15 / PCI domain / Proteasome component (PCI) domain / PCI domain profile. / Translation elongation factor EF1A/initiation factor IF2gamma, C-terminal / : / Ribosomal protein S26e signature. / S1 domain profile. / : / Ribosomal protein L41 / Ribosomal protein L41 / Ribosomal protein S26e / Ribosomal protein S26e superfamily / Ribosomal protein S26e / Ribosomal protein S21e, conserved site / Ribosomal protein S21e signature. / Ribosomal protein S12e signature. / Ribosomal protein S12e / Ribosomal protein S5, eukaryotic/archaeal / Ribosomal protein S19e, conserved site / Ribosomal protein S19e signature. / Small (40S) ribosomal subunit Asc1/RACK1 / Ribosomal protein S2, eukaryotic / Ribosomal protein S21e / Ribosomal protein S21e superfamily / Ribosomal protein S21e / 40S Ribosomal protein S10 / S27a-like superfamily / Ribosomal protein S10, eukaryotic/archaeal / Plectin/S10, N-terminal / Plectin/S10 domain / Ribosomal protein S25 / S25 ribosomal protein / Ribosomal protein S2, eukaryotic/archaeal / : / Ribosomal protein S17e, conserved site / Ribosomal protein S17e signature. / Ribosomal protein S27a / Ribosomal protein S27a / Ribosomal protein S27a / Ribosomal protein S8e subdomain, eukaryotes / Ribosomal protein S30 / Ribosomal protein S30 / 40S ribosomal protein S29/30S ribosomal protein S14 type Z / Ribosomal protein S7e signature. / Ribosomal protein S3, eukaryotic/archaeal / Ribosomal protein S19e / Ribosomal protein S3Ae, conserved site / Ribosomal protein S19e / Ribosomal protein S3Ae signature.
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / ADENOSINE-5'-TRIPHOSPHATE / PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER / METHIONINE / IRON/SULFUR CLUSTER / RNA / RNA (> 10) / RNA (> 100) / RNA (> 1000) / Eukaryotic translation initiation factor 3 subunit I ...ADENOSINE-5'-DIPHOSPHATE / ADENOSINE-5'-TRIPHOSPHATE / PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER / METHIONINE / IRON/SULFUR CLUSTER / RNA / RNA (> 10) / RNA (> 100) / RNA (> 1000) / Eukaryotic translation initiation factor 3 subunit I / Eukaryotic translation initiation factor 4C / SUI2 isoform 1 / protein-synthesizing GTPase / RLI1 isoform 1 / TIF5 isoform 1 / SUI3 isoform 1 / Eukaryotic translation initiation factor 3 subunit C / Eukaryotic translation initiation factor 3 subunit B / Eukaryotic translation initiation factor 3 subunit G / Small ribosomal subunit protein uS4A / Small ribosomal subunit protein eS17A / Small ribosomal subunit protein uS3 / Small ribosomal subunit protein uS15 / Ubiquitin-ribosomal protein eS31 fusion protein / Small ribosomal subunit protein uS11A / Small ribosomal subunit protein eS19A / Small ribosomal subunit protein eS21A / Small ribosomal subunit protein uS8A / Ubiquitin-ribosomal protein eL40A fusion protein / Small ribosomal subunit protein uS12A / Small ribosomal subunit protein eS24A / Small ribosomal subunit protein eS30A / Small ribosomal subunit protein eS4B / Small ribosomal subunit protein eS6B / Small ribosomal subunit protein eS8A / Small ribosomal subunit protein uS17A / Small ribosomal subunit protein uS9A / Small ribosomal subunit protein uS13A / Small ribosomal subunit protein eS32A / Small ribosomal subunit protein uS5 / Small ribosomal subunit protein uS7 / Small ribosomal subunit protein eS7A / Small ribosomal subunit protein uS2A / Small ribosomal subunit protein eS1A / Small ribosomal subunit protein eS27A / Small ribosomal subunit protein RACK1 / Eukaryotic translation initiation factor 3 subunit A / Small ribosomal subunit protein uS10 / OTU domain-containing protein 2 / Small ribosomal subunit protein eS26A / Small ribosomal subunit protein uS14A / Small ribosomal subunit protein eS12 / Small ribosomal subunit protein uS19 / Small ribosomal subunit protein eS10A / Small ribosomal subunit protein eS25A / Small ribosomal subunit protein eS28A
Similarity search - Component
Biological speciesSaccharomyces cerevisiae W303 (yeast)
Saccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsIkeuchi, K. / Buschauer, R. / Cheng, J. / Berninghausen, O. / Becker, T. / Beckmann, R.
Funding support Germany, Japan, 5items
OrganizationGrant numberCountry
German Research Foundation (DFG)SFB/TRR-174 Germany
German Research Foundation (DFG)BE1814/15-1 Germany
German Research Foundation (DFG)BE1814/1-1 Germany
Japan Society for the Promotion of Science (JSPS)JSPS Overseas Research Fellowships Japan
Boehringer Ingelheim Fonds (BIF)PhD fellowship Germany
CitationJournal: Nat Commun / Year: 2023
Title: Molecular basis for recognition and deubiquitination of 40S ribosomes by Otu2.
Authors: Ken Ikeuchi / Nives Ivic / Robert Buschauer / Jingdong Cheng / Thomas Fröhlich / Yoshitaka Matsuo / Otto Berninghausen / Toshifumi Inada / Thomas Becker / Roland Beckmann /
Abstract: In actively translating 80S ribosomes the ribosomal protein eS7 of the 40S subunit is monoubiquitinated by the E3 ligase Not4 and deubiquitinated by Otu2 upon ribosomal subunit recycling. Despite its ...In actively translating 80S ribosomes the ribosomal protein eS7 of the 40S subunit is monoubiquitinated by the E3 ligase Not4 and deubiquitinated by Otu2 upon ribosomal subunit recycling. Despite its importance for translation efficiency the exact role and structural basis for this translational reset is poorly understood. Here, structural analysis by cryo-electron microscopy of native and reconstituted Otu2-bound ribosomal complexes reveals that Otu2 engages 40S subunits mainly between ribosome recycling and initiation stages. Otu2 binds to several sites on the intersubunit surface of the 40S that are not occupied by any other 40S-binding factors. This binding mode explains the discrimination against 80S ribosomes via the largely helical N-terminal domain of Otu2 as well as the specificity for mono-ubiquitinated eS7 on 40S. Collectively, this study reveals mechanistic insights into the Otu2-driven deubiquitination steps for translational reset during ribosome recycling/(re)initiation.
History
DepositionJan 24, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 24, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
P: 40S ribosomal protein S0-A
Q: 40S ribosomal protein S1-A
R: 40S ribosomal protein S2
S: 40S ribosomal protein S4-B
T: 40S ribosomal protein S6-B
V: 40S ribosomal protein S8-A
W: 40S ribosomal protein S9-A
X: 40S ribosomal protein S11-A
Y: 40S ribosomal protein S13
Z: 40S ribosomal protein S14-A
a: 40S ribosomal protein S21-A
b: 40S ribosomal protein S22-A
c: 40S ribosomal protein S23-A
d: 40S ribosomal protein S24-A
e: 40S ribosomal protein S26-A
f: 40S ribosomal protein S27-A
g: 40S ribosomal protein S30-A
E: RPS15 isoform 1
B: 40S ribosomal protein S3
D: 40S ribosomal protein S10-A
F: 40S ribosomal protein S12
H: 40S ribosomal protein S16-A
I: 40S ribosomal protein S17-A
J: 40S ribosomal protein S18-A
K: 40S ribosomal protein S19-A
L: 40S ribosomal protein S20
M: 40S ribosomal protein S25-A
N: RPS29A isoform 1
O: 40S ribosomal protein S31
h: Guanine nucleotide-binding protein subunit beta-like protein
i: 40S ribosomal protein S28-A
l: Eukaryotic translation initiation factor 3 subunit I
r: Eukaryotic translation initiation factor 3 subunit G
1: tRNA
m: TIF5 isoform 1
o: Eukaryotic translation initiation factor 3 subunit A
p: Eukaryotic translation initiation factor 3 subunit B
q: Eukaryotic translation initiation factor 3 subunit C
3: mRNA
A: Eukaryotic translation initiation factor 4C
k: RLI1 isoform 1
2: 18S ribosomal RNA
C: 40S ribosomal protein S5
y: 60S ribosomal protein L40-A
n: 60S ribosomal protein L41-A
j: SUI2 isoform 1
G: protein-synthesizing GTPase
s: SUI3 isoform 1
x: OTU domain-containing protein 2
U: 40S ribosomal protein S7-A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,861,56465
Polymers1,858,87350
Non-polymers2,69115
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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40S ribosomal protein ... , 30 types, 30 molecules PQRSTVWXYZabcdefgBDFHIJKLMOiCU

#1: Protein 40S ribosomal protein S0-A / Nucleic acid-binding protein NAB1A / Small ribosomal subunit protein uS2-A


Mass: 28051.330 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P32905
#2: Protein 40S ribosomal protein S1-A / RP10A / Small ribosomal subunit protein eS1-A


Mass: 28798.467 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P33442
#3: Protein 40S ribosomal protein S2 / Omnipotent suppressor protein SUP44 / RP12 / S4 / Small ribosomal subunit protein uS5 / YS5


Mass: 27490.826 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P25443
#4: Protein 40S ribosomal protein S4-B / RP5 / S7 / Small ribosomal subunit protein eS4-B / YS6


Mass: 29469.330 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX36
#5: Protein 40S ribosomal protein S6-B / RP9 / S10 / Small ribosomal subunit protein eS6-B / YS4


Mass: 27054.486 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX38
#6: Protein 40S ribosomal protein S8-A


Mass: 22537.803 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX39
#7: Protein 40S ribosomal protein S9-A


Mass: 22487.893 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: O13516
#8: Protein 40S ribosomal protein S11-A


Mass: 17785.934 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX47
#9: Protein 40S ribosomal protein S13 / S27a / Small ribosomal subunit protein uS15 / YS15


Mass: 17059.945 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P05756
#10: Protein 40S ribosomal protein S14-A


Mass: 14562.655 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P06367
#11: Protein 40S ribosomal protein S21-A / S26 / Small ribosomal subunit protein eS21-A / YS25


Mass: 9758.829 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0C0V8
#12: Protein 40S ribosomal protein S22-A


Mass: 14650.062 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0C0W1
#13: Protein 40S ribosomal protein S23-A / RP37 / S28 / Small ribosomal subunit protein uS12-A / YS14


Mass: 16073.896 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX29
#14: Protein 40S ribosomal protein S24-A / RP50 / Small ribosomal subunit protein eS24-A


Mass: 15362.848 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX31
#15: Protein 40S ribosomal protein S26-A / Small ribosomal subunit protein eS26-A


Mass: 13538.922 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P39938
#16: Protein 40S ribosomal protein S27-A / RP61 / Small ribosomal subunit protein eS27-A / YS20


Mass: 8893.391 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P35997
#17: Protein 40S ribosomal protein S30-A / Small ribosomal subunit protein eS30-A


Mass: 7137.541 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX33
#19: Protein 40S ribosomal protein S3 / RP13 / Small ribosomal subunit protein uS3 / YS3


Mass: 26542.789 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P05750
#20: Protein 40S ribosomal protein S10-A / Small ribosomal subunit protein eS10-A


Mass: 12757.445 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: Q08745
#21: Protein 40S ribosomal protein S12 / Small ribosomal subunit protein eS12


Mass: 15488.631 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P48589
#22: Protein 40S ribosomal protein S16-A / RP61R / Small ribosomal subunit protein uS9-A


Mass: 15877.490 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX51
#23: Protein 40S ribosomal protein S17-A / RP51A / Small ribosomal subunit protein eS17-A


Mass: 15820.413 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P02407
#24: Protein 40S ribosomal protein S18-A / Small ribosomal subunit protein uS13-A


Mass: 17071.641 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX55
#25: Protein 40S ribosomal protein S19-A / RP55A / S16a / Small ribosomal subunit protein eS19-A / YP45 / YS16A


Mass: 15942.125 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P07280
#26: Protein 40S ribosomal protein S20 / Small ribosomal subunit protein uS10


Mass: 13929.044 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P38701
#27: Protein 40S ribosomal protein S25-A / RP45 / S31 / Small ribosomal subunit protein eS25-A / YS23


Mass: 12067.272 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: Q3E792
#29: Protein 40S ribosomal protein S31 / CEP76 / S37 / Small ribosomal subunit protein eS31 / YS24


Mass: 8703.462 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P05759
#31: Protein 40S ribosomal protein S28-A / S33 / Small ribosomal subunit protein eS28-A / YS27


Mass: 7605.847 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: Q3E7X9
#43: Protein 40S ribosomal protein S5 / RP14 / S2 / Small ribosomal subunit protein uS7 / YS8


Mass: 25072.600 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P26783
#50: Protein 40S ribosomal protein S7-A / RP30 / RP40 / Small ribosomal subunit protein eS7-A


Mass: 21658.209 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P26786

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Protein , 10 types, 10 molecules ENhmAkjGsx

#18: Protein RPS15 isoform 1


Mass: 16031.907 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: Q01855
#28: Protein RPS29A isoform 1


Mass: 6675.723 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P41057
#30: Protein Guanine nucleotide-binding protein subunit beta-like protein / Receptor for activated C kinase / Receptor of activated protein kinase C 1 / RACK1 / Small ...Receptor for activated C kinase / Receptor of activated protein kinase C 1 / RACK1 / Small ribosomal subunit protein RACK1


Mass: 34841.219 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P38011
#35: Protein TIF5 isoform 1


Mass: 45321.977 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A6A5Q3C7
#40: Protein Eukaryotic translation initiation factor 4C


Mass: 17462.168 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A6A5PU90
#41: Protein RLI1 isoform 1


Mass: 68433.242 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A6A5Q259
#46: Protein SUI2 isoform 1


Mass: 34763.652 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A6A5PWU9
#47: Protein protein-synthesizing GTPase


Mass: 57942.699 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast)
References: UniProt: A0A6A5Q017, protein-synthesizing GTPase
#48: Protein SUI3 isoform 1


Mass: 31631.309 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A8H4BUS5
#49: Protein OTU domain-containing protein 2


Mass: 36157.016 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae W303 (yeast) / Gene: OTU2, YHL013C / Variant: C178S / Plasmid: pKI1034 / Details (production host): p415GPDp-otu2-C178S-3xFLAG-CYC1t / Production host: Saccharomyces cerevisiae W303 (yeast) / Variant (production host): otu2 delta / References: UniProt: P38747

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Eukaryotic translation initiation factor 3 subunit ... , 5 types, 5 molecules lropq

#32: Protein Eukaryotic translation initiation factor 3 subunit I


Mass: 38803.375 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A6A5PTA5
#33: Protein Eukaryotic translation initiation factor 3 subunit G


Mass: 30547.525 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A8H8UNW9
#36: Protein Eukaryotic translation initiation factor 3 subunit A / eIF3a / Eukaryotic translation initiation factor 3 110 kDa subunit homolog / eIF3 p110 / ...eIF3a / Eukaryotic translation initiation factor 3 110 kDa subunit homolog / eIF3 p110 / Translation initiation factor eIF3 / p110 subunit homolog


Mass: 110517.641 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P38249
#37: Protein Eukaryotic translation initiation factor 3 subunit B


Mass: 88241.766 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A8H4F8W9
#38: Protein Eukaryotic translation initiation factor 3 subunit C


Mass: 93310.172 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: A0A8H4BZ29

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RNA chain , 3 types, 3 molecules 132

#34: RNA chain tRNA


Mass: 24712.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast)
#39: RNA chain mRNA


Mass: 2534.585 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast)
#42: RNA chain 18S ribosomal RNA


Mass: 579761.938 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast)

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60S ribosomal protein ... , 2 types, 2 molecules yn

#44: Protein 60S ribosomal protein L40-A / CEP52 / Large ribosomal subunit protein eL40-A


Mass: 8576.831 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CH08
#45: Protein/peptide 60S ribosomal protein L41-A / L47 / Large ribosomal subunit protein eL41-A / YL41


Mass: 3354.243 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae W303 (yeast) / References: UniProt: P0CX86

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Non-polymers , 7 types, 15 molecules

#51: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Zn
#52: Chemical ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM
#53: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Mg
#54: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
#55: Chemical ChemComp-SF4 / IRON/SULFUR CLUSTER


Mass: 351.640 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Fe4S4
#56: Chemical ChemComp-MET / METHIONINE


Type: L-peptide linking / Mass: 149.211 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H11NO2S
#57: Chemical ChemComp-GCP / PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER


Mass: 521.208 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C11H18N5O13P3 / Comment: GMP-PCP, energy-carrying molecule analogue*YM

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Yeast native 48S initiation complex (partial) with deubiquitinating enzyme Otu2 bound to ubiquitinated eS7
Type: RIBOSOME / Entity ID: #1-#50 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: W303
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 46.4 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 12059 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00691456
ELECTRON MICROSCOPYf_angle_d0.895134209
ELECTRON MICROSCOPYf_dihedral_angle_d16.34731055
ELECTRON MICROSCOPYf_chiral_restr0.05117990
ELECTRON MICROSCOPYf_plane_restr0.00411116

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