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- EMDB-20249: Structure of a mammalian small ribosomal subunit in complex with ... -

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Basic information

Entry
Database: EMDB / ID: EMD-20249
TitleStructure of a mammalian small ribosomal subunit in complex with the Israeli Acute Paralysis Virus IRES (Class 2)
Map data
Sample
  • Complex: Structure of a mammalian small ribosomal subunit in complex with the Israeli Acute Paralysis Virus IRES (Class 2)
    • RNA: x 2 types
    • Protein or peptide: x 34 types
Function / homology
Function and homology information


ribosomal subunit / DNA-(apurinic or apyrimidinic site) lyase / structural constituent of ribosome / translation
Similarity search - Function
Ribosomal protein S26e signature. / Ribosomal protein S21e signature. / Ribosomal protein S12e signature. / Ribosomal protein S19e signature. / 40S Ribosomal protein S10 / Plectin/S10, N-terminal / Plectin/S10 domain / Ribosomal protein S17e signature. / Ribosomal protein S7e signature. / Ribosomal protein S3Ae signature. ...Ribosomal protein S26e signature. / Ribosomal protein S21e signature. / Ribosomal protein S12e signature. / Ribosomal protein S19e signature. / 40S Ribosomal protein S10 / Plectin/S10, N-terminal / Plectin/S10 domain / Ribosomal protein S17e signature. / Ribosomal protein S7e signature. / Ribosomal protein S3Ae signature. / Ribosomal protein S27e signature. / Ribosomal protein S4e signature. / Ribosomal protein S8e signature. / Ribosomal S24e conserved site / Ribosomal protein S24e signature. / Ribosomal protein S24e / Ribosomal protein S24e / Ribosomal protein S6e signature. / Ribosomal protein S28e signature. / Ribosomal protein S2 signature 2. / Ubiquitin domain signature. / Ribosomal protein S3 signature. / Ribosomal protein S10 signature. / Ribosomal protein S14 signature. / Ribosomal protein S2 signature 1. / Type-2 KH domain profile. / Ribosomal protein S19 signature. / Ribosomal protein S5 signature. / Ribosomal protein S7 signature. / S5 double stranded RNA-binding domain profile. / Ribosomal protein S17 signature. / Ribosomal protein S13 signature. / Ribosomal protein S13 family profile. / Ribosomal protein S8 signature. / Ribosomal protein S4 signature. / Ribosomal protein S15 signature. / S4 RNA-binding domain profile. / Ribosomal protein S11 signature. / Ubiquitin domain profile. / Ribosomal protein S9 signature. / Ribosomal protein S12 signature. / Ribosomal protein L23/L15e core domain superfamily / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
Small ribosomal subunit protein eS32 / Small ribosomal subunit protein uS4 / Small ribosomal subunit protein eS12 / Small ribosomal subunit protein uS9 / Small ribosomal subunit protein uS10 / Small ribosomal subunit protein RACK1 / Ubiquitin-ribosomal protein eS31 fusion protein / Small ribosomal subunit protein uS15 / Small ribosomal subunit protein eS1 / Small ribosomal subunit protein eS7 ...Small ribosomal subunit protein eS32 / Small ribosomal subunit protein uS4 / Small ribosomal subunit protein eS12 / Small ribosomal subunit protein uS9 / Small ribosomal subunit protein uS10 / Small ribosomal subunit protein RACK1 / Ubiquitin-ribosomal protein eS31 fusion protein / Small ribosomal subunit protein uS15 / Small ribosomal subunit protein eS1 / Small ribosomal subunit protein eS7 / Small ribosomal subunit protein uS5 / Small ribosomal subunit protein uS12 / Small ribosomal subunit protein uS11 / 40S ribosomal protein S24 / Ubiquitin-like FUBI-ribosomal protein eS30 fusion protein / Small ribosomal subunit protein eS25 / Small ribosomal subunit protein eS26 / Small ribosomal subunit protein uS7 / Small ribosomal subunit protein uS8 / Small ribosomal subunit protein eS28 / Small ribosomal subunit protein eS8 / Small ribosomal subunit protein eS4 / Small ribosomal subunit protein eS6 / Small ribosomal subunit protein eS21 / Small ribosomal subunit protein eS19 / Small ribosomal subunit protein uS3 / Small ribosomal subunit protein uS13 / Plectin/eS10 N-terminal domain-containing protein / Small ribosomal subunit protein uS17 / Small ribosomal subunit protein eS17 / Small ribosomal subunit protein uS2 / Small ribosomal subunit protein eS27 / Small ribosomal subunit protein uS19 / Small ribosomal subunit protein uS11 / Small ribosomal subunit protein uS14
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit) / Rabbit (rabbit) / Israeli acute paralysis virus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsAcosta-Reyes FJ / Neupane R / Frank J / Fernandez IS
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM29169 United States
CitationJournal: EMBO J / Year: 2019
Title: The Israeli acute paralysis virus IRES captures host ribosomes by mimicking a ribosomal state with hybrid tRNAs.
Authors: Francisco Acosta-Reyes / Ritam Neupane / Joachim Frank / Israel S Fernández /
Abstract: Colony collapse disorder (CCD) is a multi-faceted syndrome decimating bee populations worldwide, and a group of viruses of the widely distributed Dicistroviridae family have been identified as a ...Colony collapse disorder (CCD) is a multi-faceted syndrome decimating bee populations worldwide, and a group of viruses of the widely distributed Dicistroviridae family have been identified as a causing agent of CCD. This family of viruses employs non-coding RNA sequences, called internal ribosomal entry sites (IRESs), to precisely exploit the host machinery for viral protein production. Using single-particle cryo-electron microscopy (cryo-EM), we have characterized how the IRES of Israeli acute paralysis virus (IAPV) intergenic region captures and redirects translating ribosomes toward viral RNA messages. We reconstituted two in vitro reactions targeting a pre-translocation and a post-translocation state of the IAPV-IRES in the ribosome, allowing us to identify six structures using image processing classification methods. From these, we reconstructed the trajectory of IAPV-IRES from the early small subunit recruitment to the final post-translocated state in the ribosome. An early commitment of IRES/ribosome complexes for global pre-translocation mimicry explains the high efficiency observed for this IRES. Efforts directed toward fighting CCD by targeting the IAPV-IRES using RNA-interference technology are underway, and the structural framework presented here may assist in further refining these approaches.
History
DepositionMay 27, 2019-
Header (metadata) releaseSep 18, 2019-
Map releaseSep 18, 2019-
UpdateDec 2, 2020-
Current statusDec 2, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6p4h
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20249.png

Downloads & links

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Map

FileDownload / File: emd_20249.map.gz / Format: CCP4 / Size: 115.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.233 Å
Density
Contour LevelBy AUTHOR: 0.024 / Movie #1: 0.024
Minimum - Maximum-0.112940155 - 0.2127223
Average (Standard dev.)0.00042087282 (±0.0059494916)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions312312312
Spacing312312312
CellA=B=C: 384.696 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.2331.2331.233
M x/y/z312312312
origin x/y/z0.0000.0000.000
length x/y/z384.696384.696384.696
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS312312312
D min/max/mean-0.1130.2130.000

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Supplemental data

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Mask #1

Fileemd_20249_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: #1

Fileemd_20249_additional.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_20249_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_20249_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : Structure of a mammalian small ribosomal subunit in complex with ...

EntireName: Structure of a mammalian small ribosomal subunit in complex with the Israeli Acute Paralysis Virus IRES (Class 2)
Components
  • Complex: Structure of a mammalian small ribosomal subunit in complex with the Israeli Acute Paralysis Virus IRES (Class 2)
    • RNA: 18S rRNA18S ribosomal RNA
    • Protein or peptide: uS2
    • Protein or peptide: eS1
    • Protein or peptide: uS5
    • Protein or peptide: uS3
    • RNA: IAPV-IRES
    • Protein or peptide: eS4
    • Protein or peptide: uS7
    • Protein or peptide: eS6
    • Protein or peptide: eS7
    • Protein or peptide: eS8
    • Protein or peptide: uS4
    • Protein or peptide: eS10
    • Protein or peptide: uS17
    • Protein or peptide: eS12
    • Protein or peptide: uS15
    • Protein or peptide: uS11
    • Protein or peptide: uS19
    • Protein or peptide: uS9
    • Protein or peptide: eS17
    • Protein or peptide: uS13
    • Protein or peptide: eS19
    • Protein or peptide: uS10
    • Protein or peptide: eS21
    • Protein or peptide: uS8
    • Protein or peptide: uS12
    • Protein or peptide: eS24
    • Protein or peptide: eS25
    • Protein or peptide: eS26
    • Protein or peptide: eS27
    • Protein or peptide: eS28
    • Protein or peptide: eS29
    • Protein or peptide: eS30
    • Protein or peptide: eS31
    • Protein or peptide: RACK1Receptor for activated C kinase 1
    • Protein or peptide: eL41

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Supramolecule #1: Structure of a mammalian small ribosomal subunit in complex with ...

SupramoleculeName: Structure of a mammalian small ribosomal subunit in complex with the Israeli Acute Paralysis Virus IRES (Class 2)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#36
Source (natural)Organism: Oryctolagus cuniculus (rabbit)

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Macromolecule #1: 18S rRNA

MacromoleculeName: 18S rRNA / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 602.776875 KDa
SequenceString: UACCUGGUUG AUCCUGCCAG UAGCAUAUGC UUGUCUCAAA GAUUAAGCCA UGCAUGUCUA AGUACGCACG GCCGGUACAG UGAAACUGC GAAUGGCUCA UUAAAUCAGU UAUGGUUCCU UUGGUCGCUC GCUCCUCUCC CACUUGGAUA ACUGUGGUAA U UCUAGAGC ...String:
UACCUGGUUG AUCCUGCCAG UAGCAUAUGC UUGUCUCAAA GAUUAAGCCA UGCAUGUCUA AGUACGCACG GCCGGUACAG UGAAACUGC GAAUGGCUCA UUAAAUCAGU UAUGGUUCCU UUGGUCGCUC GCUCCUCUCC CACUUGGAUA ACUGUGGUAA U UCUAGAGC UAAUACAUGC CGACGGGCGC UGACCCCCUU CGCGGGGGGG AUGCGUGCAU UUAUCAGAUC AAAACCAACC CG GUCAGCC CCUCUCCGGC CCCGGCCGGG GGGCGGGCGC CGGCGGCUUU GGUGACUCUA GAUAACCUCG GGCCGAUCGC ACG CCCCCC GUGGCGGCGA CGACCCAUUC GAACGUCUGC CCUAUCAACU UUCGAUGGUA GUCGCCGUGC CUACCAUGGU GACC ACGGG UGACGGGGAA UCAGGGUUCG AUUCCGGAGA GGGAGCCUGA GAAACGGCUA CCACAUCCAA GGAAGGCAGC AGGCG CGCA AAUUACCCAC UCCCGACCCG GGGAGGUAGU GACGAAAAAU AACAAUACAG GACUCUUUCG AGGCCCUGUA AUUGGA AUG AGUCCACUUU AAAUCCUUUA ACGAGGAUCC AUUGGAGGGC AAGUCUGGUG CCAGCAGCCG CGGUAAUUCC AGCUCCA AU AGCGUAUAUU AAAGUUGCUG CAGUUAAAAA GCUCGUAGUU GGAUCUUGGG AGCGGGCGGG CGGUCCGCCG CGAGGCGA G CCACCGCCCG UCCCCGCCCC UUGCCUCUCG GCGCCCCCUC GAUGCUCUUA GCUGAGUGUC CCGCGGGGCC CGAAGCGUU UACUUUGAAA AAAUUAGAGU GUUCAAAGCA GGCCCGAGCC GCCUGGAUAC CGCAGCUAGG AAUAAUGGAA UAGGACCGCG GUUCUAUUU UGUUGGUUUU CGGAACUGAG GCCAUGAUUA AGAGGGACGG CCGGGGGCAU UCGUAUUGCG CCGCUAGAGG U GAAAUUCU UGGACCGGCG CAAGACGGAC CAGAGCGAAA GCAUUUGCCA AGAAUGUUUU CAUUAAUCAA GAACGAAAGU CG GAGGUUC GAAGACGAUC AGAUACCGUC GUAGUUCCGA CCAUAAACGA UGCCGACCGG CGAUGCGGCG GCGUUAUUCC CAU GACCCG CCGGGCAGCU UCCGGGAAAC CAAAGUCUUU GGGUUCCGGG GGGAGUAUGG UUGCAAAGCU GAAACUUAAA GGAA UUGAC GGAAGGGCAC CACCAGGAGU GGAGCCUGCG GCUUAAUUUG ACUCAACACG GGAAACCUCA CCCGGCCCGG ACACG GACA GGAUUGACAG AUUGAUAGCU CUUUCUCGAU UCCGUGGGUG GUGGUGCAUG GCCGUUCUUA GUUGGUGGAG CGAUUU GUC UGGUUAAUUC CGAUAACGAA CGAGACUCUG GCAUGCUAAC UAGUUACGCG ACCCCCGAGC GGUCGGCGUC CCCCAAC UU CUUAGAGGGA CAAGUGGCGU UCAGCCACCC GAGAUUGAGC AAUAACAGGU CUGUGAUGCC CUUAGAUGUC CGGGGCUG C ACGCGCGCUA CACUGACUGG CUCAGCGUGU GCCUACCCUA CGCCGGCAGG CGCGGGUAAC CCGUUGAACC CCAUUCGUG AUGGGGAUCG GGGAUUGCAA UUAUUCCCCA UGAACGAGGA AUUCCCAGUA AGUGCGGGUC AUAAGCUUGC GUUGAUUAAG UCCCUGCCC UUUGUACACA CCGCCCGUCG CUACUACCGA UUGGAUGGUU UAGUGAGGCC CUCGGAUCGG CCCCGCCGGG G UCGGCCCA CGGCCCUGGC GGAGCGCUGA GAAGACGGUC GAACUUGACU AUCUAGAGGA AGUAAAAGUC GUAACAAGGU UU CCGUAGG UGAACCUGCG GAAGGAUCAU UA

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Macromolecule #6: IAPV-IRES

MacromoleculeName: IAPV-IRES / type: rna / ID: 6 / Number of copies: 1
Source (natural)Organism: Israeli acute paralysis virus
Molecular weightTheoretical: 81.572078 KDa
SequenceString: GAGCGGUUUC UGGAAUACUA UAUGUAAGUA UAGUGUUCUG GAGGCAUCAU UCUAUGGUUA CCCAUCAUUA GAGGAAAUUU CCAAUAAAC UCUGGUGUAA GGCUUAGAGU GAUGGUCGAG GUGCCCUAUU UAGGGUGAGG AGCCUCGGUG GCAGCCCCAC C AAAUCCUC ...String:
GAGCGGUUUC UGGAAUACUA UAUGUAAGUA UAGUGUUCUG GAGGCAUCAU UCUAUGGUUA CCCAUCAUUA GAGGAAAUUU CCAAUAAAC UCUGGUGUAA GGCUUAGAGU GAUGGUCGAG GUGCCCUAUU UAGGGUGAGG AGCCUCGGUG GCAGCCCCAC C AAAUCCUC UAUUGGAUAG GAACAGCUGU ACUGGGCAGU UACAGCAGUC GUAUGGUAAC ACAUGCGGCG UUCCGAAAUA CC AUGCCUG GCGAUG

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Macromolecule #2: uS2

MacromoleculeName: uS2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 33.00307 KDa
SequenceString: MSGALDVLQM KEEDVLKFLA AGTHLGGTNL DFQMEQYIYK RKSDGIYIIN LKRTWEKLLL AARAIVAIEN PADVSVISSR NTGQRAVLK FAAATGATPI AGRFTPGTFT NQIQAAFREP RLLVVTDPQA DHQPLMEASY VNLPTIALCN TDSPLHYVDI A IPCNNKGA ...String:
MSGALDVLQM KEEDVLKFLA AGTHLGGTNL DFQMEQYIYK RKSDGIYIIN LKRTWEKLLL AARAIVAIEN PADVSVISSR NTGQRAVLK FAAATGATPI AGRFTPGTFT NQIQAAFREP RLLVVTDPQA DHQPLMEASY VNLPTIALCN TDSPLHYVDI A IPCNNKGA HSVGLMWWML AQEVLRMRGT ISREHPWEVM PDLYFYRDPE EIEKEEQAAA EKAVTKEEFQ GEWTAPEPEF TA TQPEVAD WSEGMQVPSV PIQQFPTEDW SAQPATEDWS AAPTAQATEW MGTTTEWS

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Macromolecule #3: eS1

MacromoleculeName: eS1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 30.002061 KDa
SequenceString: MAVGKNKRLT KGGKKGAKKK VVDPFSKKDW YDVKAPAMFN IRNIGKTLVT RTQGTKIASD GLKGRVFEVS LADLQNDEVA FRKFKLITE DVQGKNCLTN FHGMDLTRDK MCSMVKKWQT MIEAHVDVKT TDGYLLRLFC VGFTKKRNNQ IRKTSYAQHQ Q VRQIRKKM ...String:
MAVGKNKRLT KGGKKGAKKK VVDPFSKKDW YDVKAPAMFN IRNIGKTLVT RTQGTKIASD GLKGRVFEVS LADLQNDEVA FRKFKLITE DVQGKNCLTN FHGMDLTRDK MCSMVKKWQT MIEAHVDVKT TDGYLLRLFC VGFTKKRNNQ IRKTSYAQHQ Q VRQIRKKM MEIMTREVQT NDLKEVVNKL IPDSIGKDIE KACQSIYPLH DVFVRKVKML KKPKFELGKL MELHGEGSSS GK ATGDETG AKVERADGYE PPVQESV

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Macromolecule #4: uS5

MacromoleculeName: uS5 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 27.485209 KDa
SequenceString: MADDAGAAGG PGGPGGPGNK EWLPVTKLGR LVKDMKIKSL EEIYLFSLPI KESEIIDFCL GAALKDEVLK IMPVQKQTRA GQRTRFKAF VAIGDYNGHV GLGLKCSKEV ATAIRGAIIL AKLSIVPVRR GYWGNKIGKP HTVPCKVTGR CGSVLVRLIP A PRGTGIVS ...String:
MADDAGAAGG PGGPGGPGNK EWLPVTKLGR LVKDMKIKSL EEIYLFSLPI KESEIIDFCL GAALKDEVLK IMPVQKQTRA GQRTRFKAF VAIGDYNGHV GLGLKCSKEV ATAIRGAIIL AKLSIVPVRR GYWGNKIGKP HTVPCKVTGR CGSVLVRLIP A PRGTGIVS APVPKKLLLM AGIDDCYTSA RGCTATLGNF AKATFDAISK TYSYLTPDLW KETVFTKSPY QEFTNHLMKT HT RVSVQRT QAPAVATT

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Macromolecule #5: uS3

MacromoleculeName: uS3 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 31.146607 KDa
SequenceString: MSARRRRRRA AFRRRAEPFI PISVREPLPF LSAARGGKMA VQISKKRKFV ADGIFKAELN EFLTRELAED GYSGVEVRVT PTRTEIIIL ATRTQNVLGE KGRRIRELTA VVQKRFGFPE GSVELYAEKV ATRGLCAIAQ AESLRYKLLG GLAVRRACYG V LRFIMESG ...String:
MSARRRRRRA AFRRRAEPFI PISVREPLPF LSAARGGKMA VQISKKRKFV ADGIFKAELN EFLTRELAED GYSGVEVRVT PTRTEIIIL ATRTQNVLGE KGRRIRELTA VVQKRFGFPE GSVELYAEKV ATRGLCAIAQ AESLRYKLLG GLAVRRACYG V LRFIMESG AKGCEVVVSG KLRGQRAKSM KFVDGLMIHS GDPVNYYVDT AVRHVLLRQG VLGIKVKIML PWDPSGKIGP KK PLPDHVS IVEPKDEILP TTPISEQKGG KPEPPAMPQP VPTA

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Macromolecule #7: eS4

MacromoleculeName: eS4 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 29.65892 KDa
SequenceString: MARGPKKHLK RVAAPKHWML DKLTSVFAPR PSTGPHKLRE CLPLIIFLRN KLKYALTGDE VKKICMQRFI KIDGKVRADI TYPAGFMDV ISIDKTGENF RLIYDTKGRF AVHRITPEEA KYKLCKVRKI FVGTKGIPHL VTHDARTIRY PDPLIKMNDT I QIDLETGK ...String:
MARGPKKHLK RVAAPKHWML DKLTSVFAPR PSTGPHKLRE CLPLIIFLRN KLKYALTGDE VKKICMQRFI KIDGKVRADI TYPAGFMDV ISIDKTGENF RLIYDTKGRF AVHRITPEEA KYKLCKVRKI FVGTKGIPHL VTHDARTIRY PDPLIKMNDT I QIDLETGK ITDFIKFDTG NLCMVTGGAN LGRIGVITNR ERHPGSFDVV HVKDANGNSF ATRLSNIFVI GKGNKPWISL PR GKGIRLT IAEERDKRLA AKQSSG

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Macromolecule #8: uS7

MacromoleculeName: uS7 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 22.913453 KDa
SequenceString: MTEWETAAPA VAETPDIKLF GKWSTDDVQI NDISLQDYIA VKEKYAKYLP HSAGRYAAKR FRKAQCPIVE RLTNSMMMHG RNNGKKLMT VRIVKHAFEI IHLLTGENPL QVLVNAIINS GPREDSTRIG RAGTVRRQAV DVSPLRRVNQ AIWLLCTGAR E AAFRNIKT ...String:
MTEWETAAPA VAETPDIKLF GKWSTDDVQI NDISLQDYIA VKEKYAKYLP HSAGRYAAKR FRKAQCPIVE RLTNSMMMHG RNNGKKLMT VRIVKHAFEI IHLLTGENPL QVLVNAIINS GPREDSTRIG RAGTVRRQAV DVSPLRRVNQ AIWLLCTGAR E AAFRNIKT IAECLADELI NAAKGSSNSY AIKKKDELER VAKSNR

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Macromolecule #9: eS6

MacromoleculeName: eS6 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 28.751906 KDa
SequenceString: MKLNISFPAT GCQKLIEVDD ERKLRTFYEK RMATEVAADA LGEEWKGYVV RISGGNDKQG FPMKQGVLTH GRVRLLLSKG HSCYRPRRT GERKRKSVRG CIVDANLSVL NLVIVKKGEK DIPGLTDTTV PRRLGPKRAS RIRKLFNLSK EDDVRQYVVR K PLNKEGKK ...String:
MKLNISFPAT GCQKLIEVDD ERKLRTFYEK RMATEVAADA LGEEWKGYVV RISGGNDKQG FPMKQGVLTH GRVRLLLSKG HSCYRPRRT GERKRKSVRG CIVDANLSVL NLVIVKKGEK DIPGLTDTTV PRRLGPKRAS RIRKLFNLSK EDDVRQYVVR K PLNKEGKK PRTKAPKIQR LVTPRVLQHK RRRIALKKQR TKKNKEEAAE YAKLLAKRMK EAKEKRQEQI AKRRRLSSLR AS TSKSESS QK

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Macromolecule #10: eS7

MacromoleculeName: eS7 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 22.168914 KDa
SequenceString: MFSSSAKIVK PNGEKPDEFE SGISQALLEL EMNSDLKAQL RELNITAAKE IEVGGGRKAI IIFVPVPQLK SFQKIQVRLV RELEKKFSG KHVVFIAQRR ILPKPTRKSR TKNKQKRPRS RTLTAVHDAI LEDLVFPSEI VGKRIRVKLD GSRLIKVHLD K AQQNNVEH ...String:
MFSSSAKIVK PNGEKPDEFE SGISQALLEL EMNSDLKAQL RELNITAAKE IEVGGGRKAI IIFVPVPQLK SFQKIQVRLV RELEKKFSG KHVVFIAQRR ILPKPTRKSR TKNKQKRPRS RTLTAVHDAI LEDLVFPSEI VGKRIRVKLD GSRLIKVHLD K AQQNNVEH KVETFSGVYK KLTGKDVNFE FPEFQL

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Macromolecule #11: eS8

MacromoleculeName: eS8 / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 24.134207 KDa
SequenceString: MGISRDNWHK RRKTGGKRKP YHKKRKYELG RPAANTKIGP RRIHTVRVRG GNKKYRALRL DVGNFSWGSE CCTRKTRIID VVYNASNNE LVRTKTLVKN CIVLIDSTPY RQWYESHYAL PLGRKKGAKL TPEEEEILNK KRSKKIQKKY DERKKNAKIS S LLEEQFQQ ...String:
MGISRDNWHK RRKTGGKRKP YHKKRKYELG RPAANTKIGP RRIHTVRVRG GNKKYRALRL DVGNFSWGSE CCTRKTRIID VVYNASNNE LVRTKTLVKN CIVLIDSTPY RQWYESHYAL PLGRKKGAKL TPEEEEILNK KRSKKIQKKY DERKKNAKIS S LLEEQFQQ GKLLACIASR PGQCGRADGY VLEGKELEFY LRKIKARKG

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Macromolecule #12: uS4

MacromoleculeName: uS4 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 22.641564 KDa
SequenceString: MPVARSWVCR KTYVTPRRPF EKSRLDQELK LIGEYGLRNK REVWRVKFTL AKIRKAAREL LTLDEKDPRR LFEGNALLRR LVRIGVLDE GKMKLDYILG LKIEDFLERR LQTQVFKLGL AKSIHHARVL IRQRHIRVRK QVVNIPSFIV RLDSQKHIDF S LRSPYGGG ...String:
MPVARSWVCR KTYVTPRRPF EKSRLDQELK LIGEYGLRNK REVWRVKFTL AKIRKAAREL LTLDEKDPRR LFEGNALLRR LVRIGVLDE GKMKLDYILG LKIEDFLERR LQTQVFKLGL AKSIHHARVL IRQRHIRVRK QVVNIPSFIV RLDSQKHIDF S LRSPYGGG RPGRVKRKNA KKGQGGAGAG DDEEED

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Macromolecule #13: eS10

MacromoleculeName: eS10 / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 17.156865 KDa
SequenceString:
MLMPKKNRIA IYELLFKEGV MVAKKDVHMP KHPELADKNV PNLHVMKAMQ SLKSRGYVKE QFAWRHFYWY LTNEGIQYLR DYLHLPPEI VPATLRRSRP ETGRPRPKGL EGERPARLTR GEADRDTYRR SAVPPGADKK AEAGAGSATE

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Macromolecule #14: uS17

MacromoleculeName: uS17 / type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 18.468826 KDa
SequenceString:
MADIQTERAY QKQPTIFQNK KRVLLGETGK EKLPRYYKNI GLGFKTPKEA IEGTYIDKKC PFTGNVSIRG RILSGVVTKM KMQRTIVIR RDYLHYIRKY NRFEKRHKNM SVHLSPCFRD VQIGDIVTVG ECRPLSKTVR FNVLKVTKAA GTKKQFQKF

+
Macromolecule #15: eS12

MacromoleculeName: eS12 / type: protein_or_peptide / ID: 15 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 14.538987 KDa
SequenceString:
MAEEGIAAGG VMDVNTALQE VLKTALIHDG LARGIREAAK ALDKRQAHLC VLASNCDEPM YVKLVEALCA EHQINLIKVD DNKKLGEWV GLCKIDREGK PRKVVGCSCV VVKDYGKESQ AKDVIEEYFK CKK

+
Macromolecule #16: uS15

MacromoleculeName: uS15 / type: protein_or_peptide / ID: 16 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 17.259389 KDa
SequenceString:
MGRMHAPGKG LSQSALPYRR SVPTWLKLTS DDVKEQIYKL AKKGLTPSQI GVILRDSHGV AQVRFVTGNK ILRILKSKGL APDLPEDLY HLIKKAVAVR KHLERNRKDK DAKFRLILIE SRIHRLARYY KTKRVLPPNW KYESSTASAL VA

+
Macromolecule #17: uS11

MacromoleculeName: uS11 / type: protein_or_peptide / ID: 17 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 16.302772 KDa
SequenceString:
MAPRKGKEKK EEQVISLGPQ VAEGENVFGV CHIFASFNDT FVHVTDLSGK ETICRVTGGM KVKADRDESS PYAAMLAAQD VAQRCKELG ITALHIKLRA TGGNRTKTPG PGAQSALRAL ARSGMKIGRI EDVTPIPSDS TRRKGGRRGR RL

+
Macromolecule #18: uS19

MacromoleculeName: uS19 / type: protein_or_peptide / ID: 18 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 17.049182 KDa
SequenceString:
MAEVEQKKKR TFRKFTYRGV DLDQLLDMSY EQLMQLYSAR QRRRLSRGLR RKQHSLLKRL RKAKKEAPPM EKPEVVKTHL RDMIILPEM VGSMVGVYNG KTFNQVEIKP EMIGHYLGEF SITYKPVKHG RPGIGATHSS RFIPLK

+
Macromolecule #19: uS9

MacromoleculeName: uS9 / type: protein_or_peptide / ID: 19 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 19.213465 KDa
SequenceString:
SARVAPRFPF PVVAPRGEDR HSSAAAMPSK GPLQSVQVFG RKKTATAVAH CKRGNGLIKV NGRPLEMIEP RTLQYKLLEP VLLLGKERF AGVDIRVRVK GGGHVAQIYA IRQSISKALV AYYQKYVDEA SKKEIKDILI QYDRTLLVAD PRRCESKKFG G PGARARYQ KSYR

+
Macromolecule #20: eS17

MacromoleculeName: eS17 / type: protein_or_peptide / ID: 20 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 15.552119 KDa
SequenceString:
MGRVRTKTVK KAARVIIEKY YTRLGNDFHT NKRVCEEIAI IPSKKLRNKI AGYVTHLMKR IQRGPVRGIS IKLQEEERER RDNYVPEVS ALDQEIIEVD PDTKEMLKLL DFGSLSNLQV TQPTVGMNFK TPRGAV

+
Macromolecule #21: uS13

MacromoleculeName: uS13 / type: protein_or_peptide / ID: 21 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 17.759777 KDa
SequenceString:
MSLVIPEKFQ HILRVLNTNI DGRRKIAFAI TAIKGVGRRY AHVVLRKADI DLTKRAGELT EDEVERVITI MQNPRQYKIP DWFLNRQKD VKDGKYSQVL ANGLDNKLRE DLERLKKIRA HRGLRHFWGL RVRGQHTKTT GRRGRTVGVS KKK

+
Macromolecule #22: eS19

MacromoleculeName: eS19 / type: protein_or_peptide / ID: 22 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 16.235796 KDa
SequenceString:
MPGVTVKDVN QQEFVRALAA FLKKSGKLKV PEWVDTVKLA KHKELAPYDE NWFYTRAAST ARHLYLRGGA GVGSMTKIYG GRQRNGVMP SHFSRGSKSV ARRVLQALEG LKMVEKDQDW GRKLTPQGQR DLDRIAGQVA AAKKKH

+
Macromolecule #23: uS10

MacromoleculeName: uS10 / type: protein_or_peptide / ID: 23 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 13.398763 KDa
SequenceString:
MAFKDTGKTP VEPEVAIHRI RITLTSRNVK SLEKVCADLI RGAKEKNLKV KGPVRMPTKT LRITTRKTPC GEGSKTWDRF QMRIHKRLI DLHSPSEIVK QITSISIEPG VEVEVTIADA

+
Macromolecule #24: eS21

MacromoleculeName: eS21 / type: protein_or_peptide / ID: 24 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 9.043276 KDa
SequenceString:
MQSNAGEFVD LYVPRKCSAS NRIIGAKDHA SIPMNVAEVD KVTGRFNGQS KTYAICGAIR RMGESDDSIL RLAKSHGIVS QNF

+
Macromolecule #25: uS8

MacromoleculeName: uS8 / type: protein_or_peptide / ID: 25 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 14.865555 KDa
SequenceString:
MVRMNVLADA LKSINNAEKR GKRQVLIRPC SKVIVRFLTV MMKHGYIGEF EIIDDHRAGK IVVNLTGRLN KCGVISPRFD VQLKDLEKW QNNLLPSRQF GFIVLTTSAG IMDHEEARRK HTGGKILGFF F

+
Macromolecule #26: uS12

MacromoleculeName: uS12 / type: protein_or_peptide / ID: 26 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 15.784549 KDa
SequenceString:
AGKCRGLRTA RKLRSHRRDQ KWHDKQYKKA HLGTALKANP FGGASHAKGI VLEKVGVEAK QPNSAIRKCV RVQLIKNGKK ITAFVPNDG CLNFIEENDE VLVAGFGRKG HAVGDIPGVR FKVVKVANVS LLALYKGKKE RPRS

+
Macromolecule #27: eS24

MacromoleculeName: eS24 / type: protein_or_peptide / ID: 27 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 15.548398 KDa
SequenceString:
MNDTVTIRTR KFMTNRLLQR KQMVIDVLHP GKATVPKTEI REKLAKMYKT TPDVIFVFGF RTHFGGGKTT GFGMIYDSLD YAKKNEPKH RLARHGLYEK KKTSRKQRKE RKNRMKKVRG TAKANVGAGK KEPRG

+
Macromolecule #28: eS25

MacromoleculeName: eS25 / type: protein_or_peptide / ID: 28 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 13.776224 KDa
SequenceString:
MPPKDDKKKK DAGKSAKKDK DPVNKSGGKA KKKKWSKGKV RDKLNNLVLF DKATYDKLCK EVPNYKLITP AVVSERLKIR GSLARAALQ ELLSKGLIKL VSKHRAQVIY TRNTKGGDAP AAGEDA

+
Macromolecule #29: eS26

MacromoleculeName: eS26 / type: protein_or_peptide / ID: 29 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 12.961455 KDa
SequenceString:
MTKKRRNNGR AKKGRGHVQP IRCTNCACCV PKDKAIKKFV IRNIVEAAAV RDISEVSVFD AYVLPKLYVK LHYCVSCAIH SKVVRNRSR EARKDRTPPP RFRPAGAAPP PPPKPM

+
Macromolecule #30: eS27

MacromoleculeName: eS27 / type: protein_or_peptide / ID: 30 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 9.480186 KDa
SequenceString:
MPLAKDLLHP SPEEEKRKHK KKRLVQSPNS YFMDVKCPGC YKITTVFSHA QTVVLCVGCS TVLCQPTGGK ARLTEGCSFR RKQH

+
Macromolecule #31: eS28

MacromoleculeName: eS28 / type: protein_or_peptide / ID: 31 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 7.855052 KDa
SequenceString:
MDTSRVQPIK LARVTKVLGR TGSQGQCTQV RVEFMDDTSR SIIRNVKGPV REGDVLTLLE SEREARRLR

+
Macromolecule #32: eS29

MacromoleculeName: eS29 / type: protein_or_peptide / ID: 32 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 6.690821 KDa
SequenceString:
MGHQQLYWSH PRKFGQGSRS CRVCSNRHGL IRKYGLNMCR QCFRQYAKDI GFIKLD

+
Macromolecule #33: eS30

MacromoleculeName: eS30 / type: protein_or_peptide / ID: 33 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 14.498884 KDa
SequenceString:
MQLFVRAQEL HTLEVTGRET VAQIKAHVAS LEGIAPEDQV VLLAGTPLED EATLGQCGVE ALSTLEVAGR MLGGKVHGSL ARVGKVRGQ TLKVAKQEKK KKRTGRAKRR MQYNRRFVNV VPTFGKKKGP NANS

+
Macromolecule #34: eS31

MacromoleculeName: eS31 / type: protein_or_peptide / ID: 34 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 18.004041 KDa
SequenceString:
MQIFVKTLTG KTITLEVEPS DTIENVKAKI QDKEGIPPDQ QRLIFAGKQL EDGRTLSDYN IQKESTLHLV LRLRGGAKKR KKKSYTTPK KNKHKRKKVK LAVLKYYKVD ENGKISRLRR ECPSDECGAG VFMASHFDRH YCGKCCLTYC FNKPEDK

+
Macromolecule #35: RACK1

MacromoleculeName: RACK1 / type: protein_or_peptide / ID: 35 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 35.115652 KDa
SequenceString: MTEQMTLRGT LKGHNGWVTQ IATTPQFPDM ILSASRDKTI IMWKLTRDET NYGIPQRALR GHSHFVSDVV ISSDGQFALS GSWDGTLRL WDLTTGTTTR RFVGHTKDVL SVAFSSDNRQ IVSGSRDKTI KLWNTLGVCK YTVQDESHSE WVSCVRFSPN S SNPIIVSC ...String:
MTEQMTLRGT LKGHNGWVTQ IATTPQFPDM ILSASRDKTI IMWKLTRDET NYGIPQRALR GHSHFVSDVV ISSDGQFALS GSWDGTLRL WDLTTGTTTR RFVGHTKDVL SVAFSSDNRQ IVSGSRDKTI KLWNTLGVCK YTVQDESHSE WVSCVRFSPN S SNPIIVSC GWDKLVKVWN LANCKLKTNH IGHTGYLNTV TVSPDGSLCA SGGKDGQAML WDLNEGKHLY TLDGGDIINA LC FSPNRYW LCAATGPSIK IWDLEGKIIV DELKQEVIST SSKAEPPQCT SLAWSADGQT LFAGYTDNLV RVWQVTIGTR

+
Macromolecule #36: eL41

MacromoleculeName: eL41 / type: protein_or_peptide / ID: 36 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rabbit (rabbit)
Molecular weightTheoretical: 3.473451 KDa
SequenceString:
MRAKWRKKRM RRLKRKRRKM RQRSK

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Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation #1

Preparation ID1
BufferpH: 7.5
Component:
ConcentrationFormula
20.0 mMTris-HClTris
100.0 mMKCL
8.0 mMMgCl2
2.0 mMDTT
GridModel: Quantifoil R2/2 / Material: COPPER / Support film - Film type ID: 1 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 5.0 nm / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 9.33257 kPa
Details: Plasma cleaning for both holey carbon and holey gold grids was done on a Gatan Solarus with Hydrogen (6.4 sccm gas flow) and Oxygen (27.5 sccm gas flow) and 10 W cleaning power.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
DetailsRibosomal complexes for the pre-translocated state were assembled at 240-390 nM concentration and applied to plasma treated holey carbon.

-
Sample preparation #2

Preparation ID2
BufferpH: 7.5
Component:
ConcentrationFormula
20.0 mMTris-HClTris
100.0 mMKCL
8.0 mMMgCl2
2.0 mMDTT
GridModel: Quantifoil R2/2 / Material: COPPER / Support film - Film type ID: 1 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 5.0 nm / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 9.33257 kPa
Details: Plasma cleaning for both holey carbon and holey gold grids was done on a Gatan Solarus with Hydrogen (6.4 sccm gas flow) and Oxygen (27.5 sccm gas flow) and 10 W cleaning power.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
DetailsSame sample and buffer conditions used for the holey carbon grids was used for the holey gold grids.

-
Electron microscopy

MicroscopeFEI TECNAI F30
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.26 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 31000
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Frames/image: 1-40 / Number real images: 11234 / Average exposure time: 8.0 sec. / Average electron dose: 42.09 e/Å2
Experimental equipment
Model: Tecnai F30 / Image courtesy: FEI Company

-
Image processing

Particle selectionNumber selected: 1240275
CTF correctionSoftware - Name: Gctf
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5it9

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 96826

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