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- EMDB-50509: cryo-EM structure of Mycobacterium tuberculosis sigma-B RNA polym... -

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Basic information

Entry
Database: EMDB / ID: EMD-50509
Titlecryo-EM structure of Mycobacterium tuberculosis sigma-B RNA polymerase bound to -10 promoter element ssDNA oligo - consensus map
Map dataprimary map sharpened
Sample
  • Complex: RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo
    • Protein or peptide: DNA-directed RNA polymerase subunit alpha
    • Protein or peptide: DNA-directed RNA polymerase subunit beta
    • Protein or peptide: DNA-directed RNA polymerase subunit beta'
    • Protein or peptide: DNA-directed RNA polymerase subunit omega
    • Protein or peptide: RNA polymerase sigma factor SigB
    • DNA: DNA (17-MER)
KeywordsRNA polymerase / sigma subunit / stress response / promoter element / tuberculosis / TRANSCRIPTION
Function / homology
Function and homology information


RNA polymerase core enzyme binding / Antimicrobial action and antimicrobial resistance in Mtb / sigma factor activity / DNA-directed RNA polymerase complex / peptidoglycan-based cell wall / DNA-templated transcription initiation / ribonucleoside binding / : / : / : ...RNA polymerase core enzyme binding / Antimicrobial action and antimicrobial resistance in Mtb / sigma factor activity / DNA-directed RNA polymerase complex / peptidoglycan-based cell wall / DNA-templated transcription initiation / ribonucleoside binding / : / : / : / : / : / : / DNA-directed RNA polymerase / response to heat / response to hypoxia / protein dimerization activity / response to xenobiotic stimulus / response to antibiotic / positive regulation of DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Sigma-70 factors family signature 1. / : / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 ...Sigma-70 factors family signature 1. / : / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 / RNA polymerase sigma-70 region 2 / RNA polymerase sigma-70 like domain / Sigma-70 region 2 / RNA polymerase sigma factor, region 2 / RNA polymerase sigma factor, region 3/4-like / 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 / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / 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 / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / 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 / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
RNA polymerase sigma factor SigB / 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) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.33 Å
AuthorsBrodolin K
Funding support France, 3 items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR-20-CE44-0020-01 France
Agence Nationale de la Recherche (ANR)ANR-16-CE11-0025 France
Instruct-ERIC Center (Strasbourg Centre)19348 France
CitationJournal: Nucleic Acids Res. / Year: 2025
Title: Single-stranded DNA drives sigma subunit loading onto mycobacterial RNA polymerase to unlock initiation-competent conformations
Authors: Vishwakarma RK / Marechal N / Morichaud Z / Blaise M / Margeat E / Brodolin K
History
DepositionMay 31, 2024-
Header (metadata) releaseApr 2, 2025-
Map releaseApr 2, 2025-
UpdateApr 16, 2025-
Current statusApr 16, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50509.png

Downloads & links

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Map

FileDownload / File: emd_50509.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationprimary map sharpened
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 360 pix.
= 310.32 Å		0.86 Å/pix.
x 360 pix.
= 310.32 Å		0.86 Å/pix.
x 360 pix.
= 310.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.862 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.14
Minimum - Maximum-0.813488 - 1.4035535
Average (Standard dev.)0.004259236 (±0.033726327)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 310.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Raw map

Fileemd_50509_additional_1.map
AnnotationRaw map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map A

Fileemd_50509_half_map_1.map
Annotationhalf-map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map B

Fileemd_50509_half_map_2.map
Annotationhalf-map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo

EntireName: RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo
Components
  • Complex: RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo
    • Protein or peptide: DNA-directed RNA polymerase subunit alpha
    • Protein or peptide: DNA-directed RNA polymerase subunit beta
    • Protein or peptide: DNA-directed RNA polymerase subunit beta'
    • Protein or peptide: DNA-directed RNA polymerase subunit omega
    • Protein or peptide: RNA polymerase sigma factor SigB
    • DNA: DNA (17-MER)

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Supramolecule #1: RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo

SupramoleculeName: RNA polymerase holoenzyme bound to -10 promoter element ssDNA oligo
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 406 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 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEED EPVTMYLRKQ GPGEVTAGDI VPPAGVTVHN PGMHIATLND KGKLEVELVV ERGRGYVPAV QNRASGAEIG RIPVDSIYSP ...String:
MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEED EPVTMYLRKQ GPGEVTAGDI VPPAGVTVHN PGMHIATLND KGKLEVELVV ERGRGYVPAV QNRASGAEIG RIPVDSIYSP VLKVTYKVDA TRVEQRTDFD KLILDVETKN SISPRDALAS AGKTLVELFG LARELNVEAE GIEIGPSPAE ADHIASFALP IDDLDLTVRS YNCLKREGVH TVGELVARTE SDLLDIRNFG QKSIDEVKIK LHQLGLSLKD SPPSFDPSEV AGYDVATGTW STEGAYDEQD YAETEQL

UniProtKB: DNA-directed RNA polymerase subunit alpha

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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 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYELS PIEDFSGSMS LSFSDPRFDD VKAPVDECKD KDMTYAAPLF VTAEFINNNT GEIKSQTVFM GDFPMMTEKG TFIINGTERV ...String:
MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYELS PIEDFSGSMS LSFSDPRFDD VKAPVDECKD KDMTYAAPLF VTAEFINNNT GEIKSQTVFM GDFPMMTEKG TFIINGTERV VVSQLVRSPG VYFDETIDKS TDKTLHSVKV IPSRGAWLEF DVDKRDTVGV RIDRKRRQPV TVLLKALGWT SEQIVERFGF SEIMRSTLEK DNTVGTDEAL LDIYRKLRPG EPPTKESAQT LLENLFFKEK RYDLARVGRY KVNKKLGLHV GEPITSSTLT EEDVVATIEY LVRLHEGQTT MTVPGGVEVP VETDDIDHFG NRRLRTVGEL IQNQIRVGMS RMERVVRERM TTQDVEAITP QTLINIRPVV AAIKEFFGTS QLSQFMDQNN PLSGLTHKRR LSALGPGGLS RERAGLEVRD VHPSHYGRMC PIETPEGPNI GLIGSLSVYA RVNPFGFIET PYRKVVDGVV SDEIVYLTAD EEDRHVVAQA NSPIDADGRF VEPRVLVRRK AGEVEYVPSS EVDYMDVSPR QMVSVATAMI PFLEHDDANR ALMGANMQRQ AVPLVRSEAP LVGTGMELRA AIDAGDVVVA EESGVIEEVS ADYITVMHDN GTRRTYRMRK FARSNHGTCA NQCPIVDAGD RVEAGQVIAD GPCTDDGEMA LGKNLLVAIM PWEGHNYEDA IILSNRLVEE DVLTSIHIEE HEIDARDTKL GAEEITRDIP NISDEVLADL DERGIVRIGA EVRDGDILVG KVTPKGETEL TPEERLLRAI FGEKAREVRD TSLKVPHGES GKVIGIRVFS REDEDELPAG VNELVRVYVA QKRKISDGDK LAGRHGNKGV IGKILPVEDM PFLADGTPVD IILNTHGVPR RMNIGQILET HLGWCAHSGW KVDAAKGVPD WAARLPDELL EAQPNAIVST PVFDGAQEAE LQGLLSCTLP NRDGDVLVDA DGKAMLFDGR SGEPFPYPVT VGYMYIMKLH HLVDDKIHAR STGPYSMITQ QPLGGKAQFG GQRFGEMECW AMQAYGAAYT LQELLTIKSD DTVGRVKVYE AIVKGENIPE PGIPESFKVL LKELQSLCLN VEVLSSDGAA IELREGEDED LERAAANLGI NLSRNESASV EDLA

UniProtKB: DNA-directed RNA polymerase subunit beta

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Macromolecule #4: DNA-directed RNA polymerase subunit beta'

MacromoleculeName: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: VNFFDELRIG LATAEDIRQW SYGEVKKPET INYRTLKPEK DGLFCEKIFG PTRDWECYCG KYKRVRFKGI ICERCGVEVT RAKVRRERMG HIELAAPVTH IWYFKGVPSR LGYLLDLAPK DLEKIIYFAA YVITSVDEEM RHNELSTLEA EMAVERKAVE DQRDGELEAR ...String:
VNFFDELRIG LATAEDIRQW SYGEVKKPET INYRTLKPEK DGLFCEKIFG PTRDWECYCG KYKRVRFKGI ICERCGVEVT RAKVRRERMG HIELAAPVTH IWYFKGVPSR LGYLLDLAPK DLEKIIYFAA YVITSVDEEM RHNELSTLEA EMAVERKAVE DQRDGELEAR AQKLEADLAE LEAEGAKADA RRKVRDGGER EMRQIRDRAQ RELDRLEDIW STFTKLAPKQ LIVDENLYRE LVDRYGEYFT GAMGAESIQK LIENFDIDAE AESLRDVIRN GKGQKKLRAL KRLKVVAAFQ QSGNSPMGMV LDAVPVIPPE LRPMVQLDGG RFATSDLNDL YRRVINRNNR LKRLIDLGAP EIIVNNEKRM LQESVDALFD NGRRGRPVTG PGNRPLKSLS DLLKGKQGRF RQNLLGKRVD YSGRSVIVVG PQLKLHQCGL PKLMALELFK PFVMKRLVDL NHAQNIKSAK RMVERQRPQV WDVLEEVIAE HPVLLNRAPT LHRLGIQAFE PMLVEGKAIQ LHPLVCEAFN ADFDGDQMAV HLPLSAEAQA EARILMLSSN NILSPASGRP LAMPRLDMVT GLYYLTTEVP GDTGEYQPAS GDHPETGVYS SPAEAIMAAD RGVLSVRAKI KVRLTQLRPP VEIEAELFGH SGWQPGDAWM AETTLGRVMF NELLPLGYPF VNKQMHKKVQ AAIINDLAER YPMIVVAQTV DKLKDAGFYW ATRSGVTVSM ADVLVPPRKK EILDHYEERA DKVEKQFQRG ALNHDERNEA LVEIWKEATD EVGQALREHY PDDNPIITIV DSGATGNFTQ TRTLAGMKGL VTNPKGEFIP RPVKSSFREG LTVLEYFINT HGARKGLADT ALRTADSGYL TRRLVDVSQD VIVREHDCQT ERGIVVELAE RAPDGTLIRD PYIETSAYAR TLGTDAVDEA GNVIVERGQD LGDPEIDALL AAGITQVKVR SVLTCATSTG VCATCYGRSM ATGKLVDIGE AVGIVAAQSI GEPGTQLTMR TFHQGGVGED ITGGLPRVQE LFEARVPRGK APIADVTGRV RLEDGERFYK ITIVPDDGGE EVVYDKISKR QRLRVFKHED GSERVLSDGD HVEVGQQLME GSADPHEVLR VQGPREVQIH LVREVQEVYR AQGVSIHDKH IEVIVRQMLR RVTIIDSGST EFLPGSLIDR AEFEAENRRV VAEGGEPAAG RPVLMGITKA SLATDSWLSA ASFQETTRVL TDAAINCRSD KLNGLKENVI IGKLIPAGTG INRYRNIAVQ PTEEARAAAY TIPSYEDQYY SPDFGAATGA AVPLDDYGYS DYRHHHHHH

UniProtKB: DNA-directed RNA polymerase subunit beta'

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Macromolecule #5: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MSISQSDASL AAVPAVDQFD PSSGASGGYD TPLGITNPPI DELLDRVSSK YALVIYAAKR ARQINDYYNQ LGEGILEYVG PLVEPGLQEK PLSIALREIH ADLLEHTEGE

UniProtKB: DNA-directed RNA polymerase subunit omega

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Macromolecule #6: RNA polymerase sigma factor SigB

MacromoleculeName: RNA polymerase sigma factor SigB / type: protein_or_peptide / ID: 6 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGSSHHHHHH SSGLVPRGSH MADAPTRATT SRVDSDLDAQ SPAADLVRVY LNGIGKTALL NAAGEVELAK RIEAGLYAEH LLETRKRLGE NRKRDLAAVV RDGEAARRHL LEANLRLVVS LAKRYTGRGM PLLDLIQEGN LGLIRAMEKF DYTKGFKFST YATWWIRQAI ...String:
MGSSHHHHHH SSGLVPRGSH MADAPTRATT SRVDSDLDAQ SPAADLVRVY LNGIGKTALL NAAGEVELAK RIEAGLYAEH LLETRKRLGE NRKRDLAAVV RDGEAARRHL LEANLRLVVS LAKRYTGRGM PLLDLIQEGN LGLIRAMEKF DYTKGFKFST YATWWIRQAI TRGMADQSRT IRLPVHLVEQ VNKLARIKRE MHQHLGREAT DEELAAESGI PIDKINDLLE HSRDPVSLDM PVGSEEEAPL GDFIEDAEAM SAENAVIAEL LHTDIRSVLA TLDEREHQVI RLRFGLDDGQ PRTLDQIGKL FGLSRERVRQ IERDVMSKLR HGERADRLRS YAS

UniProtKB: RNA polymerase sigma factor SigB

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Macromolecule #7: DNA (17-MER)

MacromoleculeName: DNA (17-MER) / type: dna / ID: 7 / Details: synthetic oligonucleotide / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
SequenceString:
TGCGTATAAT GTGTGGA

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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

Concentration6 mg/mL
BufferpH: 7.9
GridModel: Quantifoil R2/2 / Material: GOLD / Mesh: 200 / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 55.735 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE / Details: ab-initio
Final reconstructionNumber classes used: 1 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.33 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 167825
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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