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- EMDB-14560: Cryo-EM structure of Mycobacterium tuberculosis RNA polymerase core -

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Basic information

Entry
Database: EMDB / ID: EMD-14560
TitleCryo-EM structure of Mycobacterium tuberculosis RNA polymerase core
Map dataprimary map of RNAP core
Sample
  • Complex: DNA-dependent RNA polymerase core
    • Protein or peptide: DNA-directed RNA polymerase subunit alphaPolymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit betaPolymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit beta'Polymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit omegaPolymerase
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
Function / homology
Function and homology information


Antimicrobial action and antimicrobial resistance in Mtb / peptidoglycan-based cell wall / DNA-directed RNA polymerase complex / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / response to antibiotic / DNA-templated transcription / magnesium ion binding ...Antimicrobial action and antimicrobial resistance in Mtb / peptidoglycan-based cell wall / DNA-directed RNA polymerase complex / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / response to antibiotic / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase beta subunit, bacterial-type / RNA polymerase Rpb6 ...DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase beta subunit, bacterial-type / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb1, domain 3 superfamily / RNA polymerase Rpb1, clamp domain superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 4 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 5 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, RBP11-like subunit / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit omega / DNA-directed RNA polymerase subunit beta' / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase subunit alpha
Similarity search - Component
Biological speciesMycobacterium tuberculosis H37Rv (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.08 Å
AuthorsBrodolin K
Funding support France, 1 items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR-16-CE11-0025 France
Citation
Journal: Nat Commun / Year: 2023
Title: Structural basis of the mycobacterial stress-response RNA polymerase auto-inhibition via oligomerization
Authors: Morichaud Z / Trapani S / Vishwakarma RK / Chaloin L / Lionne C / Lai-Kee-Him J / Bron P / Brodolin K
#1: Journal: Biorxiv / Year: 2022
Title: Structural basis of the mycobacterial stress-response RNA polymerase auto-inhibition via oligomerization
Authors: Morichaud Z / Trapani S / Vishwakarma R / Chaloin L / Lionne C / Lai-Kee-Him J / Bron P / Brodolin K
History
DepositionMar 18, 2022-
Header (metadata) releaseFeb 8, 2023-
Map releaseFeb 8, 2023-
UpdateFeb 8, 2023-
Current statusFeb 8, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_14560.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationprimary map of RNAP core
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 320 pix.
= 352. Å
1.1 Å/pix.
x 320 pix.
= 352. Å
1.1 Å/pix.
x 320 pix.
= 352. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 0.24
Minimum - Maximum-0.4918646 - 1.3637666
Average (Standard dev.)-0.00060012966 (±0.04853187)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 352.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : DNA-dependent RNA polymerase core

EntireName: DNA-dependent RNA polymerase core
Components
  • Complex: DNA-dependent RNA polymerase core
    • Protein or peptide: DNA-directed RNA polymerase subunit alphaPolymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit betaPolymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit beta'Polymerase
    • Protein or peptide: DNA-directed RNA polymerase subunit omegaPolymerase
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION

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Supramolecule #1: DNA-dependent RNA polymerase core

SupramoleculeName: DNA-dependent RNA polymerase core / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1-#4
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Molecular weightTheoretical: 400 KDa

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Macromolecule #1: DNA-directed RNA polymerase subunit alpha

MacromoleculeName: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: ATCC 25618 / H37Rv
Molecular weightTheoretical: 37.745328 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEE DEPVTMYLRK QGPGEVTAGD IVPPAGVTVH NPGMHIATLN DKGKLEVELV VERGRGYVPA VQNRASGAEI G RIPVDSIY ...String:
MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEE DEPVTMYLRK QGPGEVTAGD IVPPAGVTVH NPGMHIATLN DKGKLEVELV VERGRGYVPA VQNRASGAEI G RIPVDSIY SPVLKVTYKV DATRVEQRTD FDKLILDVET KNSISPRDAL ASAGKTLVEL FGLARELNVE AEGIEIGPSP AE ADHIASF ALPIDDLDLT VRSYNCLKRE GVHTVGELVA RTESDLLDIR NFGQKSIDEV KIKLHQLGLS LKDSPPSFDP SEV AGYDVA TGTWSTEGAY DEQDYAETEQ L

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Macromolecule #2: DNA-directed RNA polymerase subunit beta

MacromoleculeName: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 2 / Details: L2E3G4C5 -> V / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: ATCC 25618 / H37Rv
Molecular weightTheoretical: 129.602344 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYEL SPIEDFSGSM SLSFSDPRFD DVKAPVDECK DKDMTYAAPL FVTAEFINNN TGEIKSQTVF MGDFPMMTEK G TFIINGTE ...String:
MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYEL SPIEDFSGSM SLSFSDPRFD DVKAPVDECK DKDMTYAAPL FVTAEFINNN TGEIKSQTVF MGDFPMMTEK G TFIINGTE RVVVSQLVRS PGVYFDETID KSTDKTLHSV KVIPSRGAWL EFDVDKRDTV GVRIDRKRRQ PVTVLLKALG WT SEQIVER FGFSEIMRST LEKDNTVGTD EALLDIYRKL RPGEPPTKES AQTLLENLFF KEKRYDLARV GRYKVNKKLG LHV GEPITS STLTEEDVVA TIEYLVRLHE GQTTMTVPGG VEVPVETDDI DHFGNRRLRT VGELIQNQIR VGMSRMERVV RERM TTQDV EAITPQTLIN IRPVVAAIKE FFGTSQLSQF MDQNNPLSGL THKRRLSALG PGGLSRERAG LEVRDVHPSH YGRMC PIET PEGPNIGLIG SLSVYARVNP FGFIETPYRK VVDGVVSDEI VYLTADEEDR HVVAQANSPI DADGRFVEPR VLVRRK AGE VEYVPSSEVD YMDVSPRQMV SVATAMIPFL EHDDANRALM GANMQRQAVP LVRSEAPLVG TGMELRAAID AGDVVVA EE SGVIEEVSAD YITVMHDNGT RRTYRMRKFA RSNHGTCANQ CPIVDAGDRV EAGQVIADGP CTDDGEMALG KNLLVAIM P WEGHNYEDAI ILSNRLVEED VLTSIHIEEH EIDARDTKLG AEEITRDIPN ISDEVLADLD ERGIVRIGAE VRDGDILVG KVTPKGETEL TPEERLLRAI FGEKAREVRD TSLKVPHGES GKVIGIRVFS REDEDELPAG VNELVRVYVA QKRKISDGDK LAGRHGNKG VIGKILPVED MPFLADGTPV DIILNTHGVP RRMNIGQILE THLGWCAHSG WKVDAAKGVP DWAARLPDEL L EAQPNAIV STPVFDGAQE AELQGLLSCT LPNRDGDVLV DADGKAMLFD GRSGEPFPYP VTVGYMYIMK LHHLVDDKIH AR STGPYSM ITQQPLGGKA QFGGQRFGEM ECWAMQAYGA AYTLQELLTI KSDDTVGRVK VYEAIVKGEN IPEPGIPESF KVL LKELQS LCLNVEVLSS DGAAIELREG EDEDLERAAA NLGINLSRNE SASVEDLA

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Macromolecule #3: DNA-directed RNA polymerase subunit beta'

MacromoleculeName: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: ATCC 25618 / H37Rv
Molecular weightTheoretical: 147.438344 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: VNFFDELRIG LATAEDIRQW SYGEVKKPET INYRTLKPEK DGLFCEKIFG PTRDWECYCG KYKRVRFKGI ICERCGVEVT RAKVRRERM GHIELAAPVT HIWYFKGVPS RLGYLLDLAP KDLEKIIYFA AYVITSVDEE MRHNELSTLE AEMAVERKAV E DQRDGELE ...String:
VNFFDELRIG LATAEDIRQW SYGEVKKPET INYRTLKPEK DGLFCEKIFG PTRDWECYCG KYKRVRFKGI ICERCGVEVT RAKVRRERM GHIELAAPVT HIWYFKGVPS RLGYLLDLAP KDLEKIIYFA AYVITSVDEE MRHNELSTLE AEMAVERKAV E DQRDGELE ARAQKLEADL AELEAEGAKA DARRKVRDGG EREMRQIRDR AQRELDRLED IWSTFTKLAP KQLIVDENLY RE LVDRYGE YFTGAMGAES IQKLIENFDI DAEAESLRDV IRNGKGQKKL RALKRLKVVA AFQQSGNSPM GMVLDAVPVI PPE LRPMVQ LDGGRFATSD LNDLYRRVIN RNNRLKRLID LGAPEIIVNN EKRMLQESVD ALFDNGRRGR PVTGPGNRPL KSLS DLLKG KQGRFRQNLL GKRVDYSGRS VIVVGPQLKL HQCGLPKLMA LELFKPFVMK RLVDLNHAQN IKSAKRMVER QRPQV WDVL EEVIAEHPVL LNRAPTLHRL GIQAFEPMLV EGKAIQLHPL VCEAFNADFD GDQMAVHLPL SAEAQAEARI LMLSSN NIL SPASGRPLAM PRLDMVTGLY YLTTEVPGDT GEYQPASGDH PETGVYSSPA EAIMAADRGV LSVRAKIKVR LTQLRPP VE IEAELFGHSG WQPGDAWMAE TTLGRVMFNE LLPLGYPFVN KQMHKKVQAA IINDLAERYP MIVVAQTVDK LKDAGFYW A TRSGVTVSMA DVLVPPRKKE ILDHYEERAD KVEKQFQRGA LNHDERNEAL VEIWKEATDE VGQALREHYP DDNPIITIV DSGATGNFTQ TRTLAGMKGL VTNPKGEFIP RPVKSSFREG LTVLEYFINT HGARKGLADT ALRTADSGYL TRRLVDVSQD VIVREHDCQ TERGIVVELA ERAPDGTLIR DPYIETSAYA RTLGTDAVDE AGNVIVERGQ DLGDPEIDAL LAAGITQVKV R SVLTCATS TGVCATCYGR SMATGKLVDI GEAVGIVAAQ SIGEPGTQLT MRTFHQGGVG EDITGGLPRV QELFEARVPR GK APIADVT GRVRLEDGER FYKITIVPDD GGEEVVYDKI SKRQRLRVFK HEDGSERVLS DGDHVEVGQQ LMEGSADPHE VLR VQGPRE VQIHLVREVQ EVYRAQGVSI HDKHIEVIVR QMLRRVTIID SGSTEFLPGS LIDRAEFEAE NRRVVAEGGE PAAG RPVLM GITKASLATD SWLSAASFQE TTRVLTDAAI NCRSDKLNGL KENVIIGKLI PAGTGINRYR NIAVQPTEEA RAAAY TIPS YEDQYYSPDF GAATGAAVPL DDYGYSDYRH HHHHH

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Macromolecule #4: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: ATCC 25618 / H37Rv
Molecular weightTheoretical: 11.85114 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MSISQSDASL AAVPAVDQFD PSSGASGGYD TPLGITNPPI DELLDRVSSK YALVIYAAKR ARQINDYYNQ LGEGILEYVG PLVEPGLQE KPLSIALREI HADLLEHTEG E

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Macromolecule #5: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 5 / Number of copies: 2 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #6: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 6 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration1.4 mg/mL
BufferpH: 7.9
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 7.0 µm / Nominal defocus min: 0.5 µm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 3064 / Average electron dose: 49.6 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 721752
Startup modelType of model: NONE / Details: ab-inito reconstuction
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 2
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.08 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 55008
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-7z8q:
Cryo-EM structure of Mycobacterium tuberculosis RNA polymerase core

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