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- EMDB-12179: Staphylococcus aureus 30S ribosomal subunit in presence of spermi... -

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Basic information

Entry
Database: EMDB / ID: EMD-12179
TitleStaphylococcus aureus 30S ribosomal subunit in presence of spermidine (head only)
Map data
Sample
  • Complex: Staphylococcus aureus 30S ribosomal subunit in presence of spermidine (head only)
    • RNA: 16S ribosomal RNA
    • Protein or peptide: 30S ribosomal protein S2
    • Protein or peptide: 30S ribosomal protein S3
    • Protein or peptide: 30S ribosomal protein S7
    • Protein or peptide: 30S ribosomal protein S9
    • Protein or peptide: 30S ribosomal protein S10
    • Protein or peptide: 30S ribosomal protein S13
    • Protein or peptide: 30S ribosomal protein S14 type ZRibosome
    • Protein or peptide: 30S ribosomal protein S19
Function / homology
Function and homology information


ribosomal small subunit assembly / cytosolic small ribosomal subunit / small ribosomal subunit / tRNA binding / rRNA binding / ribosome / structural constituent of ribosome / translation / ribonucleoprotein complex / mRNA binding ...ribosomal small subunit assembly / cytosolic small ribosomal subunit / small ribosomal subunit / tRNA binding / rRNA binding / ribosome / structural constituent of ribosome / translation / ribonucleoprotein complex / mRNA binding / RNA binding / zinc ion binding / cytosol
Similarity search - Function
Ribosomal protein S14, type Z / Ribosomal protein S14/S29 / Ribosomal protein S3, bacterial-type / Ribosomal protein S19, bacterial-type / Ribosomal protein S7, bacterial/organellar-type / Ribosomal protein S13, bacterial-type / Ribosomal protein S9, bacterial/plastid / Ribosomal protein S2, bacteria/mitochondria/plastid / Ribosomal protein S2 signature 2. / K Homology domain ...Ribosomal protein S14, type Z / Ribosomal protein S14/S29 / Ribosomal protein S3, bacterial-type / Ribosomal protein S19, bacterial-type / Ribosomal protein S7, bacterial/organellar-type / Ribosomal protein S13, bacterial-type / Ribosomal protein S9, bacterial/plastid / Ribosomal protein S2, bacteria/mitochondria/plastid / Ribosomal protein S2 signature 2. / K Homology domain / K homology RNA-binding domain / Ribosomal protein S3, conserved site / Ribosomal protein S14, conserved site / Ribosomal protein S10, conserved site / K Homology domain, type 2 / Ribosomal protein S3, C-terminal / Ribosomal protein S3, C-terminal domain superfamily / Ribosomal protein S15/S19, conserved site / KH domain / Ribosomal protein S19/S15 / Ribosomal protein S19/S15, superfamily / Ribosomal protein S10 / Ribosomal protein S3, C-terminal domain / Ribosomal protein S3 signature. / Ribosomal protein S10 signature. / Ribosomal protein S14 signature. / Ribosomal protein S7, conserved site / K homology domain superfamily, prokaryotic type / Ribosomal protein S19 / Ribosomal protein S2 signature 1. / Ribosomal protein S13, conserved site / Ribosomal protein S2, conserved site / Ribosomal protein S13 / 30s ribosomal protein S13, C-terminal / Ribosomal protein S2 / Ribosomal protein S2, flavodoxin-like domain superfamily / Ribosomal protein S14 / Ribosomal protein S2 / Type-2 KH domain profile. / Ribosomal protein S13/S18 / Ribosomal protein S19 signature. / K homology domain-like, alpha/beta / Ribosomal protein S14p/S29e / Ribosomal protein S7 signature. / Ribosomal protein S10p/S20e / Ribosomal protein S13-like, H2TH / Ribosomal protein S9, conserved site / Ribosomal protein S10 domain / Ribosomal protein S10 domain superfamily / Ribosomal protein S13 signature. / Ribosomal protein S5/S7 / Ribosomal protein S7 domain / Ribosomal protein S7 domain superfamily / Ribosomal protein S13 family profile. / Ribosomal protein S10p/S20e / Ribosomal protein S9 / Ribosomal protein S7p/S5e / Ribosomal protein S9/S16 / Ribosomal protein S9 signature. / Ribosomal protein S5 domain 2-type fold, subgroup / Ribosomal protein S5 domain 2-type fold
Similarity search - Domain/homology
Small ribosomal subunit protein uS7 / Small ribosomal subunit protein uS19 / Small ribosomal subunit protein uS3 / Small ribosomal subunit protein uS14B / Small ribosomal subunit protein uS13 / Small ribosomal subunit protein uS9 / Small ribosomal subunit protein uS2 / Small ribosomal subunit protein uS10
Similarity search - Component
Biological speciesStaphylococcus aureus subsp. aureus NCTC 8325 (bacteria) / Staphylococcus aureus (strain NCTC 8325) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsBelinite M / Khusainov I / Marzi S / Romby P / Yusupov M / Hashem Y
Funding support France, 2 items
OrganizationGrant numberCountry
Laboratories of Excellence (LabEx)ANR-10-LABX-0036 NETRNA France
Agence Nationale de la Recherche (ANR)ANR-16-CE11-0007-01 France
CitationJournal: Front Mol Biosci / Year: 2021
Title: Stabilization of Ribosomal RNA of the Small Subunit by Spermidine in .
Authors: Margarita Belinite / Iskander Khusainov / Heddy Soufari / Stefano Marzi / Pascale Romby / Marat Yusupov / Yaser Hashem /
Abstract: Cryo-electron microscopy is now used as a method of choice in structural biology for studying protein synthesis, a process mediated by the ribosome machinery. In order to achieve high-resolution ...Cryo-electron microscopy is now used as a method of choice in structural biology for studying protein synthesis, a process mediated by the ribosome machinery. In order to achieve high-resolution structures using this approach, one needs to obtain homogeneous and stable samples, which requires optimization of ribosome purification in a species-dependent manner. This is especially critical for the bacterial small ribosomal subunit that tends to be unstable in the absence of ligands. Here, we report a protocol for purification of stable 30 S from the Gram-positive bacterium and its cryo-EM structures: in presence of spermidine at a resolution ranging between 3.4 and 3.6 Å and in its absence at 5.3 Å. Using biochemical characterization and cryo-EM, we demonstrate the importance of spermidine for stabilization of the 30 S preserving favorable conformation of the helix 44.
History
DepositionJan 6, 2021-
Header (metadata) releaseDec 8, 2021-
Map releaseDec 8, 2021-
UpdateDec 8, 2021-
Current statusDec 8, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0282
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.0282
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7bge
  • Surface level: 0.0282
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_12179.png

Downloads & links

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Map

FileDownload / File: emd_12179.map.gz / Format: CCP4 / Size: 75.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.19 Å
Density
Contour LevelBy AUTHOR: 0.0282 / Movie #1: 0.0282
Minimum - Maximum-0.18268038 - 0.32439134
Average (Standard dev.)0.00038017353 (±0.0062601836)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions270270270
Spacing270270270
CellA=B=C: 321.30002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.191.191.19
M x/y/z270270270
origin x/y/z0.0000.0000.000
length x/y/z321.300321.300321.300
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS270270270
D min/max/mean-0.1830.3240.000

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Supplemental data

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Sample components

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Entire : Staphylococcus aureus 30S ribosomal subunit in presence of spermi...

EntireName: Staphylococcus aureus 30S ribosomal subunit in presence of spermidine (head only)
Components
  • Complex: Staphylococcus aureus 30S ribosomal subunit in presence of spermidine (head only)
    • RNA: 16S ribosomal RNA
    • Protein or peptide: 30S ribosomal protein S2
    • Protein or peptide: 30S ribosomal protein S3
    • Protein or peptide: 30S ribosomal protein S7
    • Protein or peptide: 30S ribosomal protein S9
    • Protein or peptide: 30S ribosomal protein S10
    • Protein or peptide: 30S ribosomal protein S13
    • Protein or peptide: 30S ribosomal protein S14 type ZRibosome
    • Protein or peptide: 30S ribosomal protein S19

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Supramolecule #1: Staphylococcus aureus 30S ribosomal subunit in presence of spermi...

SupramoleculeName: Staphylococcus aureus 30S ribosomal subunit in presence of spermidine (head only)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Staphylococcus aureus subsp. aureus NCTC 8325 (bacteria)

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Macromolecule #1: 16S ribosomal RNA

MacromoleculeName: 16S ribosomal RNA / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Staphylococcus aureus subsp. aureus NCTC 8325 (bacteria)
Molecular weightTheoretical: 503.218031 KDa
SequenceString: UUUUCUGGAG AGUUUGAUCC UGGCUCAGGA UGAACGCUGG CGGCGUGCCU AAUACAUGCA AGUCGAGCGA ACGGACGAGA AGCUUGCUU CUCUGAUGUU AGCGGCGGAC GGGUGAGUAA CACGUGGAUA ACCUACCUAU AAGACUGGGA UAACUUCGGG A AACCGUAG ...String:
UUUUCUGGAG AGUUUGAUCC UGGCUCAGGA UGAACGCUGG CGGCGUGCCU AAUACAUGCA AGUCGAGCGA ACGGACGAGA AGCUUGCUU CUCUGAUGUU AGCGGCGGAC GGGUGAGUAA CACGUGGAUA ACCUACCUAU AAGACUGGGA UAACUUCGGG A AACCGUAG CUAAUACCGG AUAAUAUUUU GAACCGCAUG GUUCAAAAGU GAAAGACGGU CUUGCUGUCA CUUAUAGAUG GA UCCGCGC UGCAUUAGCU AGUUGGUAAG GUAACGGCUU ACCAAGGCAA CGAUGCAUAG CCGACCUGAG AGGGUGAUCG GCC ACACUG GAACUGAGAC ACGGUCCAGA CUCCUACGGG AGGCAGCAGU AGGGAAUCUU CCGCAAUGGG CGAAAGCCUG ACGG AGCAA CGCCGCGUGA GUGAUGAAGG UCUUCGGAUC GUAAAACUCU GUUAUUAGGG AAGAACAUAU GUGUAAGUAA CUGUG CACA UCUUGACGGU ACCUAAUCAG AAAGCCACGG CUAACUACGU GCCAGCAGCC GCGGUAAUAC GUAGGUGGCA AGCGUU AUC CGGAAUUAUU GGGCGUAAAG CGCGCGUAGG CGGUUUUUUA AGUCUGAUGU GAAAGCCCAC GGCUCAACCG UGGAGGG UC AUUGGAAACU GGAAAACUUG AGUGCAGAAG AGGAAAGUGG AAUUCCAUGU GUAGCGGUGA AAUGCGCAGA GAUAUGGA G GAACACCAGU GGCGAAGGCG ACUUUCUGGU CUGUAACUGA CGCUGAUGUG CGAAAGCGUG GGGAUCAAAC AGGAUUAGA UACCCUGGUA GUCCACGCCG UAAACGAUGA GUGCUAAGUG UUAGGGGGUU UCCCGCCCCU UAGUGCUGCA GCUAACGCAU UAAGCACUC CGCCUGGGGA GUACGACCGC AAGGUUGAAA CUCAAAGGAA UUGACGGGGA CCCGCACAAG CGGUGGAGCA U GUGGUUUA AUUCGAAGCA ACGCGAAGAA CCUUACCAAA UCUUGACAUC CUUUGACAAC UCUAGAGAUA GAGCCUUCCC CU UCGGGGG ACAAAGUGAC AGGUGGUGCA UGGUUGUCGU CAGCUCGUGU CGUGAGAUGU UGGGUUAAGU CCCGCAACGA GCG CAACCC UUAAGCUUAG UUGCCAUCAU UAAGUUGGGC ACUCUAAGUU GACUGCCGGU GACAAACCGG AGGAAGGUGG GGAU GACGU CAAAUCAUCA UGCCCCUUAU GAUUUGGGCU ACACACGUGC UACAAUGGAC AAUACAAAGG GCAGCGAAAC CGCGA GGUC AAGCAAAUCC CAUAAAGUUG UUCUCAGUUC GGAUUGUAGU CUGCAACUCG ACUACAUGAA GCUGGAAUCG CUAGUA AUC GUAGAUCAGC AUGCUACGGU GAAUACGUUC CCGGGUCUUG UACACACCGC CCGUCACACC ACGAGAGUUU GUAACAC CC GAAGCCGGUG GAGUAACCUU UUAGGAGCUA GCCGUCGAAG GUGGGACAAA UGAUUGGGGU GAAGUCGUAA CAAGGUAG C CGUAUCGGAA GGUGCGGCUG GAUCACCUCC UUUCU

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Macromolecule #2: 30S ribosomal protein S2

MacromoleculeName: 30S ribosomal protein S2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 29.136369 KDa
SequenceString: MAVISMKQLL EAGVHFGHQT RRWNPKMKKY IFTERNGIYI IDLQKTVKKV DEAYNFLKQV SEDGGQVLFV GTKKQAQESV KSEAERAGQ FYINQRWLGG LLTNYKTISK RIKRISEIEK MEEDGLFEVL PKKEVVELKK EYDRLIKFLG GIRDMKSMPQ A LFVVDPRK ...String:
MAVISMKQLL EAGVHFGHQT RRWNPKMKKY IFTERNGIYI IDLQKTVKKV DEAYNFLKQV SEDGGQVLFV GTKKQAQESV KSEAERAGQ FYINQRWLGG LLTNYKTISK RIKRISEIEK MEEDGLFEVL PKKEVVELKK EYDRLIKFLG GIRDMKSMPQ A LFVVDPRK ERNAIAEARK LNIPIVGIVD TNCDPDEIDY VIPANDDAIR AVKLLTAKMA DAILEGQQGV SNEEVAAEQN ID LDEKEKS EETEATEE

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Macromolecule #3: 30S ribosomal protein S3

MacromoleculeName: 30S ribosomal protein S3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 24.143867 KDa
SequenceString: MGQKINPIGL RVGIIRDWEA KWYAEKDFAS LLHEDLKIRK FIDNELKEAS VSHVEIERAA NRINIAIHTG KPGMVIGKGG SEIEKLRNK LNALTDKKVH INVIEIKKVD LDARLVAENI ARQLENRASF RRVQKQAITR AMKLGAKGIK TQVSGRLGGA D IARAEQYS ...String:
MGQKINPIGL RVGIIRDWEA KWYAEKDFAS LLHEDLKIRK FIDNELKEAS VSHVEIERAA NRINIAIHTG KPGMVIGKGG SEIEKLRNK LNALTDKKVH INVIEIKKVD LDARLVAENI ARQLENRASF RRVQKQAITR AMKLGAKGIK TQVSGRLGGA D IARAEQYS EGTVPLHTLR ADIDYAHAEA DTTYGKLGVK VWIYRGEVLP TKNTSGGGK

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Macromolecule #4: 30S ribosomal protein S7

MacromoleculeName: 30S ribosomal protein S7 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 17.826555 KDa
SequenceString:
MPRKGSVPKR DVLPDPIHNS KLVTKLINKI MLDGKRGTAQ RILYSAFDLV EQRSGRDALE VFEEAINNIM PVLEVKARRV GGSNYQVPV EVRPERRTTL GLRWLVNYAR LRGEKTMEDR LANEILDAAN NTGGAVKKRE DTHKMAEANK AFAHYRW

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Macromolecule #5: 30S ribosomal protein S9

MacromoleculeName: 30S ribosomal protein S9 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 14.856987 KDa
SequenceString:
MTLAQVEYRG TGRRKNSVAR VRLVPGEGNI TVNNRDVREY LPFESLILDL NQPFDVTETK GNYDVLVNVH GGGFTGQAQA IRHGIARAL LEADPEYRGS LKRAGLLTRD PRMKERKKPG LKAARRSPQF SKR

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Macromolecule #6: 30S ribosomal protein S10

MacromoleculeName: 30S ribosomal protein S10 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus subsp. aureus NCTC 8325 (bacteria)
Molecular weightTheoretical: 11.60052 KDa
SequenceString:
MAKQKIRIRL KAYDHRVIDQ SAEKIVETAK RSGADVSGPI PLPTEKSVYT IIRAVHMYKD SREQFEQRTH KRLIDIVNPT PKTVDALMG LNLPSGVDIE IKL

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Macromolecule #7: 30S ribosomal protein S13

MacromoleculeName: 30S ribosomal protein S13 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 13.747919 KDa
SequenceString:
MARIAGVDIP REKRVVISLT YIYGIGTSTA QKILEEANVS ADTRVKDLTD DELGRIREVV DGYKVEGDLR RETNLNIKRL MEISSYRGI RHRRGLPVRG QKTKNNARTR KGPVKTVANK KK

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Macromolecule #8: 30S ribosomal protein S14 type Z

MacromoleculeName: 30S ribosomal protein S14 type Z / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 7.317769 KDa
SequenceString:
MAKTSMVAKQ QKKQKYAVRE YTRCERCGRP HSVYRKFKLC RICFRELAYK GQIPGVRKAS W

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Macromolecule #9: 30S ribosomal protein S19

MacromoleculeName: 30S ribosomal protein S19 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Staphylococcus aureus (strain NCTC 8325) (bacteria)
Strain: NCTC 8325
Molecular weightTheoretical: 10.639309 KDa
SequenceString:
MARSIKKGPF VDEHLMKKVE AQEGSEKKQV IKTWSRRSTI FPNFIGHTFA VYDGRKHVPV YVTEDMVGHK LGEFAPTRTF KGHVADDKK TRR

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 3.0 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Gctf (ver. 1.06)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5li0

Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 529602

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